TY  - JOUR
AB  - NPM1 is a multifunctional phosphoprotein with key roles in ribosome biogenesis amongst its many functions. NPM1 gene mutations drive 30% of acute myeloid leukemia (AML) cases. The mutations disrupt a nucleolar localization signal (NoLS) and create a novel nuclear export signal (NES), leading to cytoplasmic displacement of the protein (NPM1c). NPM1c mutations prime hematopoietic progenitors to leukemic transformation, but their precise molecular consequences remain elusive. Here, we first examine the effects of isolated NPM1c mutations on the global proteome of pre-leukemic hematopoietic stem and progenitor cells (HSPCs) using conditional knock-in Npm1cA/+ mice. We discover that many proteins involved in ribosome biogenesis are significantly depleted in these murine HSPCs, but also importantly in human NPM1-mutant AMLs. In line with this, we found that pre-leukemic Npm1cA/+ HSPCs display higher sensitivity to RNA polymerase I inhibitors, including Actinomycin D (ActD), compared to Npm1+/+ cells. Combination treatment with ActD and Venetoclax inhibited the growth and colony forming ability of pre-leukemic and leukemic NPM1c+ cells, whilst low-dose ActD treatment was able to re-sensitize resistant NPM1c+ cells to Venetoclax. Furthermore, using data from CRISPR dropout screens, we identified and validated TSR3, a 40S ribosomal maturation factor whose knock-out preferentially inhibited the proliferation of NPM1c+ AML cells by activating a p53-dependent apoptotic response. Similarly to low-dose ActD treatment, TSR3 depletion could partially restore sensitivity to Venetoclax in therapy-resistant NPM1c+ AML models. Our findings propose that targeted disruption of ribosome biogenesis should be explored as a therapeutic strategy against NPM1-mutant AML.
AU  - Damaskou, A.*
AU  - Wilson, R.*
AU  - Gozdecka, M.*
AU  - Giotopoulos, G.*
AU  - Asby, R.*
AU  - Eleftheriou, M.*
AU  - Gu, M.*
AU  - Récher, C.*
AU  - Mansat-De Mas, V.*
AU  - Vergez, F.*
AU  - Sahal, A.*
AU  - Vick, B.
AU  - Papachristou, E.K.*
AU  - Sawle, A.*
AU  - Yankova, E.*
AU  - Dudek, M.*
AU  - Liu, X.*
AU  - Russell, J.*
AU  - Rak, J.*
AU  - Hilcenko, C.*
AU  - D'Santos, C.S.*
AU  - Jeremias, I.
AU  - Sarry, J.E.*
AU  - Tzelepis, K.*
AU  - Huntly, B.J.*
AU  - Warren, A.J.*
AU  - Tavana, O.*
AU  - Vassiliou, G.S.*
C1  - 75014
C2  - 57697
CY  - Radarweg 29, 1043 Nx Amsterdam, Netherlands
SP  - 1239-1252
TI  - Posttranscriptional depletion of ribosome biogenesis factors engenders therapeutic vulnerabilities in NPM1-mutant AML.
JO  - Blood
VL  - 146
IS  - 10
PB  - Elsevier
PY  - 2025
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The success of targeted therapies for hematological malignancies has heralded their potential as both salvage treatment and early treatment lines, reducing the need for high-dose, intensive, and often toxic chemotherapeutic regimens. For young patients with classic Hodgkin lymphoma (cHL), immunotherapies provide the possibility to lessen long-term, treatment-related toxicities. However, suitable therapeutic targets are lacking. By integrating single-cell dissection of the tumor landscape and an in-depth, single-cell–based off-tumor antigen prediction, we identify CD86 as a promising therapeutic target in cHL. CD86 is highly expressed on Hodgkin and Reed-Sternberg cancer cells and cHL-specific tumor-associated macrophages. We reveal CD86–CTLA-4 as a key suppressive pathway in cHL, driving T-cell exhaustion. Cellular therapies targeting CD86 had extraordinary efficacy in vitro and in vivo and were safe in immunocompetent mouse models without compromising bacterial host defense in sepsis models. Our results prove the potential value of anti-CD86 immunotherapies for treating cHL.
AU  - Gottschlich, A.*
AU  - Grünmeier, R.*
AU  - Hoffmann, G.V.*
AU  - Nandi, S.*
AU  - Kavaka, V.*
AU  - Müller, P.J.*
AU  - Jobst, J.*
AU  - Oner, A.*
AU  - Kaiser, R.*
AU  - Gärtig, J.*
AU  - Piseddu, I.*
AU  - Frenz-Wiessner, S.*
AU  - Fairley, S.D.*
AU  - Schulz, H.*
AU  - Igl, V.*
AU  - Janert, T.A.*
AU  - Di Fina, L.*
AU  - Mulkers, M.*
AU  - Thomas, M.
AU  - Briukhovetska, D.*
AU  - Simnica, D.*
AU  - Carlini, E.*
AU  - Tsiverioti, C.A.*
AU  - Trefny, M.P.*
AU  - Lorenzini, T.*
AU  - Märkl, F.*
AU  - Mesquita, P.*
AU  - Brabenec, R.
AU  - Strzalkowski, T.*
AU  - Stock, S.*
AU  - Michaelides, S.*
AU  - Hellmuth, J.C.*
AU  - Thelen, M.*
AU  - Reinke, S.N.*
AU  - Klapper, W.*
AU  - Gelebart, P.F.*
AU  - Nicolai, L.*
AU  - Marr, C.
AU  - Beltrán, E.*
AU  - Megens, R.T.A.*
AU  - Klein, C.*
AU  - Baran-Marszak, F.*
AU  - Rosenwald, A.*
AU  - von Bergwelt-Baildon, M.*
AU  - Bröckelmann, P.J.*
AU  - Endres, S.
AU  - Kobold, S.
C1  - 73445
C2  - 56838
CY  - Radarweg 29, 1043 Nx Amsterdam, Netherlands
SP  - 1536-1552
TI  - Dissection of single-cell landscapes for the development of chimeric antigen receptor T cells in Hodgkin lymphoma.
JO  - Blood
VL  - 145
IS  - 14
PB  - Elsevier
PY  - 2025
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Telomere length shortening has been associated with genomic instability and acquisition of molecular lesions, but these processes have not been systematically studied across large cohorts of myeloid neoplasia (MN). As proof of concept for a novel, cross-validated WGS-based method of telomere content (TC) determination combined with mutations, transcriptomics, and functional assays, we studied TC in correlation with specific molecular features of a large cohort (n=1804) of MN patients including acute myeloid leukemia (AML) and myelodysplastic syndrome. When compared to healthy subjects and patients with non-clonal diseases such as persistent polyclonal B cell lymphocytosis, both MN and non-malignant controls with clonal disease, such as paroxysmal nocturnal hemoglobinuria and aplastic anemia, exhibited decreased TC. Furthermore, we show that TC is lowered in adult MN abrogating correlation with age with considerable TC diversification among certain morphologic and molecular subtypes. For instance, AML harbored the lowest TC. Furthermore, MN originating from a more mature cell of origin (e.g., APL), and those characterized by hyperproliferative driver mutations (e.g., RAS pathway genes) had lower TC, possibly indicating a loss of telomere maintenance capacity. In contrast, MN subtypes arising in a context of profound genetic alterations, such as TP53 mutations and complex karyotype, exhibited a relatively higher/preserved TC compared to other mutations. This phenomenon did not involve alternative lengthening processes but was rather consistent with an increased TC due to preserved activity of the telomerase complex. Our results describe a common and genotype-specific telomeric make-up of a large cohort of patients with MN providing a molecular benchmark for future therapeutic targeting of the telomere machinery.
AU  - Guarnera, L.*
AU  - Wahida, A.
AU  - Gurnari, C.*
AU  - Hutter, S.*
AU  - Stainczyk, S.A.*
AU  - Williams, N.*
AU  - Durmaz, A.*
AU  - Kubota, Y.*
AU  - Bravo-Perez, C.*
AU  - Kawashima, N.*
AU  - Orland, M.D.*
AU  - Pagliuca, S.*
AU  - Huang, Y.*
AU  - LaFramboise, T.*
AU  - Visconte, V.*
AU  - Walter, W.*
AU  - Meggendorfer, M.*
AU  - Kern, W.*
AU  - Westermann, F.*
AU  - Feuerbach, L.*
AU  - Haferlach, T.*
AU  - Maciejewski, J.P.*
C1  - 75606
C2  - 58052
TI  - Determining telomere content and genomics of myeloid neoplasia by whole-genome sequencing.
JO  - Blood
PY  - 2025
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Immunotherapy has become standard of care in the treatment of diffuse large B-cell lymphoma (DLBCL). Changes in immunophenotypes observed at first diagnosis predict therapy outcome but little is known about the resolution of these alterations in remission. Comprehensive characterization of immune changes from fresh, peripheral whole blood revealed a functionally relevant increase of myeloid-derived suppressor cells, reduced naïve T-cells, and an increase of activated and terminally differentiated T-cells before treatment which aggravated after therapy. Suggesting causal relation, injection of lymphoma in mice induced similar changes in the murine T cells. Distinct immune imprints were found in breast cancer and AML survivors. Identified alterations persisted beyond five years of ongoing complete remission and in DLBCL correlated with increased pro-inflammatory markers such as IL-6, B2M, or sCD14. The chronic inflammation was associated with functionally blunted T-cell immunity against SARS-CoV-2-specific peptides and reduced responses correlated with reduced Tn-cells. Persisting inflammation was confirmed by deep sequencing and by cytokine profiles, together pointing towards a compensatory activation of innate immunity. The persisting, lymphoma-induced immune alterations in remission may explain long-term complications, have implications for vaccine strategies, and are likely relevant for immunotherapies.
AU  - Pelzl, R.*
AU  - Benintende, G.*
AU  - Gsottberger, F.*
AU  - Scholz, J.K.*
AU  - Ruebner, M.*
AU  - Yao, H.*
AU  - Wendland, K.*
AU  - Rejeski, K.*
AU  - Altmann, H.*
AU  - Petkovic, S.*
AU  - Mellenthin, L.*
AU  - Kübel, S.*
AU  - Schmiedeberg, M.*
AU  - Klein, P.*
AU  - Petrera, A.
AU  - Baur, R.*
AU  - Eckstein, S.*
AU  - Hoepffner-Grundy, S.*
AU  - Röllig, C.*
AU  - Subklewe, M.*
AU  - Huebner, H.*
AU  - Schett, G.*
AU  - Mackensen, A.*
AU  - Laurenti, L.*
AU  - Graw, F.*
AU  - Völkl, S.*
AU  - Nganou-Makamdop, K.*
AU  - Müller, F.*
C1  - 74551
C2  - 57506
CY  - Radarweg 29, 1043 Nx Amsterdam, Netherlands
SP  - 1300-1313
TI  - Large B-cell lymphoma imprints a dysfunctional immune phenotype that persists years after treatment.
JO  - Blood
VL  - 146
IS  - 11
PB  - Elsevier
PY  - 2025
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - In this issue of Blood, Sam et al demonstrate the translational potential of CD19-targeted CD28-mediated costimulation to ameliorate the therapeutic potential of the anti-CD19-anti-CD3 T-cell bispecific engager glofitamab in preclinical models, buttressing the rationale for an ongoing phase 1 clinicalstudy.(1)
AU  - Kobold, S.
C1  - 70714
C2  - 55560
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 2115-2116
TI  - Giving T-cell bispecifics a helping hand.
JO  - Blood
VL  - 143
IS  - 21
PB  - Amer Soc Hematology
PY  - 2024
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - RNA-binding proteins (RBPs) form a large and diverse class of factors, many members of which are overexpressed in hematologic malignancies. RBPs participate in various processes of messenger RNA (mRNA) metabolism and prevent harmful DNA:RNA hybrids or R-loops. Here, we report that PIWIL4, a germ stem cell-associated RBP belonging to the RNase H-like superfamily, is overexpressed in patients with acute myeloid leukemia (AML) and is essential for leukemic stem cell function and AML growth, but dispensable for healthy human hematopoietic stem cells. In AML cells, PIWIL4 binds to a small number of known piwi-interacting RNA. Instead, it largely interacts with mRNA annotated to protein-coding genic regions and enhancers that are enriched for genes associated with cancer and human myeloid progenitor gene signatures. PIWIL4 depletion in AML cells downregulates the human myeloid progenitor signature and leukemia stem cell (LSC)-associated genes and upregulates DNA damage signaling. We demonstrate that PIWIL4 is an R-loop resolving enzyme that prevents R-loop accumulation on a subset of AML and LSC-associated genes and maintains their expression. It also prevents DNA damage, replication stress, and activation of the ATR pathway in AML cells. PIWIL4 depletion potentiates sensitivity to pharmacological inhibition of the ATR pathway and creates a pharmacologically actionable dependency in AML cells.
AU  - Bamezai, S.*
AU  - Pulikkottil, A.J.*
AU  - Yadav, T.*
AU  - Vegi, N.M.*
AU  - Mueller, J.*
AU  - Mark, J.*
AU  - Mandal, T.*
AU  - Feder, K.*
AU  - Ihme, S.*
AU  - Song, C.*
AU  - Rosler, R.*
AU  - Wiese, S.*
AU  - Hoell, J.I.*
AU  - Kloetgen, A.*
AU  - Karsan, A.*
AU  - Kumari, A.*
AU  - Wojenski, L.*
AU  - Sinha, A.U.*
AU  - González-Menéndez, I.*
AU  - Quintanilla-Martinez, L.*
AU  - Donato, E.*
AU  - Trumpp, A.*
AU  - Kruse, E.
AU  - Hamperl, S.
AU  - Zou, L.*
AU  - Rawat, V.P.S.*
AU  - Buske, C.*
C1  - 67770
C2  - 54248
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 90-105
TI  - A noncanonical enzymatic function of PIWIL4 maintains genomic integrity and leukemic growth in AML.
JO  - Blood
VL  - 142
IS  - 1
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Aberrant expression of stem-cell-associated genes is a common feature in acute myeloid leukemia (AML) and is linked to leukemic self-renewal and therapy resistance. Using AF10-rearranged leukemia as a prototypical example of the recurrently activated "stemness" network in AML, we screened for chromatin regulators that sustain its expression. We deployed a CRISPR-Cas9 screen with a bespoke domain-focused library and identified several novel chromatin-modifying complexes as regulators of the TALE domain transcription factor MEIS1, a key leukemia stem cell (LSC)-associated gene. CRISPR droplet sequencing revealed that many of these MEIS1 regulators coordinately controlled the transcription of several AML oncogenes. In particular, we identified a novel role for the Tudor-domain containing chromatin reader protein SGF29 in the transcription of AML oncogenes. Furthermore, SGF29 deletion impaired leukemogenesis in models representative of multiple AML subtypes in multiple AML subtype models. Our studies reveal a novel role for SGF29 as a non-oncogenic dependency in AML and identify the SGF29 Tudor domain as an attractive target for drug discovery.
AU  - Barbosa, K.O.*
AU  - Deshpande, A.*
AU  - Perales, M.E.*
AU  - Xiang, P.*
AU  - Murad, R.*
AU  - Pramod, A.B.*
AU  - Minkina, A.*
AU  - Robertson, N.A.*
AU  - Schischlik, F.*
AU  - Lei, X.*
AU  - Sun, Y.*
AU  - Brown, A.*
AU  - Amend, D.
AU  - Jeremias, I.
AU  - Doench, J.G.*
AU  - Humphries, K.K.*
AU  - Ruppin, E.*
AU  - Shendure, J.A.*
AU  - Mali, P.*
AU  - Adams, P.D.*
AU  - Deshpande, A.J.*
C1  - 68873
C2  - 53732
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 697-712
TI  - Transcriptional control of leukemogenesis by the chromatin reader SGF29.
JO  - Blood
VL  - 143
IS  - 8
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Frenz-Wiessner, S.*
AU  - Fairley, S.*
AU  - Buser, M.
AU  - Goek, I.*
AU  - Salewskij, K.*
AU  - Jonsson, G.*
AU  - zu Putlitz, B.*
AU  - Petersheim, D.*
AU  - Illig, D.*
AU  - Li, Y.*
AU  - Chen, P.*
AU  - Kalauz, M.*
AU  - Conca, R.*
AU  - Sterr, M.
AU  - Geuder, J.*
AU  - Mizoguchi, Y.*
AU  - Kotlarz, D.*
AU  - Rudelius, M.*
AU  - Penninger, J.M.*
AU  - Marr, C.
AU  - Klein, C.*
C1  - 70127
C2  - 55095
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Human Induced Pluripotent Stem Cell-Derived Bone Marrow Organoids to Model Hematopoietic Development.
JO  - Blood
VL  - 142
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Ghalandary
AU  - Gao, Y.
AU  - Amend, D.
AU  - Kutkaite, G.
AU  - Vick, B.
AU  - Spiekermann, K.*
AU  - Rothenberg-Thurley, M.*
AU  - Metzeler, K.H.*
AU  - Marcinek, A.*
AU  - Subklewe, M.*
AU  - Menden, M.P.
AU  - Jurinovic, V.
AU  - Bahrami, E.
AU  - Jeremias, I.
C1  - 66481
C2  - 52826
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 955-960
TI  - WT1 and DNMT3A play essential roles in the growth of certain patient AML cells in mice.
JO  - Blood
VL  - 141
IS  - 8
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The mechanisms of coordinated changes in proteome composition and their relevance for the differentiation of neutrophil granulocytes are not well studied. Here, we discover two novel human genetic defects in SRPRA and SRP19, constituents of the mammalian co-translational targeting machinery and characterize their role in neutrophil granulocyte differentiation. We systematically study the proteome of neutrophil granulocytes from patients with variants in the signal recognition particle (SRP) genes, HAX1, and ELANE and identify global as well as specific proteome aberrations. Using in vitro differentiation of human induced pluripotent stem cells and in vivo zebrafish models, we study the effects of SRP-deficiency on neutrophil granulocyte development. In a heterologous cell-based inducible protein expression system, we validate the effects conferred by SRP dysfunction for selected proteins that we identified in our proteome screen. Thus, SRP-dependent protein processing, intracellular trafficking and homeostasis are critically important for the differentiation of neutrophil granulocytes.
AU  - Linder, M.I.*
AU  - Mizoguchi, Y.*
AU  - Hesse, S.*
AU  - Csaba, G.*
AU  - Tatematsu, M.*
AU  - Łyszkiewicz, M.*
AU  - Ziętara, N.*
AU  - Jeske, T.*
AU  - Hastreiter, M.*
AU  - Rohlfs, M.*
AU  - Liu, Y.*
AU  - Grabowski, P.*
AU  - Ahomaa, K.
AU  - Maier-Begandt, D.*
AU  - Schwestka, M.*
AU  - Pazhakh, V.*
AU  - Isiaku, A.*
AU  - Briones Miranda, B.*
AU  - Blombery, P.*
AU  - Saito, M.K.*
AU  - Rusha, E.*
AU  - Alizadeh, Z.*
AU  - Pourpak, Z.*
AU  - Kobayashi, M.*
AU  - Rezaei, N.*
AU  - Unal, E.*
AU  - Hauck, F.*
AU  - Drukker, M.
AU  - Walzog, B.*
AU  - Rappsilber, J.*
AU  - Zimmer, R.*
AU  - Lieschke, G.J.*
AU  - Klein, C.*
C1  - 66497
C2  - 52825
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 645-658
TI  - Human genetic defects in SRP19 and SRPRA cause severe congenital neutropenia with distinctive proteome changes.
JO  - Blood
VL  - 141
IS  - 6
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Fms like tyrosine kinase 3 (FLT3) is often overexpressed or constitutively activated by internal tandem duplication (ITD) and tyrosine kinase domain (TKD) mutations in acute myeloid leukemia (AML). Despite the use of receptor tyrosine kinase inhibitors (TKI) in FLT3-ITD positive AML, the prognosis of patients is still poor and further improvement of therapy is required. Targeting FLT3 independent of mutations by antibody‑drug‑conjugates (ADCs) is a promising strategy for AML therapy. Here, we report the development and preclinical characterization of a novel FLT3‑targeting ADC, 20D9-ADC, which was generated by applying the innovative P5 conjugation technology. In vitro, 20D9‑ADC mediated potent cytotoxicity to Ba/F3 cells expressing transgenic FLT3 or FLT3-ITD, to AML cell lines and to FLT3-ITD positive patient derived xenograft AML cells. In vivo, 20D9‑ADC treatment led to a significant tumor reduction and even durable complete remission in AML xenograft models. Further, 20D9‑ADC demonstrated no severe hematotoxicity in in vitro colony formation assays using concentrations that were cytotoxic in AML cell line treatment. The combination of 20D9-ADC with the TKI midostaurin showed strong synergy in vitro and in vivo, leading to reduction of aggressive AML cells below the detection limit. Our data indicate that targeting FLT3 with an advanced new-generation ADC is a promising and potent antileukemic strategy, especially when combined with FLT3-TKI in FLT3‑ITD positive AML.
AU  - Roas, M.*
AU  - Vick, B.
AU  - Kasper, M.A.*
AU  - Able, M.*
AU  - Polzer, H.*
AU  - Gerlach, M.*
AU  - Kremmer, E.
AU  - Hecker, J.S.*
AU  - Schmitt, S.*
AU  - Stengl, A.*
AU  - Waller, V.*
AU  - Hohmann, N.*
AU  - Festini, M.*
AU  - Ludwig, A.E.*
AU  - Rohrbacher, L.*
AU  - Herold, T.*
AU  - Subklewe, M.*
AU  - Götze, K.S.*
AU  - Hackenberger, C.P.R.*
AU  - Schumacher, D.*
AU  - Helma-Smets, J.*
AU  - Jeremias, I.
AU  - Leonhardt, H.*
AU  - Spiekermann, K.*
C1  - 66008
C2  - 53051
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1023-1035
TI  - Targeting FLT3 by new-generation antibody-drug-conjugate in combination with kinase inhibitors for treatment of AML.
JO  - Blood
VL  - 141
IS  - 9
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Patients with hypomorphic mutations in RAG1 or RAG2 genes present as either Omenn syndrome or atypical combined immunodeficiency with a wide phenotypic range. Hematopoietic stem cell transplantation (HSCT) is potentially curative, but data are scarce. We report on a worldwide cohort of 60 patients with hypomorphic RAG variants who underwent HSCT, 78% of whom experienced infections (29% active at HSCT), 72% had autoimmunity, and 18% had granulomas pretransplant. These complications are frequently associated with organ damage. Eight individuals (13%) were diagnosed by newborn screening or family history. HSCT was performed at a median of 3.4 years (range 0.3-42.9 years) from matched unrelated donors, matched sibling or matched family donors, or mismatched donors in 48%, 22%, and 30% of the patients, respectively. Grafts were T-cell depleted in 15 cases (25%). Overall survival at 1 and 4 years was 77.5% and 67.5% (median follow-up of 39 months). Infection was the main cause of death. In univariable analysis, active infection, organ damage pre-HSCT, T-cell depletion of the graft, and transplant from a mismatched family donor were predictive of worse outcome, whereas organ damage and T-cell depletion remained significant in multivariable analysis (hazard ratio [HR] = 6.01, HR = 8.46, respectively). All patients diagnosed by newborn screening or family history survived. Cumulative incidences of acute and chronic graft-versus-host disease were 35% and 22%, respectively. Cumulative incidences of new-onset autoimmunity was 15%. Immune reconstitution, particularly recovery of naïveCD4+ T cells, was faster and more robust in patients transplanted before 3.5 years of age, and without organ damage. These findings support the indication for early transplantation.
AU  - Schuetz, C.*
AU  - Gerke, J.S.*
AU  - Ege, M.J.
AU  - Walter, J.*
AU  - Kusters, M.*
AU  - Worth, A.*
AU  - Kanakry, J.A.*
AU  - Dimitrova, D.*
AU  - Wolska-Kuśnierz, B.*
AU  - Chen, K.*
AU  - Unal, E.*
AU  - Karakukcu, M.*
AU  - Pashchenko, O.*
AU  - Leiding, J.*
AU  - Kawai, T.*
AU  - Amrolia, P.J.*
AU  - Berghuis, D.*
AU  - Buechner, J.*
AU  - Buchbinder, D.*
AU  - Cowan, M.J.*
AU  - Gennery, A.R.*
AU  - Güngör, T.*
AU  - Heimall, J.*
AU  - Miano, M.*
AU  - Meyts, I.*
AU  - Morris, E.C.*
AU  - Riviere, J.*
AU  - Sharapova, S.O.*
AU  - Shaw, P.J.*
AU  - Slatter, M.*
AU  - Hönig, M.*
AU  - Veys, P.*
AU  - Fischer, A.*
AU  - Cavazzana, M.*
AU  - Moshous, D.*
AU  - Schulz, A.*
AU  - Albert, M.H.*
AU  - Puck, J.M.*
AU  - Lankester, A.C.*
AU  - Notarangelo, L.D.*
AU  - Neven, B.*
C1  - 67074
C2  - 53448
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 713-724
TI  - Hypomorphic RAG deficiency: Impact of disease burden on survival and thymic recovery argues for early diagnosis and HSCT.
JO  - Blood
VL  - 141
IS  - 7
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Constitutive MALT1 activity drives survival of malignant lymphomas addicted to chronic B-cell receptor (BCR) signaling, oncogenic CARD11, or the API2-MALT1 (also BIRC3::MALT1) fusion oncoprotein. While MALT1 scaffolding induces NF-kB-dependent survival signaling, MALT1 protease function is thought to augment NF-kB activation by cleaving signaling mediators and transcriptional regulators in B-cell lymphomas. However, the pathological role of MALT1 protease function in lymphomagenesis is not well understood. Here, we show that TRAF6 controls MALT1-dependent activation of NF-kB transcriptional responses, but is dispensable for MALT1 protease activation driven by oncogenic CARD11. To uncouple enzymatic and non-enzymatic functions of MALT1, we analyzed TRAF6-dependent and -independent as well as MALT1 protease-dependent gene expression profiles downstream of oncogenic CARD11 and API2-MALT1. The data hint that by cleaving and inactivating the RNA binding proteins Regnase-1 and Roquin-1/2, MALT1 protease induces post-transcriptional upregulation of many genes including NFKBIZ/IkBz, NFKBID/IkBNS and ZC3H12A/Regnase-1 in activated B-cell-like diffuse large B-cell lymphoma (ABC DLBCL). We demonstrate that oncogene-driven MALT1 activity in ABC DLBCL cells regulates NFKBIZ and NFKBID induction on mRNA level via releasing a brake imposed by Regnase-1 and Roquin-1/2. Furthermore, MALT1 protease drives post-transcriptional gene induction in the context of the API2-MALT1 fusion created by the recurrent t(11;18)(q21;q21) translocation in mucosa-associated lymphoid tissue (MALT) lymphoma. Thus, MALT1 paracaspase acts as a bifurcation point for enhancing transcriptional and post-transcriptional gene expression in malignant lymphomas. Moreover, the identification of MALT1 protease selective target genes provides specific biomarkers for the clinical evaluation of MALT1 inhibitors.
AU  - Wimberger, N.
AU  - Ober, F.
AU  - Avar, G.
AU  - Grau, M.*
AU  - Xu, W.*
AU  - Lenz, G.*
AU  - Menden, M.P.
AU  - Krappmann, D.
C1  - 67954
C2  - 54432
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1985-2001
TI  - Oncogene-induced MALT1 protease activity drives post-transcriptional gene expression in malignant lymphomas.
JO  - Blood
VL  - 142
IS  - 23
PB  - Amer Soc Hematology
PY  - 2023
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - BCL-2 inhibition has been shown to be effective in acute myeloid leukemia (AML) in combination with hypomethylating agents or low-dose cytarabine. However, resistance and relapse represent major clinical challenges. Therefore, there is an unmet need to overcome resistance to current venetoclax-based strategies. We performed high-throughput drug screening to identify effective combination partners for venetoclax in AML. Overall, 64 antileukemic drugs were screened in 31 primary high-risk AML samples with or without venetoclax. Gilteritinib exhibited the highest synergy with venetoclax in FLT3 wild-type AML. The combination of gilteritinib and venetoclax increased apoptosis, reduced viability, and was active in venetoclax-azacitidine–resistant cell lines and primary patient samples. Proteomics revealed increased FLT3 wild-type signaling in specimens with low in vitro response to the currently used venetoclax-azacitidine combination. Mechanistically, venetoclax with gilteritinib decreased phosphorylation of ERK and GSK3B via combined AXL and FLT3 inhibition with subsequent suppression of the antiapoptotic protein MCL-1. MCL-1 downregulation was associated with increased MCL-1 phosphorylation of serine 159, decreased phosphorylation of threonine 161, and proteasomal degradation. Gilteritinib and venetoclax were active in an FLT3 wild-type AML patient-derived xenograft model with TP53 mutation and reduced leukemic burden in 4 patients with FLT3 wild-type AML receiving venetoclax-gilteritinib off label after developing refractory disease under venetoclax-azacitidine. In summary, our results suggest that combined inhibition of FLT3/AXL potentiates venetoclax response in FLT3 wild-type AML by inducing MCL-1 degradation. Therefore, the venetoclax-gilteritinib combination merits testing as a potentially active regimen in patients with high-risk FLT3 wild-type AML.
AU  - Janssen, M.*
AU  - Schmidt, C.*
AU  - Bruch, P.M.*
AU  - Blank, M.F.*
AU  - Rohde, C.*
AU  - Waclawiczek, A.*
AU  - Heid, D.*
AU  - Renders, S.*
AU  - Göllner, S.*
AU  - Vierbaum, L.*
AU  - Besenbeck, B.*
AU  - Herbst, S.A.*
AU  - Knoll, M.*
AU  - Kolb, C.*
AU  - Przybylla, A.*
AU  - Weidenauer, K.*
AU  - Ludwig, A.K.*
AU  - Fabre, M.*
AU  - Gu, M.*
AU  - Schlenk, R.F.*
AU  - Stölzel, F.*
AU  - Bornhäuser, M.*
AU  - Röllig, C.*
AU  - Platzbecker, U.*
AU  - Baldus, C.*
AU  - Serve, H.*
AU  - Sauer, T.*
AU  - Raffel, S.*
AU  - Pabst, C.*
AU  - Vassiliou, G.S.*
AU  - Vick, B.
AU  - Jeremias, I.
AU  - Trumpp, A.*
AU  - Krijgsveld, J.*
AU  - Müller-Tidow, C.*
AU  - Dietrich, S.*
C1  - 66901
C2  - 53350
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 2594–2610
TI  - Venetoclax synergizes with gilteritinib in FLT3 wild-type high-risk acute myeloid leukemia by suppressing MCL-1.
JO  - Blood
VL  - 140
IS  - 24
PB  - Amer Soc Hematology
PY  - 2022
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - T-cell–recruiting bispecific molecule therapy has yielded promising results in patients with hematologic malignancies; however, resistance and subsequent relapse remains a major challenge. T-cell exhaustion induced by persistent antigen stimulation or tonic receptor signaling has been reported to compromise outcomes of T-cell–based immunotherapies. The impact of continuous exposure to bispecifics on T-cell function, however, remains poorly understood. In relapsed/refractory B-cell precursor acute lymphoblastic leukemia patients, 28-day continuous infusion with the CD19xCD3 bispecific molecule blinatumomab led to declining T-cell function. In an in vitro model system, mimicking 28-day continuous infusion with the half-life–extended CD19xCD3 bispecific AMG 562, we identified hallmark features of exhaustion arising over time. Continuous AMG 562 exposure induced progressive loss of T-cell function (day 7 vs day 28 mean specific lysis: 88.4% vs 8.6%; n = 6; P = .0003). Treatment-free intervals (TFIs), achieved by AMG 562 withdrawal, were identified as a powerful strategy for counteracting exhaustion. TFIs induced strong functional reinvigoration of T cells (continuous vs TFI-specific lysis on day 14: 34.9% vs 93.4%; n = 6; P < .0001) and transcriptional reprogramming. Furthermore, use of a TFI led to improved T-cell expansion and tumor control in vivo. Our data demonstrate the relevance of T-cell exhaustion in bispecific antibody therapy and highlight that T cells can be functionally and transcriptionally rejuvenated with TFIs. In view of the growing number of bispecific molecules being evaluated in clinical trials, our findings emphasize the need to consider and evaluate TFIs in application schedules to improve clinical outcomes.
AU  - Philipp, N.*
AU  - Kazerani, M.*
AU  - Nicholls, A.*
AU  - Vick, B.
AU  - Wulf, J.*
AU  - Straub, T.*
AU  - Scheurer, M.*
AU  - Muth, A.*
AU  - Hänel, G.*
AU  - Nixdorf, D.*
AU  - Sponheimer, M.*
AU  - Ohlmeyer, M.*
AU  - Lacher, S.M.*
AU  - Brauchle, B.*
AU  - Marcinek, A.*
AU  - Rohrbacher, L.*
AU  - Leutbecher, A.*
AU  - Rejeski, K.*
AU  - Weigert, O.*
AU  - von Bergwelt-Baildon, M.*
AU  - Theurich, S.*
AU  - Kischel, R.*
AU  - Jeremias, I.
AU  - Bücklein, V.*
AU  - Subklewe, M.*
C1  - 66186
C2  - 52622
SP  - 1104-1118
TI  - T-cell exhaustion induced by continuous bispecific molecule exposure is ameliorated by treatment-free intervals.
JO  - Blood
VL  - 140
IS  - 10
PY  - 2022
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Antibody-based immunotherapy is a promising strategy for targeting chemo-resistant leukemic cells. However, classical antibody-based approaches are restricted to targeting lineage-specific cell-surface antigens. By targeting intracellular antigens, a large number of other leukemia-associated targets would become accessible. In this study, we evaluated a novel T-cell bispecific (TCB) antibody, generated using CrossMab and knob-into-holes technology, containing a bivalent T-cell receptor-like binding domain that recognizes the RMFPNAPYL peptide derived from the intracellular tumor antigen Wilms' tumor 1 (WT1) in the context of human leukocyte antigen (HLA) A*02. Binding to CD3ε recruits T cells irrespective of their T-cell receptor specificity. WT1-TCB elicited antibody-mediated T-cell cytotoxicity against AML cell lines in a WT1- and HLA-restricted manner. Specific lysis of primary AML cells was mediated in ex vivo long-term co-cultures utilizing allogenic (mean specific lysis: 67±6% after 13-14 days; ±SEM; n=18) or autologous, patient-derived T cells (mean specific lysis: 54±12% after 11-14 days; ±SEM; n=8). WT1-TCB-treated T cells exhibited higher cytotoxicity against primary AML cells than an HLA-A*02 RMF-specific T-cell clone. Combining WT1-TCB with the immunomodulatory drug lenalidomide further enhanced antibody-mediated T-cell cytotoxicity against primary AML cells (mean specific lysis on day 3-4: 45.4±9.0% vs 70.8±8.3%; p=0.015; ±SEM; n=9-10). In vivo, WT1-TCB-treated humanized mice bearing SKM-1 tumors showed a significant and dose-dependent reduction in tumor growth. In summary, we show that WT1-TCB facilitates potent in vitro, ex vivo and in vivo killing of AML cell lines and primary AML cells; these results led to the initiation of a phase I trial in patients with r/r AML (NCT04580121).
AU  - Augsberger, C.*
AU  - Hänel, G.*
AU  - Xu, W.*
AU  - Pulko, V.*
AU  - Hanisch, L.J.*
AU  - Augustin, A.*
AU  - Challier, J.*
AU  - Hunt, K.
AU  - Vick, B.
AU  - Rovatti, P.E.*
AU  - Krupka, C.*
AU  - Rothe, M.*
AU  - Schönle, A.*
AU  - Sam, J.*
AU  - Lezan, E.*
AU  - Ducret, A.*
AU  - Ortiz-Franyuti, D.*
AU  - Walz, A.C.*
AU  - Benz, J.*
AU  - Bujotzek, A.*
AU  - Lichtenegger, F.S.*
AU  - Gassner, C.*
AU  - Carpy, A.*
AU  - Lyamichev, V.*
AU  - Patel, J.*
AU  - Konstandin, N.P.*
AU  - Tunger, A.*
AU  - Schmitz, M.*
AU  - von Bergwelt-Baildon, M.*
AU  - Spiekermann, K.*
AU  - Vago, L.*
AU  - Jeremias, I.
AU  - Marrer-Berger, E.*
AU  - Umaña, P.*
AU  - Klein, C.*
AU  - Subklewe, M.*
C1  - 62554
C2  - 50939
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 2655-2669
TI  - Targeting intracellular WT1 in AML with a novel RMF-peptide-MHC specific T-cell bispecific antibody.
JO  - Blood
VL  - 138
IS  - 25
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Leukemias bearing fusions of the AF10/MLLT10 gene are associated with poor prognosis, and therapies targeting these fusion proteins are lacking. To understand mechanisms underlying AF10 fusion-mediated leukemogenesis, we generated inducible mouse models of AML driven by the most common AF10 fusion proteins, PICALM/CALM-AF10 and KMT2A/MLL-AF10, and performed comprehensive characterization of the disease using transcriptomic, epigenomic, proteomic, and functional genomic approaches. Our studies provide a detailed map of gene networks and protein interactors associated with key AF10 fusions involved in leukemia. Specifically, we report that AF10 fusions activate a cascade of JAK/STAT-mediated inflammatory signaling through direct recruitment of JAK1 kinase. Inhibition of the JAK/STAT signaling by genetic Jak1 deletion or through pharmacological JAK/STAT inhibition elicited potent anti-oncogenic effects in mouse and human models of AF10 fusion AML. Collectively, our study identifies JAK1 as a tractable therapeutic target in AF10-rearranged leukemias.
AU  - Chen, B.R.*
AU  - Deshpande, A.*
AU  - Barbosa, K.O.*
AU  - Kleppe, M.*
AU  - Lei, X.*
AU  - Yeddula, N.*
AU  - Sánchez Vela, P.*
AU  - Campos, A.R.*
AU  - Wechsler-Reya, R.J.*
AU  - Bagchi, A.*
AU  - Meshinchi, S.*
AU  - Eaves, C.J.*
AU  - Jeremias, I.
AU  - Haferlach, T.*
AU  - Frank, D.A.*
AU  - Ronai, Z.A.*
AU  - Chanda, S.*
AU  - Armstrong, S.A.*
AU  - Adams, P.*
AU  - Levine, R.L.*
C1  - 61557
C2  - 50346
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 3403-3415
TI  - A JAK/STAT-mediated inflammatory signaling cascade drives oncogenesis In AF10-rearranged AML.
JO  - Blood
VL  - 137
IS  - 24
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The expression of ZAP-70 in a subset of CLL patients strongly correlates with a more aggressive clinical course, though the exact underlying mechanisms remain elusive. The ability of ZAP-70 to enhance B cell receptor (BCR) signaling, independently of its kinase function, is considered to contribute. Here we employed RNA-sequencing and proteomic analyses of primary cells differing only in their expression of ZAP-70 to further define how ZAP-70 increases aggressiveness of CLL. We identified that ZAP-70 is directly required for cell survival in the absence of an overt BCR signal, which can compensate for ZAP-70 deficiency as an anti-apoptotic signal. In addition, the expression of ZAP-70 regulates the transcription of factors regulating recruitment and activation of T cells, such as CCL3, CCL4 and IL4I1. Quantitative mass spectrometry of double-cross linked ZAP-70 complexes further demonstrated constitutive and direct protein-protein interactions between ZAP-70 and BCR-signaling components. Unexpectedly, ZAP-70 also binds to ribosomal proteins, which is not dependent on, but further increased by BCR-stimulation. Importantly, decreased expression of ZAP-70 significantly reduced MYC-expression and global protein synthesis, providing evidence that ZAP-70 contributes to translational dysregulation in CLL. In conclusion, ZAP-70 constitutively promotes cell survival, microenvironment-interactions and protein synthesis in CLL cells, likely to improve cellular fitness and to further drive disease progression.
AU  - Chen, J.*
AU  - Sathiaseelan, V.*
AU  - Moore, A.D.*
AU  - Tan, S.*
AU  - Chilamakuri, C.S.*
AU  - Roamio Franklin, V.N.*
AU  - Shahsavari, A.*
AU  - Jakwerth, C.A.
AU  - Hake, S.B.*
AU  - Warren, A.J.*
AU  - Mohorianu, I.*
AU  - D'Santos, C.S.*
AU  - Ringshausen, I.*
C1  - 61488
C2  - 50289
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 3629-3640
TI  - ZAP-70 constitutively regulates gene expression and protein synthesis in chronic lymphocytic leukemia.
JO  - Blood
VL  - 137
IS  - 26
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Primary immunodeficiencies in the co-stimulatory molecule CD27 and its ligand CD70 predispose for pathologies of uncontrolled Epstein Barr virus (EBV) infection in nearly all affected patients. We demonstrate that both depletion of CD27 positive cells and antibody blocking of CD27 interaction with CD70 causes uncontrolled EBV infection in mice with reconstituted human immune system components. While overall CD8+ T cell expansion and composition is unaltered after antibody blocking of CD27, only some EBV specific CD8+ T cell responses, exemplified by early lytic EBV antigen BMLF1 specific CD8+ T cells are inhibited in their proliferation and killing of EBV transformed B cells. This suggests that CD27 is not required for all CD8+ T cell expansions and cytotoxicity, but for a subset of CD8+ T cell responses that protect us from EBV infection.
AU  - Deng, Y.*
AU  - Chatterjee, B.*
AU  - Zens, K.*
AU  - Zdimerova, H.*
AU  - Müller, A.*
AU  - Schuhmachers, P.*
AU  - Ligeon, L.A.*
AU  - Bongiovanni, A.*
AU  - Capaul, R.*
AU  - Zbinden, A.*
AU  - Holler, A.*
AU  - Stauss, H.J.*
AU  - Hammerschmidt, W.
AU  - Münz, C.*
C1  - 61793
C2  - 50454
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 3225-3236
TI  - CD27 is required for protective lytic EBV antigen specific CD8+ T cell expansion.
JO  - Blood
VL  - 137
IS  - 23
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Vacuolar protein sorting 45 homolog (VPS45), a member of the Sec1/Munc18 (SM) family, has been implicated in the regulation of endosomal trafficking. VPS45 deficiency in human patients results in congenital neutropenia, bone marrow fibrosis, and extramedullary renal hematopoiesis. Detailed mechanisms of the VPS45 function are unknown. Here, we show an essential role of mammalian VPS45 in maintaining the intracellular organization of endolysosomal vesicles and promoting recycling of cell-surface receptors. Loss of VPS45 causes defective Rab5-to-Rab7 conversion resulting in trapping of cargos in early endosomes and impaired delivery to lysosomes. In this context, we demonstrate aberrant trafficking of the G-CSF receptor (G-CSFR) in the absence of VPS45. Furthermore, we find that lack of VPS45 in mice is not compatible with embryonic development. Thus, we identify mammalian VPS45 as a critical regulator of trafficking through the endosomal system and early embryogenesis of mice.
AU  - Frey, L.*
AU  - Ziętara, N.*
AU  - Lyszkiewicz, M.*
AU  - Marquardt, B.*
AU  - Mizoguchi, Y.*
AU  - Linder, M.I.*
AU  - Liu, Y.*
AU  - Giesert, F.
AU  - Wurst, W.
AU  - Dahlhoff, M.*
AU  - Schneider, M.*
AU  - Wolf, E.*
AU  - Somech, R.*
AU  - Klein, C.*
C1  - 61200
C2  - 49796
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1932-1944
TI  - Mammalian VPS45 orchestrates trafficking through the endosomal system.
JO  - Blood
VL  - 137
IS  - 14
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Clonal hematopoiesis (CH) is an age-related condition predisposing to blood cancer and cardiovascular disease (CVD). Murine models demonstrate CH-mediated altered immune function and proinflammation. Low-grade inflammation has been implicated in the pathogenesis of osteoarthritis (OA), the main indication for total hip arthroplasty (THA). THA-derived hip bones serve as a major source of 'healthy' hematopoietic cells in experimental hematology. We prospectively investigated frequency and clinical associations of CH in 200 patients without known hematologic disease undergoing THA. Prevalence of CH was 50%, including 77 patients with CH of indeterminate potential (CHIP, defined as somatic variants with allele frequencies [VAF] ≥2%), and 23 patients harboring CH with lower mutation burden (VAF 1-2%). Most commonly mutated genes were DNMT3A (29.5%), TET2 (15.0%) and ASXL1 (3.5%). CHIP significantly associated with lower hemoglobin, higher mean corpuscular volume, prior/present malignant disease, and CVD. Strikingly, we observed a previously unreported association of CHIP with autoimmune diseases (AID; multivariate adjusted odds ratio, 6.6; 95% confidence interval [1.7, 30]; p=0.0081). These findings underscore the association between CH and inflammatory diseases. Our results have considerable relevance for management of patients with OA and AID or mild anemia, and question use of hip bone-derived cells as 'healthy' experimental controls.
AU  - Hecker, J.S.*
AU  - Hartmann, L.*
AU  - Riviere, J.*
AU  - Buck, M.C.*
AU  - van der Garde, M.*
AU  - Rothenberg-Thurley, M.*
AU  - Fischer, L.*
AU  - Winter, S.*
AU  - Ksienzyk, B.*
AU  - Ziemann, F.*
AU  - Solovey, M.*
AU  - Rauner, M.*
AU  - Tsourdi, E.*
AU  - Sockel, K.*
AU  - Schneider, M.*
AU  - Kubasch, A.S.*
AU  - Nolde, M.*
AU  - Hausmann, D.*
AU  - Paulus, A.C.*
AU  - Lützner, J.*
AU  - Roth, A.*
AU  - Bassermann, F.*
AU  - Spiekermann, K.*
AU  - Marr, C.
AU  - Hofbauer, L.C.*
AU  - Platzbecker, U.*
AU  - Metzeler, K.H.*
AU  - Götze, K.S.*
C1  - 62330
C2  - 50709
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1727-1732
TI  - CHIP & HIPs: Clonal hematopoiesis is common in hip arthroplasty patients and associates with autoimmune disease.
JO  - Blood
VL  - 138
IS  - 18
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Biomedical applications of deep 
      learning algorithms rely on large, expert annotated data sets. The 
      classification of bone marrow cell cytomorphology, an important 
      cornerstone of hematological diagnosis, is still done manually thousands
       of times every day, due to a lack of datasets and trained models.We
       apply convolutional neural networks (CNNs) to a large dataset of 
      171,374 microscopic cytological images taken from bone marrow smears of 
      945 patients diagnosed with a variety of hematological diseases. The 
      dataset is the largest expert-annotated pool of bone marrow cytology 
      images available in the literature so far. It allows us to train 
      high-quality classifiers of leukocyte cytomorphology that identify a 
      wide range of diagnostically relevant cell species at high precision and
       recall.Our CNNs outcompete previous feature-based approaches and 
      provide a proof-of-concept to the classification problem of single bone 
      marrow cells.This work is a step towards automated 
      evaluation of bone marrow cell morphology using state-of-the-art image 
      classification algorithms. The underlying dataset represents both an 
      educational resource as well as a reference for future AI-based 
      approaches to bone marrow cytomorphology.
AU  - Matek, C.
AU  - Krappe, S.*
AU  - Münzenmayer, C.*
AU  - Haferlach, T.*
AU  - Marr, C.
C1  - 62507
C2  - 50937
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1917-1927
TI  - Highly accurate differentiation of bone marrow cell morphologies using deep neuralnetworks on a large image dataset
JO  - Blood
VL  - 138
IS  - 20
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Orchestrated recruitment of neutrophils to inflamed tissue is essential during initiation of inflammation. Inflamed areas are usually hypoxic, and adaptation to reduced oxygen pressure is typically mediated by hypoxia pathway proteins. However, it is still unclear how these factors influence the migration of neutrophils to and at the site of inflammation either during their transmigration through the blood-endothelial cell barrier, or their motility in the interstitial space. Here, we reveal that activation of the Hypoxia Inducible Factor-2 (HIF2α) due to deficiency of HIF-prolyl hydroxylase domain protein-2 (PHD2) boosts neutrophil migration specifically through highly confined microenvironments. In vivo, the increased migratory capacity of PHD2-deficient neutrophils resulted in massive tissue accumulation in models of acute local inflammation. Using systematic RNAseq analyses and mechanistic approaches, we identified RhoA, a cytoskeleton organizer, as the central downstream factor that mediates HIF2α-dependent neutrophil motility. Thus, we propose that the here identified novel PHD2-HIF2α-RhoA axis is vital to the initial stages of inflammation as it promotes neutrophil movement through highly confined tissue landscapes.
AU  - Sormendi, S.*
AU  - Deygas, M.*
AU  - Sinha, A.*
AU  - Bernard, M.L.M.*
AU  - Krüger, A.*
AU  - Koutzelis, I.*
AU  - Le Lay, G.M.*
AU  - Sáez, P.J.*
AU  - Gerlach, M.*
AU  - Franke, K.*
AU  - Meneses, A.*
AU  - Kräter, M.*
AU  - Palladini, A.
AU  - Guck, J.*
AU  - Coskun, Ü.
AU  - Chavakis, T.*
AU  - Vargas, P.*
AU  - Wielockx, B.*
C1  - 61487
C2  - 50288
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 3416-3427
TI  - HIF2α is a direct regulator of neutrophil motility.
JO  - Blood
VL  - 137
IS  - 24
PB  - Amer Soc Hematology
PY  - 2021
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Anti-CD19 chimeric antigen receptor (CAR) T cells showed significant antileukemic activity in B-precursor acute lymphoblastic leukemia (ALL). Allogeneic, HLA-mismatched off-the-shelf third-party donors may offer ideal fitness of the effector cells, but carry the risk of graft-versus-host disease. Knockout (KO) of the endogenous T-cell receptor (TCR) in CD19-CAR-T cells may be a promising solution. Here, we induced a CRISPR/Cas9-mediated KO of the TCR chain in combination with a second-generation retroviral CAR transduction including a 4-1BB costimulatory domain in primary T cells. This tandem engineering led to a highly functional population of TCR-KO-CAR-T cells with strong activation (CD25, interferon gamma), proliferation, and specific killing upon CD19 target recognition. TCR-KO-CART cells had a balanced phenotype of central memory and effector memory T cells. KO of the endogenous TCR in T cells strongly ablated alloreactivity in comparison with TCR-expressing T cells. In a patient-derived xenograft model of childhood ALL TCR-KO-CAR-T cells clearly controlled CD19(+) leukemia burden and improved survival in vivo. However, coexpression of endogenous TCR plus CAR led to superior persistence of T cells and significantly prolonged leukemia control in vivo, confirmed by a second in vivo model using the leukemia cell line NALM6. These results point toward an essential role of the endogenous TCR for longevity of the response at the price of alloreactivity. In conclusion, anti-CD19 CAR T cells with a CRISPR/Cas9-mediated TCR-KO are promising candidates for nonmatched thirdparty adoptive T-cell transfer with high antileukemic functionality in the absence of alloreactivity, but long-term persistence in vivo is better in the presence of the endogenous TCR.
AU  - Stenger, D.*
AU  - Stief, T.A.*
AU  - Kaeuferle, T.*
AU  - Willier, S.*
AU  - Rataj, F.*
AU  - Schober, K.*
AU  - Vick, B.
AU  - Lotfi, R.*
AU  - Wagner, B.*
AU  - Gruenewald, T.G.P.*
AU  - Kobold, S.*
AU  - Busch, D.H.*
AU  - Jeremias, I.
AU  - Blaeschke, F.*
AU  - Feuchtinger, T.*
C1  - 60350
C2  - 49212
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1407-1418
TI  - Endogenous TCR promotes in vivo persistence of CD19-CAR-T cells compared to a CRISPR/Cas9-mediated TCR knockout CAR.
JO  - Blood
VL  - 136
IS  - 12
PB  - Amer Soc Hematology
PY  - 2020
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Bahrami, E.
AU  - Becker, M.
AU  - Wirth, A.-K.
AU  - Schmid, J.P.
AU  - Herold, T.
AU  - Öllinger, R.*
AU  - Rad, R.*
AU  - Jeremias, I.
C1  - 57952
C2  - 48012
TI  - A CRISPR/Cas9 library screen
      in patients’ leukemia cells in vivo.
JO  - Blood
VL  - 134
PY  - 2019
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - FLT3, DNMT3A, and NPM1 are the most frequently mutated genes in cytogenetically normal acute myeloid leukemia (AML), but little is known about how these mutations synergize upon cooccurrence. Here we show that triple-mutated AML is characterized by high leukemia stem cell (LSC) frequency, an aberrant leukemia-specific GPR56highCD34low immunophenotype, and synergistic upregulation of Hepatic Leukemia Factor (HLF). Cell sorting based on the LSC marker GPR56 allowed isolation of triple-mutated from DNMT3A/NPM1 double-mutated subclones. Moreover, in DNMT3A R882-mutated patients, CpG hypomethylation at the HLF transcription start site correlated with high HLF mRNA expression, which was itself associated with poor survival. Loss of HLF via CRISPR/Cas9 significantly reduced the CD34+GPR56+ LSC compartment of primary human triple-mutated AML cells in serial xenotransplantation assays. HLF knockout cells were more actively cycling when freshly harvested from mice, but rapidly exhausted when reintroduced in culture. RNA sequencing of primary human triple-mutated AML cells after shRNA-mediated HLF knockdown revealed the NOTCH target Hairy and Enhancer of Split 1 (HES1) and the cyclin-dependent kinase inhibitor CDKN1C/p57 as novel targets of HLF, potentially mediating these effects. Overall, our data establish HLF as a novel LSC regulator in this genetically defined high-risk AML subgroup.
AU  - Garg, S.*
AU  - Reyes-Palomares, A.*
AU  - He, L.*
AU  - Bergeron, A.*
AU  - Lavallée, V.P.*
AU  - Lemieux, S.*
AU  - Gendron, P.*
AU  - Rohde, C.*
AU  - Xia, J.*
AU  - Jagdhane, P.*
AU  - Müller-Tidow, C.*
AU  - Lipka, D.B.*
AU  - Imren, S.*
AU  - Humphries, R.K.*
AU  - Waskow, C.*
AU  - Vick, B.
AU  - Jeremias, I.
AU  - Richard-Carpentier, G.*
AU  - Hébert, J.*
AU  - Sauvageau, G.*
AU  - Zaugg, J.*
AU  - Barabé, F.*
AU  - Pabst, C.*
C1  - 56035
C2  - 46785
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 263-276
TI  - Hepatic leukemia factor is a novel leukemic stem cell regulator in DNMT3A, NPM1, and FLT3-ITD triple-mutated AML.
JO  - Blood
VL  - 134
IS  - 3
PB  - Amer Soc Hematology
PY  - 2019
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - CD30 is expressed on a variety of B-cell lymphomas, such as Hodgkin lymphoma, primary effusion lymphoma, and a diffuse large B-cell lymphoma subgroup. In normal tissues, CD30 is expressed on some activated B and T lymphocytes. However, the physiological function of CD30 signaling and its contribution to the generation of CD30(+) lymphomas are still poorly understood. To gain a better understanding of CD30 signaling in B cells, we studied the expression of CD30 in different murine B-cell populations. We show that B1 cells expressed higher levels of CD30 than B2 cells and that CD30 was upregulated in IRF4(+) plasmablasts (PBs). Furthermore, we generated and analyzed mice expressing a constitutively active CD30 receptor in B lymphocytes. These mice displayed an increase in B1 cells in the peritoneal cavity (PerC) and secondary lymphoid organs as well as increased numbers of plasma cells (PCs). TI-2 immunization resulted in a further expansion of B1 cells and PCs. We provide evidence that the expanded B1 population in the spleen included a fraction of PBs. CD30 signals seemed to enhance PC differentiation by increasing activation of NF-kappa B and promoting higher levels of phosphorylated STAT3 and STAT6 and nuclear IRF4. In addition, chronic CD30 signaling led to B-cell lymphomagenesis in aged mice. These lymphomas were localized in the spleen and PerC and had a B1-like/plasmablastic phenotype. We conclude that our mouse model mirrors chronic B-cell activation with increased numbers of CD30(+) lymphocytes and provides experimental proof that chronic CD30 signaling increases the risk of B-cell lymphomagenesis.
AU  - Sperling, S.
AU  - Fiedler, P.
AU  - Lechner, M.
AU  - Pollithy, A.
AU  - Ehrenberg, S.
AU  - Schiefer, A.I.*
AU  - Kenner, L.*
AU  - Feuchtinger, A.
AU  - Kühn, R.*
AU  - Swinerd, G.*
AU  - Schmidt-Supprian, M.*
AU  - Strobl, L.J.
AU  - Zimber-Strobl, U.
C1  - 55826
C2  - 46591
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 2597-2609
TI  - Chronic CD30 signaling in B cells results in lymphomagenesis by driving the expansion of plasmablasts and B1 cells.
JO  - Blood
VL  - 133
IS  - 24
PB  - Amer Soc Hematology
PY  - 2019
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Burchert, A.*
AU  - Bug, G.*
AU  - Finke, J.*
AU  - Stelljes, M.*
AU  - Röllig, C.*
AU  - Waesch, R.*
AU  - Bornhaeuser, M.*
AU  - Berg, T.*
AU  - Lang, F.*
AU  - Ehninger, G.*
AU  - Serve, H.*
AU  - Zeiser, R.*
AU  - Wagner, E.*
AU  - Kroeger, N.*
AU  - Wolschke, C.*
AU  - Schleuning, M.*
AU  - Elmaagacli, A.*
AU  - Goetze, K.S.*
AU  - Schmid, C.*
AU  - Jost, E.*
AU  - Wolf, D.*
AU  - Boehm, A.*
AU  - Thiede, C.*
AU  - Haferlach, T.*
AU  - Bethge, W.*
AU  - Harnisch, S.*
AU  - Wittenberg, M.*
AU  - Rospleszcz, S.
AU  - Neubauer, A.*
AU  - Brugger, M.
AU  - Strauch, K.
AU  - Schade-Brittinger, C.*
AU  - Metzelder, S.K.*
C1  - 55631
C2  - 46403
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Sorafenib as maintenance therapy post allogeneic stem cell transplantation for FLT3-ITD positive AML: Results from the randomized, double-blind, placebo-controlled multicentre sormain trial.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Danisch, S.*
AU  - Slabik, C.*
AU  - Zeidler, R.
AU  - Ganser, A.*
AU  - Stripecke, R.*
C1  - 55591
C2  - 46270
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - CAR-T cells targeting gp350 recognize immortalized cells latently infected with EBV.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - De Braekeleer, E.*
AU  - Hsu, J.*
AU  - Hervieu, C.*
AU  - Tzelepis, K.*
AU  - Indraccolo, S.*
AU  - Warren, A.J.*
AU  - Jeremias, I.
AU  - Antoran, D.F.*
AU  - Jones, P.H.*
AU  - Goodell, M.A.*
AU  - Vassiliou, G.S.*
C1  - 55628
C2  - 46405
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - The nucleotide kinase nadk is required for ROS detoxification and constitutes a metabolic vulnerability of NOTCH1-driven T-ALL.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Ebinger, S.
AU  - Vick, B.
AU  - Spiekermann, K.*
AU  - Jeremias, I.
C1  - 55629
C2  - 46411
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Long-term dormant cells in acute myeloid leukemia patient-derived xenografts display reversible treatment resistance, but are not enriched for leukemia-initiating cells.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The CD33-targeting bispecific T-cell engager (BiTE) AMG 330 proved to be highly efficient in mediating cytolysis of acute myeloid leukemia (AML) cells in vitro and in mouse models. Yet, T-cell activation is correlated with upregulation of programmed cell death-ligand 1 (PD-L1) and other inhibitory checkpoints on AML cells that confer adaptive immune resistance. PD-1 and PD-L1 blocking agents may counteract T-cell dysfunction, however, at the expense of broadly distributed immune-related adverse events (irAEs). We developed a bifunctional checkpoint inhibitory T cell-engaging (CiTE) antibody that combines T-cell redirection to CD33 on AML cells with locally restricted immune checkpoint blockade. This is accomplished by fusing the extracellular domain of PD-1 (PD-1(ex)), which naturally holds a low affinity to PD-L1, to an alpha CD3. alpha CD33 BiTE-like scaffold. By a synergistic effect of checkpoint blockade and avidity-dependent binding, the PD-1(ex) attachment increases T-cell activation (3.3-fold elevation of interferon-g) and leads to efficient and highly selective cytotoxicity against CD33(+)PD-L1(+) cell lines (50% effective concentration 5 2.3-26.9 pM) as well as patient-derived AML cells (n = 8). In a murine xenograft model, the CiTE induces complete AML eradication without initial signs of irAEs as measured by body weight loss. We conclude that our molecule preferentially targets AML cells, whereas high-affinity blockers, such as clinically approved anticancer agents, also address PD-L1(+) non-AML cells. By combining the high efficacy of T-cell engagers with immune checkpoint blockade in a single molecule, we expect to minimize irAEs associated with the systemic application of immune checkpoint inhibitors and suggest high therapeutic potential, particularly for patients with relapsed/refractory AML.
AU  - Hermann, M.*
AU  - Krupka, C.*
AU  - Deiser, K.*
AU  - Brauchle, B.*
AU  - Marcinek, A.*
AU  - Ogrinc Wagner, A.*
AU  - Rataj, F.*
AU  - Mocikat, R.
AU  - Metzeler, K.H.*
AU  - Spiekermann, K.*
AU  - Kobold, S.*
AU  - Fenn, N.C.*
AU  - Hopfner, K.P.*
AU  - Subklewe, M.*
C1  - 54444
C2  - 45593
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 2484-2494
TI  - Bifunctional PD-1 x alpha CD3 x alpha CD33 fusion protein reverses adaptive immune escape in acute myeloid leukemia.
JO  - Blood
VL  - 132
IS  - 23
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Hermann, M.*
AU  - Krupka, C.*
AU  - Deiser, K.*
AU  - Lindl, B.*
AU  - Mocikat, R.
AU  - Metzeler, K.H.*
AU  - Spiekermann, K.*
AU  - Kobold, S.*
AU  - Fenn, N.C.*
AU  - Hopfner, K.*
AU  - Subklewe, M.*
C1  - 55592
C2  - 46269
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Development of a bifunctional checkpoint inhibitory T cell engager (CiTE) to reverse adaptive immune escape in AML.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Kuhn, L.
AU  - Zapf, S.S.
AU  - Djermanovic, K.
AU  - Strobl, D.C.
AU  - Weih, F.*
AU  - Blum, H.*
AU  - Weigert, O.*
AU  - Strobl, L.J.
AU  - Zimber-Strobl, U.
C1  - 55630
C2  - 46404
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - The non-canonical NF-kappaB Signaling Pathway Contributes to the Expansion and Lymphomagenesis of CD40-activated B Cells.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Kyncl, M.*
AU  - Bast, L.
AU  - Henkel, L.*
AU  - Theis, F.J.
AU  - Oostendorp, R.A.J.*
AU  - Marr, C.
AU  - Goetze, K.S.*
C1  - 55593
C2  - 46268
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Data driven computational modeling of hematopoiesis in myelodysplastic syndromes unveils differences in hematopoietic stem cell kinetics compared to age-matched healthy controls.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Lenk, L.*
AU  - Vogiatzi, F.*
AU  - Carlet, M.
AU  - Vokuhl, C.*
AU  - Cario, G.*
AU  - Schrappe, M.*
AU  - Jeremias, I.
AU  - Fuhrmann, S.*
AU  - Hobeika, E.*
AU  - Jumaa, H.*
AU  - Alsadeq, A.*
AU  - Schewe, D.M.*
C1  - 55626
C2  - 46407
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - CD79a is associated with central nervous system infiltration of pediatric B-cell precursor acute lymphoblastic leukemia.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Passerini, V.*
AU  - Bösl, M.R.
AU  - Silkenstedt, E.*
AU  - Osterode, E.*
AU  - Heide, M.*
AU  - Kridel, R.*
AU  - Ziegenhain, C.*
AU  - Staiger, A.M.*
AU  - Ott, G.*
AU  - Szczepanowski, M.*
AU  - Richter, J.*
AU  - Klapper, W.*
AU  - Rosenwald, A.*
AU  - Enard, W.*
AU  - Hiddemann, W.*
AU  - Bararia, D.*
AU  - Zimber-Strobl, U.
AU  - Weigert, O.*
C1  - 55589
C2  - 46267
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - PARP14 is a novel therapeutic target in STAT6 mutant follicular lymphoma.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Stief, S.M.*
AU  - Hanneforth, A.*
AU  - Mattes, R.*
AU  - Weser, S.*
AU  - Vick, B.
AU  - Bartoschek, M.D.*
AU  - Moreno, H.D.*
AU  - Liu, W.-H.
AU  - Ksienzyk, B.*
AU  - Rothenberg-Thurley, M.*
AU  - Quentmeier, H.*
AU  - Hiddemann, W.*
AU  - Metzeler, K.H.*
AU  - Schotta, G.*
AU  - Bultmann, S.*
AU  - Jeremias, I.
AU  - Leonhardt, H.*
AU  - Spiekermann, K.*
C1  - 55590
C2  - 46271
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Loss of KDM6A confers drug resistance in acute myeloid leukemia.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Stripecke, R.*
AU  - Danisch, S.*
AU  - Slabik, C.*
AU  - Zeidler, R.
AU  - Hammerschmidt, W.
AU  - Feuerhake, F.*
AU  - Ganser, A.*
C1  - 55627
C2  - 46406
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
TI  - Pembrolizumab therapy exacerbates EBV-induced infections and tumors in a long-term fully humanized mouse model.
JO  - Blood
VL  - 132
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Many hemostatic factors are associated with age and age-related diseases; however, much remains unknown about the biological mechanisms linking aging and hemostatic factors. DNA methylation is a novel means by which to assess epigenetic aging, which is a measure of age and the aging processes as determined by altered epigenetic states. We used a meta-analysis approach to examine the association between measures of epigenetic aging and hemostatic factors, as well as a clotting time measure. For fibrinogen, we performed European and African ancestry-specific meta-analyses which were then combined via a random effects meta-analysis. For all other measures we could not estimate ancestry-specific effects and used a single fixed effects meta-analysis. We found that 1-year higher extrinsic epigenetic age as compared with chronological age was associated with higher fibrinogen (0.004 g/L/y; 95% confidence interval, 0.001-0.007; P = .01) and plasminogen activator inhibitor 1 (PAI-1; 0.13 U/mL/y; 95% confidence interval, 0.07-0.20; P = 6.6 x 10(-5)) concentrations, as well as lower activated partial thromboplastin time, a measure of clotting time. We replicated PAI-1 associations using an independent cohort. To further elucidate potential functional mechanisms, we associated epigenetic aging with expression levels of the PAI-1 protein encoding gene (SERPINE1) and the 3 fibrinogen subunit-encoding genes (FGA, FGG, and FGB) in both peripheral blood and aorta intima-media samples. We observed associations between accelerated epigenetic aging and transcription of FGG in both tissues. Collectively, our results indicate that accelerated epigenetic aging is associated with a procoagulation hemostatic profile, and that epigenetic aging may regulate hemostasis in part via gene transcription.
AU  - Ward-Caviness, C.K.
AU  - Huffman, J.E.*
AU  - Evertt, K.*
AU  - Germain, M.*
AU  - van Dongen, J.*
AU  - Hill, W.D.*
AU  - Jhun, M.A.*
AU  - Brody, J.A.*
AU  - Ghanbari, M.*
AU  - Du, L.*
AU  - Roetker, N.S.*
AU  - de Vries, P.S.*
AU  - Waldenberger, M.
AU  - Gieger, C.
AU  - Wolf, P.
AU  - Prokisch, H.
AU  - Koenig, W.*
AU  - O'Donnell, C.J.*
AU  - Levy, D.*
AU  - Liu, C.*
AU  - Truong, V.*
AU  - Wells, P.S.*
AU  - Trégouët, D.A.*
AU  - Tang, W.*
AU  - Morrison, A.C.*
AU  - Boerwinkle, E.*
AU  - Wiggins, K.L.*
AU  - McKnight, B.*
AU  - Guo, X.*
AU  - Psaty, B.M.*
AU  - Sotoodehnia, N.*
AU  - Boomsa, D.I.*
AU  - Willemsen, G.*
AU  - Ligthart, L.*
AU  - Deary, I.J.*
AU  - Zhao, W.*
AU  - Ware, E.B.*
AU  - Kardia, S.L.R.*
AU  - van Meurs, J.B.J.*
AU  - Uitterlinden, A.G.*
AU  - Franco, O.H.*
AU  - Eriksson, P.*
AU  - Franco-Cereceda, A.*
AU  - Pankow, J.S.*
AU  - Johnson, A.D.*
AU  - Gagnon, F.*
AU  - Morange, P.E.*
AU  - de Geus, E.J.C.*
AU  - Starr, J.M.*
AU  - Smith, J.A.*
AU  - Dehghan, A.*
AU  - Björck, H.M.*
AU  - Smith, N.L.*
AU  - Peters, A.
C1  - 54003
C2  - 45195
CY  - 2021 L St Nw, Suite 900, Washington, Dc 20036 Usa
SP  - 1842-1850
TI  - DNA methylation age is associated with an altered hemostatic profile in a multiethnic meta-analysis.
JO  - Blood
VL  - 132
IS  - 17
PB  - Amer Soc Hematology
PY  - 2018
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - There is high interest in understanding the mechanisms that drive self-renewal of stem cells. HOXB4 is one of the few transcription factors that can amplify long-term repopulating hematopoietic stem cells in a controlled way. Here we show in mice that this characteristic of HOXB4 depends on a proline-rich sequence near the N terminus, which is unique among HOX genes and highly conserved in higher mammals. Deletion of this domain substantially enhanced the oncogenicity of HOXB4, inducing acute leukemia in mice. Conversely, insertion of the domain into Hoxa9 impaired leukemogenicity of this homeobox gene. These results indicate that proline-rich stretches attenuate the potential of stem cell active homeobox genes to acquire oncogenic properties.
AU  - Cusan, M.
AU  - Vegi, N.M.*
AU  - Mulaw, M.A.*
AU  - Bamezai, S.*
AU  - Kaiser, L.M.*
AU  - Deshpande, A.J.*
AU  - Greif, P.A.
AU  - Quintanilla-Fend, L.*
AU  - Göllner, S.*
AU  - Müller-Tidow, C.*
AU  - Humphries, K.R.*
AU  - Armstrong, S.A.*
AU  - Hiddemann, W.
AU  - Feuring-Buske, M.*
AU  - Buske, C.*
C1  - 51892
C2  - 43560
SP  - 319-323
TI  - Controlled stem cell amplification by HOXB4 depends on its unique proline-rich region near the N terminus.
JO  - Blood
VL  - 129
IS  - 3
PY  - 2017
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Mantle cell lymphoma (MCL) is a mature B-cell lymphoma characterized by poor clinical outcome. Recent studies revealed the importance of B-cell receptor (BCR) signaling in maintaining MCL survival. However, it remains unclear which role MALT1, an essential component of the CARD11-BCL10-MALT1 complex that links BCR signaling to the NF-κB pathway, plays in the biology of MCL. Here we show that a subset of MCLs is addicted to MALT1, as its inhibition by either RNA or pharmacologic interference induced cytotoxicity both in vitro and in vivo. Gene expression profiling following MALT1 inhibition demonstrated that MALT1 controls an MYC-driven gene expression network predominantly through increasing MYC protein stability. Thus, our analyses identify a previously unappreciated regulatory mechanism of MYC expression. Investigating primary mouse splenocytes, we could demonstrate that MALT1-induced MYC regulation is not restricted to MCL, but represents a common mechanism. MYC itself is pivotal for MCL survival because its downregulation and pharmacologic inhibition induced cytotoxicity in all MCL models. Collectively, these results provide a strong mechanistic rationale to investigate the therapeutic efficacy of targeting the MALT1-MYC axis in MCL patients.
AU  - Dai, B.*
AU  - Grau, M.*
AU  - Juilland, M.*
AU  - Klener, P.*
AU  - Höring, E.*
AU  - Molinsky, J.*
AU  - Schimmack, G.
AU  - Aukema, S.M.*
AU  - Hoster, E.*
AU  - Vogt, N.*
AU  - Staiger, A.M.*
AU  - Erdmann, T.*
AU  - Xu, W.*
AU  - Erdmann, K.*
AU  - Dzyuba, N.*
AU  - Madle, H.*
AU  - Berdel, W.E.*
AU  - Trneny, M.*
AU  - Dreyling, M.*
AU  - Jöhrens, K.*
AU  - Lenz, P.*
AU  - Rosenwald, A.*
AU  - Siebert, R.*
AU  - Tzankov, A.*
AU  - Klapper, W.*
AU  - Anagnostopoulos, I.*
AU  - Krappmann, D.
AU  - Ott, G.*
AU  - Thome, M.*
AU  - Lenz, G.*
C1  - 50723
C2  - 42471
CY  - Washington
SP  - 333-346
TI  - B-cell receptor-driven MALT1 activity regulates MYC signaling in mantle cell lymphoma.
JO  - Blood
VL  - 129
IS  - 3
PB  - Amer Soc Hematology
PY  - 2017
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Controlled regulation of lineage decisions is imperative for hematopoiesis. Yet, the molecular mechanisms underlying hematopoietic lineage choices are poorly defined. CSF-1, the cytokine acting as the principal regulator of monocyte/macrophage development, has been shown to be able to instruct the lineage choice of uncommitted granulocyte macrophage progenitors towards a monocyte/macrophage fate. However, the intracellular signaling pathways involved are unknown. CSF-1 activates a multitude of signaling pathways resulting in a pleiotropic cellular response. The precise role of individual pathways within this complex and redundant signaling network is dependent on cellular context and not well understood. Here, we addressed which CSF-1-activated pathways are involved in transmitting the lineage-instructive signal in primary bone marrow-derived granulocyte macrophage progenitors. While its loss is compensated for by alternative signaling activation mechanisms, Src family kinase signaling is sufficient to transmit the CSF-1 lineage instructive signal. Moreover, c-Src activity is sufficient to drive macrophage fate, even in non-myeloid cells.
AU  - Endele, M.
AU  - Loeffler, D.
AU  - Kokkaliaris, K.D.
AU  - Hilsenbeck, O.
AU  - Skylaki, S.
AU  - Hoppe, P.S.
AU  - Schambach, A.*
AU  - Stanley, E.R.*
AU  - Schroeder, T.
C1  - 50658
C2  - 42835
SP  - 1691-1701
TI  - CSF-1-induced Src signaling can instruct monocytic lineage choice.
JO  - Blood
VL  - 129
IS  - 12
PY  - 2017
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Maintaining cellular redox balance is vital for cell survival and tissue homoeostasis since imbalanced production of ROS may lead to oxidative stress and cell death. The anti-oxidant enzyme glutathione peroxidase 4 (Gpx4) is a key regulator of oxidative stress-induced cell death. We show that mice with deletion of Gpx4 in hematopoietic cells develop anemia and that it is essential for preventing RIP3 dependent necroptosis in erythroid precursor cells. Absence of Gpx4 leads to functional inactivation of caspase 8 by glutathionylation. This results in necroptosis, which occurs independently of TNFα activation. While genetic ablation of Rip3 normalizes reticulocyte maturation and prevents anemia, ROS accumulation and lipid peroxidation in Gpx4 deficient cells remain high. Our results demonstrate that ROS and lipid hydroperoxides function as so far unrecognized unconventional upstream signaling activators of RIP3-dependent necroptosis.
AU  - Canli, Ö.*
AU  - Alankus, Y.B.*
AU  - Grootjans, S.*
AU  - Vegi, N.*
AU  - Hültner, L.
AU  - Hoppe, P.S.
AU  - Schroeder, T.*
AU  - Vandenabeele, P.*
AU  - Bornkamm, G.W.
AU  - Greten, F.R.*
C1  - 47180
C2  - 39139
CY  - Washington
SP  - 139-148
TI  - Glutathione peroxidase 4 prevents necroptosis in mouse erythroid precursors.
JO  - Blood
VL  - 127
IS  - 1
PB  - Amer Soc Hematology
PY  - 2016
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The Bruton tyrosine kinase (BTK) inhibitor ibrutinib induces responses in 70% of patients with relapsed and refractory mantle cell lymphoma (MCL). Intrinsic resistance can occur through activation of the nonclassical NF-κB pathway and acquired resistance may involve the BTK C481S mutation. Outcomes after ibrutinib failure are dismal, indicating an unmet medical need. We reasoned that newer heat shock protein 90 (HSP90) inhibitors could overcome ibrutinib resistance by targeting multiple oncogenic pathways in MCL. HSP90 inhibition induced the complete degradation of both BTK and IκB kinase α in MCL lines and CD40-dependent B cells, with downstream loss of MAPK and nonclassical NF-κB signaling. A proteome-wide analysis in MCL lines and an MCL patient-derived xenograft identified a restricted set of targets from HSP90 inhibition that were enriched for factors involved in B-cell receptor and JAK/STAT signaling, the nonclassical NF-κB pathway, cell-cycle regulation, and DNA repair. Finally, multiple HSP90 inhibitors potently killed MCL lines in vitro, and the clinical agent AUY922 was active in vivo against both patient-derived and cell-line xenografts. Together, these findings define the HSP90-dependent proteome in MCL. Considering the disappointing clinical activity of HSP90 inhibitors in other contexts, trials in patients with MCL will be essential for defining the efficacy of and mechanisms of resistance after ibrutinib failure.
AU  - Jacobsen, C.*
AU  - Kopp, N.*
AU  - Layer, J.*
AU  - Redd, R.A.*
AU  - Tschuri, S.*
AU  - Haebe, S.*
AU  - van Bodegom, D.*
AU  - Bird, L.*
AU  - Christie, A.L.*
AU  - Christodoulou, A.N.*
AU  - Saur, A.*
AU  - Tivey, T.*
AU  - Zapf, S.S.
AU  - Bararia, D.*
AU  - Zimber-Strobl, U.
AU  - Rodig, S.J.*
AU  - Weigert, O.*
AU  - Weinstock, D.M.*
C1  - 50305
C2  - 42082
CY  - Washington
SP  - 2517-2526
TI  - HSP90 inhibition overcomes ibrutinib resistance in mantle cell lymphoma.
JO  - Blood
VL  - 128
IS  - 21
PB  - Amer Soc Hematology
PY  - 2016
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The maintenance of hematopoietic stem cells (HSCs) during ex vivo culture is an important prerequisite for their therapeutic manipulation. However, despite intense research, culture conditions for robust maintenance of HSCs are still missing. Cultured HSCs are quickly lost, preventing their improved analysis and manipulation. Identification of novel factors supporting HSC ex vivo maintenance is therefore necessary. Co-culture with the AFT024 stroma cell line is capable of maintaining HSCs ex vivo long-term, but the responsible molecular players remain unknown. Here, we use continuous long-term single-cell observation to identify the HSC behavioral signature under supportive or non-supportive stroma co-cultures. We report early HSC survival as a major characteristic of HSC-maintaining conditions. Behavioral screening after manipulation of candidate molecules revealed that the extracellular matrix protein dermatopontin (Dpt) is involved in HSC maintenance. DPT knock-down in supportive stroma impaired HSC survival, while ectopic expression of the Dpt gene or protein in non-supportive conditions restored HSC survival. Supplementing defined stroma- and serum-free culture conditions with recombinant DPT protein improved HSC clonogenicity. These findings illustrate a previously uncharacterized role of Dpt in maintaining HSCs ex vivo.
AU  - Kokkaliaris, K.D.
AU  - Drew, E.
AU  - Endele, M.
AU  - Loeffler, D.
AU  - Hoppe, P.S.
AU  - Hilsenbeck, O.
AU  - Schauberger, B.
AU  - Hinzen, C.
AU  - Skylaki, S.
AU  - Theodorou, M.
AU  - Kieslinger, M.
AU  - Lemischka, I.*
AU  - Moore, K.*
AU  - Schroeder, T.
C1  - 48980
C2  - 41503
CY  - Washington
SP  - 1181-1192
TI  - Identification of factors promoting ex vivo maintenance of mouse hematopoietic stem cells by long-term single-cell quantification.
JO  - Blood
VL  - 128
IS  - 9
PB  - Amer Soc Hematology
PY  - 2016
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Human CD34(+) hematopoietic stem and progenitor cells (HSPCs) can reconstitute a human hemato-lymphoid system when transplanted into immunocompromised mice. While fetal liver- and cord blood-derived CD34(+) cells lead to high engraftment levels, engraftment of mobilized, adult donor-derived CD34(+) cells has remained poor. We generated so-called MSTRG and MISTRG hu-manized mice on a Rag2(-/-)Il2rg(-/-) background carrying a transgene for human SIRPα and human homologues of the cytokines macrophage-colony stimulating factor, thrombopoietin, with or without interleukin-3 and granulocyte-macrophage colony stimulating factor under murine promotors. Here we transplanted mobilized peripheral blood CD34(+) cells in sub-lethally irradiated newborn and adult recipients. Human hematopoietic engraftment levels were significantly higher in bone marrow, spleen and peripheral blood in newborn transplanted MSTRG/MISTRG as compared to non-obese diabetic/severe combined immunodeficient Il2rg(-/-) or human SIRPα-transgenic Rag2(-/-)Il2rg(-/-) recipients. Furthermore newborn transplanted MSTRG/MISTRG mice supported higher engraftment levels of human phenotypically defined HSPCs in bone marrow, T-cells in the thymus, and myeloid cells in non-hematopoietic organs such as liver, lung, colon and skin, approximating the levels in the human system. Similar results were obtained in adult recipient mice. Thus, human cytokine knock-in mice might open new avenues for personalized studies of human (patho)physiology of the hematopoietic and immune system in vivo.
AU  - Saito, Y.*
AU  - Ellegast, J.M.*
AU  - Rafiei, A.*
AU  - Song, Y.*
AU  - Kull, D.
AU  - Heikenwälder, M.
AU  - Rongvaux, A.*
AU  - Halene, S.*
AU  - Flavell, R.A.*
AU  - Manz, M.G.*
C1  - 49289
C2  - 41725
CY  - Washington
SP  - 1829-1833
TI  - Peripheral blood CD34+ cells efficiently engraft human cytokine knock-in mice.
JO  - Blood
VL  - 128
IS  - 14
PB  - Amer Soc Hematology
PY  - 2016
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Deep venous thrombosis (DVT) is one of the most common cardiovascular diseases, but its pathophysiology remains incompletely understood. While sterile inflammation has recently been shown to boost coagulation during DVT, the underlying molecular mechanisms are not fully resolved, which could potentially identify new anti-inflammatory approaches to prophylaxis and therapy of DVT. Using a mouse model of venous thrombosis induced by flow reduction in the vena cava inferior we identified blood-derived high-mobility group box 1 protein (HMGB1) - a prototypical mediator of sterile inflammation - to be a master regulator of the prothrombotic cascade involving platelets and myeloid leukocytes fostering occlusive DVT formation. Transfer of platelets into Hmgb1(-/-) chimeras showed that this cell type is the major source of HMGB1, exposing reduced HMGB1 on their surface upon activation thereby enhancing the recruitment of monocytes. Activated leukocytes in turn support oxidation of HMGB1 unleashing its prothrombotic activity and promoting platelet aggregation. This potentiates the amount of HMGB1 and further nurtures the accumulation and activation of monocytes through RAGE and TLR2, leading to local delivery of monocyte-derived tissue factor and cytokines. Moreover, disulfide HMGB1 facilitates formation of prothrombotic neutrophil extracellular traps (NETs) mediated by RAGE, exposing additional HMGB1 on their extracellular DNA strands. Eventually, a vicious circle of coagulation and inflammation is set in motion leading to obstructive DVT formation. Therefore, platelet derived disulfide HMGB1 is a central mediator of the sterile inflammatory process in venous thrombosis and could be an attractive target for an anti-inflammatory approach for DVT prophylaxis.
AU  - Stark, K.*
AU  - Philippi, V.*
AU  - Stockhausen, S.*
AU  - Busse, J.*
AU  - Antonelli, A.*
AU  - Miller, M.*
AU  - Schubert, I.*
AU  - Hoseinpour, P.*
AU  - Chandraratne, S.*
AU  - von Brühl, M.L.*
AU  - Gärtner, F.*
AU  - Lorenz, M.*
AU  - Agresti, A.*
AU  - Coletti, R.*
AU  - Antoine, D.J.*
AU  - Heermann, R.*
AU  - Jung, K.*
AU  - Reese, S.*
AU  - Laitinen, I.*
AU  - Schwaiger, M.*
AU  - Walch, A.K.
AU  - Sperandio, M.*
AU  - Nawroth, P.P.*
AU  - Reinhardt, C.*
AU  - Jäckel, S.*
AU  - Bianchi, M.E.*
AU  - Massberg, S.*
C1  - 49346
C2  - 41758
CY  - Washington
SP  - 2435-2449
TI  - Disulfide HMGB1 derived from platelets coordinates venous thrombosis in mice.
JO  - Blood
VL  - 128
IS  - 20
PB  - Amer Soc Hematology
PY  - 2016
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Since their discovery in patients with autosomal dominant (AD) chronic mucocutaneous candidiasis (CMC) in 2011, heterozygous STAT1 gain-of-function (GOF) mutations have increasingly been identified worldwide. The clinical spectrum associated with them needed to be delineated. We enrolled 274 patients from 167 kindreds originating from 40 countries from five continents. Demographic data, clinical features, immunological parameters, treatment, and outcome were recorded. The median age of the 274 patients was 22 years (range: 1 - 71 years); 98% of them had CMC, with a median age at onset of one year (range: 0 - 24 years). Patients often displayed bacterial (74%) infections, mostly due to Staphylococcus aureus (36%), including the respiratory tract and the skin in 47% and 28% of patients, respectively, and viral (38%) infections, mostly due to Herpesviridae (83%) and affecting the skin in 32% of patients. Invasive fungal infections (10%), mostly caused by Candida spp. (29%), and mycobacterial disease (6%) caused by Mycobacterium tuberculosis, environmental mycobacteria, or BCG vaccines, were less common. Many patients had autoimmune manifestations (37%), including hypothyroidism (22%), type 1 diabetes (4%), blood cytopenia (4%), and systemic lupus erythematosus (2%). Invasive infections (25%), cerebral aneurysms (6%), and cancers (6%) were the strongest predictors of poor outcome. CMC persisted in 39% of the 202 patients receiving prolonged antifungal treatment. Circulating IL-17A-producing T-cell count was low for most (82%) but not all of the patients tested. STAT1 GOF mutations underlie AD CMC, as well as an unexpectedly wide range of other clinical features, including not only a variety of infectious and autoimmune diseases, but also cerebral aneurysms and carcinomas that confer a poor prognosis.
AU  - Toubiana, J.*
AU  - Okada, S.*
AU  - Hiller, J.
AU  - Oleastro, M.*
AU  - Lagos Gomez, M.*
AU  - Aldave Becerra, J.C.*
AU  - Ouachée-Chardin, M.*
AU  - Fouyssac, F.*
AU  - Girisha, K.M.*
AU  - Etzioni, A.*
AU  - van Montfrans, J.*
AU  - Camcioglu, Y.*
AU  - Kerns, L.A.*
AU  - Belohradsky, B.*
AU  - Blanche, S.*
AU  - Bousfiha, A.*
AU  - Rodriguez-Gallego, C.*
AU  - Meyts, I.*
AU  - Kisand, K.*
AU  - Reichenbach, J.*
AU  - Renner, E.D.*
AU  - Rosenzweig, S.*
AU  - Grimbacher, B.*
AU  - van de Veerdonk, F.L.*
AU  - Traidl-Hoffmann, C.
AU  - Picard, C.*
AU  - Maródi, L.*
AU  - Morio, T.*
AU  - Kobayashi, M.*
AU  - Lilic, D.*
AU  - Milner, J.D.*
AU  - Holland, S.*
AU  - Casanova, J.L.*
AU  - Puel, A.*
C1  - 48558
C2  - 41185
CY  - Washington
SP  - 3154-3164
TI  - Heterozygous STAT1 gain-of-function mutations underlie an unexpectedly broad clinical phenotype: An international survey of 274 patients from 167 kindreds.
JO  - Blood
VL  - 127
IS  - 25
PB  - Amer Soc Hematology
PY  - 2016
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Expression profiling and next generation sequencing have enabled a detailed knowledge on alterations present in tumors from individual patients. In contrast, only limited understanding exists on the role that each alteration plays for the existing tumor. Of direct clinical interest, genes are of special interest which harbor an essential function for tumor maintenance and growth as they represent putative targets for anti-cancer therapy.
      
      	Characterizing gene functions is demanding regarding both techniques and resources. Questions on gene function are often studied in established tumor cell lines, although establishing cell lines from primary tumors is rarely successful in acute leukemias. Primary acute leukemia cells poorly grow in vitro and established acute leukemias cell lines rely on additional mutations enabling in vitro growth, making them doubtful models to study genes with essential function.
      
      	To bridge the gap, we aimed at studying gene function in the complex environment of individual tumors. As primary acute leukemia cells are unable to grow in vitro, we used the orthotopic model of patient-derived xenograft (PDX) leukemia and amplified cells in mice.
      
      	We established a novel technique to manipulate distinct signaling proteins in PDX cells using lentiviruses and knockdown. We expressed small hairpin RNA (shRNA) in the background of micro RNA 30 (miR30) under control of a Pol II promoter and 3 prime of dsRED as molecular marker. This approach closely links expression of the shRNA to the fluorochrome and resulted in a potent and stable knockdown. We expressed a control shRNA targeting Renilla luciferase and several shRNA sequences targeting XIAP. In order to discriminate different derivative cell populations within a single mouse, we co-expressed a second fluorochrome from a second plasmid so that green cells harbored a knockdown of XIAP, while blue cells harbored the control construct, thus allowing in vivo outcompete proliferation assays. We called our new approach "genetically engineered PDX (GEPDX)" models in parallel to genetically engineered mouse models (GEMM).
      
      	We used our new technology to study the role of XIAP, the X-linked inhibitor of apoptosis for acute lymphoblastic leukemia (ALL), the single most frequent tumor in children. XIAP is frequently and highly overexpressed in hematological malignancies and its up-regulation was shown to be associated with inferior prognosis of patients in different tumors. Nevertheless XIAP's role for tumor maintenance remains unclear.
      
      	In several preB- and T-cell ALL cell lines, potent and stable knockdown of XIAP did not alter cell proliferation in vitro or upon xeno-transplantation in vivo. Thus expression levels of XIAP seem irrelevant for spontaneous proliferation of established acute leukemia cell lines in vitro and in vivo.
      
      	We next studied PDX ALL cells growing in mice as model closer related to patients. We generated GEPDX cells from two children with relapse of a B precursor ALL with either knockdown of XIAP or control knockdown together with the appropriate molecular color markers. When blue and green GEPDX cells harboring control or XIAP knockdown, respectively, were co-transplanted into mice at a 1:1 ratio in a competitive outcompete assay, control transfected cells significantly overgrew or even eliminated cells with XIAP knockdown in both samples studied. GEPDX cells with knockdown of XIAP showed a significant and dose-dependent growth disadvantage in vivo compared to control cells indicating that XIAP played an essential role for PDX cells growing in vivo.
      
      	Thus, our novel technique of genetic engineering in PDX cells revealed an essential role of XIAP for tumor maintenance and growth in patients' tumor cells making XIAP an attractive therapeutic target in ALL. As established ALL cell lines were unable to unravel this important role of XIAP, GEPDX might be superior to cell lines for identifying genes with essential function. GEPDX represent a powerful new tool to characterize the complex environment of individual patients' tumor cells in vivo, the function of the many lesions and alterations described by expression profiling and next generation sequencing.
AU  - Carlet, M.
AU  - Vergalli, J.
AU  - Finkenzeller, C.
AU  - Grunert, M.
AU  - Jeremias, I.
C1  - 48029
C2  - 39857
CY  - Washington
TI  - The novel technique of Genetically Engineered Patient-Derived Xenografts (GEPDX) reveals that the X-linked Inhibitor of Apoptosis Protein (XIAP) plays an essential role for maintenance and growth of patients' acute lymphoblastic leukemia in vivo.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Postremission therapy for acute myeloid leukemia (AML) is critical for elimination of minimal residual disease (MRD). In patients not eligible for allogeneic stem cell transplantation, alternative treatment options are needed. Therapeutic vaccination with autologous dendritic cells (DCs) loaded with leukemia-associated antigens (LAAs) is a promising treatment strategy to induce anti-leukemic immune responses and to eradicate chemorefractory cells. We have developed a GMP-compliant 3-day protocol including a TLR7/8 agonist to differentiate monocytes of intensively pretreated AML patients into next-generation DCs.
      
      	A phase I/II proof-of-concept study has been initiated using next-generation DCs as postremission therapy of AML patients with a non-favorable genetic risk profile in CR after intensive induction therapy (NCT01734304). DCs are loaded with in vitro transcribed RNA encoding the LAAs WT1 and PRAME as well as CMVpp65 as adjuvant and surrogate antigen. Patients are vaccinated intradermally with 5x106 DCs of each antigen species up to 10 times within 26 weeks. The primary endpoint of the phase I/II trial is feasibility and safety of the vaccination. Secondary endpoints are immunological responses and disease control.
      
      	Based on the safety and toxicity profile of the phase I trial (n=6), phase II has been initiated. In total, 10 patients have been enrolled into the study. DCs of sufficient number and quality were generated from leukapheresis in 8/9 cases. DCs exhibited an immune-stimulatory profile based on high surface expression of positive costimulatory molecules, the capacity to secrete IL-12p70, the migration towards a chemokine gradient and processing and presentation of antigen. 5 patients have completed the vaccination schedule; the 6th and 7th patient have received 7/10 and 4/10 vaccinations, respectively. We observed delayed-type hypersensitivity (DTH) responses at the vaccination site in 6/6 patients, accompanied by slight erythema and indurations at the injection site, but no grade III/IV toxicities. TCR repertoire analysis by next-generation sequencing revealed an enrichment of particular clonotypes at DTH sites. Limited by HLA restriction, we have so far analyzed 4 patients by multimer staining. All of them mounted DC vaccination-specific T cell responses: We detected an increase of WT1-specific T cells in one patient and strong expansion/induction of CMVpp65-specific T cells in one CMV-seropositive and two CMV-seronegative patients. In an individual treatment attempt, an enrolled patient with impending relapse was treated with a combination of DC vaccination and 5-azacytidine, resulting in MRD conversion. Long-term disease control and immunological responses are studied in the ongoing phase II trial.
      
      	We conclude that vaccination with next-generation LAA-expressing DCs in AML is feasible, safe and induces anti-leukemia-specific immune responses in vivo.
AU  - Deiser, K.
AU  - Lichtenegger, F.S.
AU  - Schnorfeil, F.M.
AU  - Köhnke, T.*
AU  - Altmann, T.*
AU  - Buecklein, V.*
AU  - Moosmann, A.
AU  - Brueggemann, M.*
AU  - Wagner, B.*
AU  - Hiddemann, W.*
AU  - Bigalke, I.*
AU  - Kvalheim, G.*
AU  - Subklewe, M.
C1  - 48032
C2  - 39854
CY  - Washington
TI  - Next-generation dendritic cell vaccination in postremission therapy of AML: Results of a clinical phase I trial.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Treatment-resistant cells determine prognosis and outcome of cancer patients as they induce relapse with poor outcome. Novel therapeutic options are urgently needed to eradicate chemo-resistant tumor cells. Towards this aim, a deep understanding is required on mechanisms determining treatment-resistance in vivo and the ability to induce relapse.
      
      	Here, we aimed to identify and characterize the challenging rare subpopulation of drug resistant cells which survive in vivo chemo-therapy and are able to induce relapse.
      
      	As technical approach, we used the individualized mouse model of acute lymphoblastic leukemia (ALL) and amplified primary tumor cells in mice to generate patient-derived xenograft (PDX) cells. Upon genetic engineering by lentiviral transduction, PDX ALL cells expressed the three transgenes NGFR, a red fluorochrome and luciferase. While recombinant luciferase was used for in vivo imaging, a combined MACS/FACS procedure based on the expression of the transgenes enabled enriching PDX ALL cells from murine bone marrow by a factor above 10,000. Staining of PDX cells with CFSE was used to discriminate between highly proliferative and dormant tumor cells in vivo.
      
      	We treated mice harboring triple-transgenic, CFSE labeled PDX ALL cells with conventional chemotherapy; while in vivo treatment decreased the number of highly proliferative cells by more than 1 order of magnitude, the amount of dormant cells remained completely unchanged. Isolated, drug resistant cells revealed leukemia propagating potential and induced leukemia upon transplantation into next generation mice. Thus and using dormancy as an anchor, we could identify, isolate and enrich a subpopulation of treatment-resistant PDX ALL cells which might mimic relapse-inducing cells at minimal residual disease in patients.
      
      	We next aimed at characterizing the expression profile of these cells and were able to isolate single cells out of the low number of dormant cells to perform single cell RNA sequencing. Dormant, drug-resistant cells showed increased expression of several adhesion molecules suggesting an increased dependence on the bone marrow environment. Upon using gene set enrichment analyses, drug-resistant cells showed a highly similar expression profile to primary high risk leukemia subpopulations such as primary high risk ALL cells, the subpopulation of CD34 positive CML cells and leukemic or benign hematopoietic stem cells.
      
      	Taken together, treatment-resistant PDX ALL stem cells isolated and enriched from mice showed increased expression of adhesion molecules and resemble primary tumor cells of high risk subpopulations. These cells represent valuable tools to increase our understanding of mechanisms in minimal residual disease and relapse in patients. Our model will help to develop novel therapies which eliminate treatment resistant cells, prevent disease relapse and increase the prognosis of patients with ALL.
AU  - Ebinger, S.
AU  - Erbey, O.
AU  - Tiedt, S.
AU  - Ziegenhain, C.*
AU  - Alves, C.C.
AU  - Enard, W.*
AU  - Jeremias, I.
C1  - 48025
C2  - 39861
CY  - Washington
TI  - Single cell RNA sequencing reveals increased adhesion signals in treatment-resistant tumor stem cells in a preclinical mouse model of genetically engineered patient-derived acute lymphoblastic leukemia.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Acute lymphoblastic leukemia (ALL) consists of genetically heterogeneous cell subpopulations, but little is known about how genetic differences lead to functional differences between the clones. Of major clinical importance, aggressive, treatment-resistant and putatively relapse-inducing subclones need to be identified and require effective eradication by treatment. The most aggressive subpopulation likely determines prognosis and outcome in each patient. We aimed at characterizing on a functional as well as on a genetic level single stem cell clones derived from patients' samples growing in mice and to combine the results of both levels in order to learn which genetic characteristics are associated with adverse functional behavior. We transplanted primary tumor cells from a 5-year old girl with hyperdiploid ALL involving a trisomic X chromosome at first relapse into severely immune-compromised mice and lentivirally modified them to express the fluorochromes red, blue and green at different amounts and combinations (RGB marking, Weber et al., 2012). Eight single stem cell clones were generated by limiting dilution transplantation and their uniqueness was verified by ligation-mediated (LM) PCR. We functionally compared the single stem cell clones between each other by re-mixing them in a single mouse for in vivo assays; analysis was performed one-by-one for each clone by flow cytometry where they could be distinguished from each other using their unique color codes. Clones showed clear differences in proliferation rate with faster and slower growing clones, independently whether 2 or 5 clones were mixed. When mice harboring clone mixtures were treated with conventional chemotherapy, clonal composition changed markedly and resistant clones overgrew sensitive clones indicating selective clonal responses and clonal advantage. A clone which showed especially slow growth in vivo was most resistant to in vivo treatment with Glucocorticoids. The slowly proliferating, Glucocorticoid-resistant clone had lost the additional X chromosome, which was present in all other clones and the bulk and showed a distinct DNA-methylation pattern analyzed by 450K arrays (illumina). In exome analysis, the clone showed 11 unique alterations including a single nucleotide variant in the oncogene USP6. We are currently performing RNA sequencing analysis to assess the differential gene expression in the clones. Taken together, genetic multicolor marking PDX ALL cells in the individualized xenograft mouse model allowed generating viable single cell clones for genetic functional characterization in vivo. Within the heterogeneous tumor bulk, an subclone existed which showed slow tumor growth and drug resistance which was associated with distinct genetic characteristics. Our studies allow the challenging functional characterization of subclones in vivo in order to develop efficient novel treatment approaches to eliminate aggressive stem cell clones in ALL.
AU  - Finkenzeller, C.
AU  - Carlet, M.
AU  - Vosberg, S.*
AU  - Greif, P.A.*
AU  - Jeremias, I.
C1  - 48026
C2  - 39860
CY  - Washington
TI  - Functional diversity of single stem cell clones in patients' acute lymphoblastic leukemia growing in mice: An adverse subclone with distinct DNA-methylation pattern, slow growth in vivo and drug resistance.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Introduction: Mantle cell lymphoma (MCL) is characterized by t(11;14) resulting in a constitutive cyclin D1 overexpression. The cyclin D1-CDK4/6 complex inactivates Rb through phosphorylation, leading to G1/S-phase transition. Therefore, inhibition of CDK4/6 is an efficient and rational approach to overcome cell cycle dysregulation in MCL. We evaluated the efficiency of the novel CDK4/6 inhibitor abemaciclib in various MCL cell lines and in primary MCL cells in combination with cytarabine (AraC) and ibrutinib.
      
      	Material & Methods: MCL cell lines (Granta 519, JeKo-1, Maver-1, Mino) and primary MCL cells were exposed to abemaciclib alone and combined with AraC or ibrutinib. Cells were pretreated with abemaciclib and exposed to AraC or ibrutinib with or without consecutive wash-out of the CDK4/6 inhibitor. Proliferation and viability were measured by tryptan blue staining and Cell Titer Glo assay. Flow cytometry was used for cell-cycle (PI-staining) and apoptosis analysis (Annexin V PE/7AAD-staining). Western Blot analysis showed protein expression and phosphorylation status of various downstream proteins.
      
      	Results: Abemaciclib inhibited cell proliferation by induction of early G1-arrest. Western Blot analysis revealed reduced phosphorylation of Rb on serine 795 without changes in CDK 4 and cyclin D1 expression, in line with reversible cell cycle arrest. IC50-values of sensitive cell lines (JeKo-1, Maver-1, Mino) were <30 nM after 72 h. We observed an almost complete and reversible G1-arrest in all sensitive cell lines by FACS analysis (JeKo-1: G1-phase +51,7 %; S/G2-phase -51,7 % at 31,25 nM after 24 h; G1-phase +35,4 %; S/G2-phase -34,8 % after 72 h), whereas cell viability was not reduced.
      
      	Wash-out of abemaciclib after 24 h resulted in synchronized S-phase entry in all sensitive cell lines (e.g. Mino: G1-phase -20,4 %; S-phase +30,5 %). The sequential combination of abemaciclib followed by AraC showed strong synergy in Mino cells (CI=0,22 for 31,25 nM abemaciclib and 3,33 µM cytarabine). In contrast, simultaneous exposure to abemaciclib had a protective effect against AraC treatment in all sensitive cell lines, due to an ongoing G1-arrest (Mino: CI=-0,19 for 31,25 nM abemaciclib and 3,33 µM AraC).
      
      	In primary MCL cells, 31,25 nM of abemaciclib had no impact on cell death. Moreover, no sensitization to AraC was observed as all cells were resting in G0-phase.
      
      	The combination of abemaciclib induced G1 arrest and ibrutinib had additive or synergistic effects in sensitive cell lines (JeKo-1, Mino and Maver).
      
      	Conclusion: The novel CDK4/6 inhibitor abemaciclib causes reversible G1 cell cycle arrest without loss of viability at low nanomolar doses. Rationale drug combinations exploiting the sequential effect may achieve major benefits, but drug interactions are complex: Pretreatment with abemaciclib sensitizes MCL cell line cells to AraC whereas simultaneous application protects them from AraC treatment. Further analyses explore the interaction with other targeted approaches (inhibitors of the B-cell receptor pathway) to better understand the underlying molecular mechanisms.
AU  - Fischer, L.
AU  - Schnaiter, A.
AU  - Freysoldt, B.
AU  - Irger, M.
AU  - Zimmermann, Y.
AU  - Hutter, G.
AU  - Hiddemann, W.
AU  - Dreyling, M.
C1  - 48024
C2  - 39862
CY  - Washington
TI  - The novel CDK4/6-inhibitor abemaciclib induces early G1-arrest in MCL cell lines, sensitizes cells to cytarabine treatment and is additive with ibrutinib.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Introduction: Mantle cell lymphoma (MCL) comprises about 6% of all non-Hodgkin's lymphoma with a median survival of 3-5 years. Constitutional activation of the mTOR/AKT pathway has been identified in the majority of cases (Rudelius, Blood 2006). The pro-viral insertion in murine (PIM) lymphoma proteins are serine/threonine kinases which play a critical role in cell survival as well as proliferation and identifies a high risk patient cohort with MCL (Hsi, Leuk Lymphoma 2008). In this study we evaluated the efficiency and mode of action of a dual PIM/PI3K (IBL-202) and a triple PIM/PI3K/mTOR inhibitor (IBL-301) in MCL cell lines and primary cells.
      
      	Methods: MCL cell lines (Granta 519, Jeko-1, Rec-1 and Mino), as well as primary MCL cells were exposed to the combined PIM-kinase/PI3K (IBL-202) and the PIM-kinase/PI3K/mTOR Inhibitor (IBL-301). Cell proliferation (trypan blue staining), cell apoptosis (Annexin V PE/7-AAD staining) and cell cycle (FACS) were investigated. Protein expression and phosphorylation status of different downstream proteins (Akt, GSK-3β, 4EBP1) as well as markers of apoptosis (PARP, Caspase 9) were analysed after 1h, 4h, 8h and 24h. Cell viability was assessed by CellTiter-Glo® assay after 48h.
      
      	Results: Both inhibitors led to G1 arrest. At 500 nM, the triple inhibitor IBL-301 (19,8%) is in average slightly more efficacious than the dual-inhibitor IBL-202 (13,5%). Accordingly, IBL-301 had a much higher impact on cell proliferation than IBL-202 in all tested MCL cell lines (reduction by 48 - 93% vs 22 - 87%), possibly due to its mTOR inhibitory potential, although it may be also a more potent inhibitor of PIM and PI3K kinases. In addition, treatment with IBL-202 and IBL-301 induced cell apoptosis in Jeko-1, Rec-1 and Mino. Again, rate of apoptosis by IBL-301 was much higher (e.g. JEKO: 56% vs 13%) and could be achieved at lower concentrations in comparison to IBL-202. The differential impact on apoptosis could be confirmed based on PARP and Caspase 9 cleavage, which was higher after treatment with IBL-301 after 24h. In Jeko-1, Granta-519 and Mino both agents led to de-phosphorylation of Akt. Interestingly, this effect was more prominent in IBL-301 treated cell lines, supporting the mode of action via the PI3K-AKT pathway of both inhibitors. De-phosphorylation of GSK-3β was observed in all tested MCL cell lines with both inhibitors already during the first hour of exposure and was reversible thereafter.
      
      	Primary MCL cells of 2 patients were treated with 62.5 nM IBL-202, 31.25 nM IBL-301 and single inhibitors of PIM (2.5 µM AZD1208), PI3K (1.25 µM idelalisib) and mTOR (5 nM temsirolimus). Viability after 48h was reduced by about 70% following IBL-301 exposure compared to 39% in IBL-202 treated samples. Both combined inhibitors were more potent than any of the single inhibitors. IBL-301 and IBL-202 decreased viability in a similar way as the combination of AZD1208, idelalisib +/- temsirolimus. Normal lymphocytes tolerated both inhibitors in various concentrations (62,5 - 500 nM).
      
      	Conclusions: Triple inhibition of PIM kinases, PI3K and mTOR is very efficient in MCL cell lines as well as in primary MCL cells, exceeding dual inhibition of PIM kinases and PI3K. Thus, cotargeting PIM kinases, PI3K and mTOR may be a promising novel approach for clinical development in MCL.
AU  - Freysoldt, B.
AU  - Schnaiter, A.
AU  - Fischer, L.
AU  - O'Neill, M.*
AU  - Zimmermann, Y.
AU  - Hutter, G.
AU  - Hiddemann, W.
AU  - Dreyling, M.
C1  - 48028
C2  - 39858
CY  - Washington
TI  - Cotargeting of PIM, PI3K and Mtor in Mantle Cell Lymphoma (MCL).
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Garz, A.-K.*
AU  - Habringer, S.*
AU  - Wolf, S.*
AU  - Vick, B.
AU  - Weickert, M.-T.*
AU  - Jeremias, I.
AU  - Spiekermann, K.*
AU  - Peschel, C.*
AU  - Oostendorp, R.A.J.*
AU  - Keller, U.*
AU  - Goetze, K.S.*
C1  - 48068
C2  - 39888
CY  - Washington
TI  - Azacitidine in combination with the selective FLT3 kinase inhibitor crenolanib effectively disrupts stromal protection of CD34+Leukemia-Initiating Cells (LIC) in FLT3-ITD+ Acute Myeloid Leukemia (AML).
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Human monocytes are subdivided into classical, intermediate and non-classical subsets but there is no unequivocal strategy to dissect the latter two cell types. We show herein that the cell surface marker 6-sulfo LacNAc (slan) can define slan-positive CD14+CD16++ non-classical monocytes and slan-negative CD14++CD16+ intermediate monocytes. Gene expression profiling confirms that slan-negative intermediate monocytes show highest expression levels of MHC class II genes, while a differential ubiquitin signature is a novel feature of the slan approach. In unsupervised hierarchical clustering the slan-positive non-classical monocytes cluster with monocytes and are clearly distinct from CD1c-positive dendritic cells. In clinical studies we show a selective increase of the slan-positive intermediate monocytes to >100 cells/μl in patients with sarcoidosis and a 5-fold depletion of the slan-positive monocytes in patients with hereditary diffuse leukodystrophy with axonal spheroids (HDLS), which is caused by macrophage colony stimulating factor (M-CSF) receptor mutations. These data demonstrate that the slan-definition of CD16-positive monocyte subsets is informative in molecular studies and in clinical settings.
AU  - Hofer, T.P.
AU  - Zawada, A.M.*
AU  - Frankenberger, M.
AU  - Skokann, K.
AU  - Satzl, A.A.*
AU  - Gesierich, W.*
AU  - Schuberth, M.*
AU  - Levin, J.*
AU  - Danek, A.*
AU  - Rotter, B.*
AU  - Heine, G.H.*
AU  - Ziegler-Heitbrock, L.
C1  - 46937
C2  - 39454
SP  - 2601-2610
TI  - Slan-defined subsets of CD16-positive monocytes: Impact of granulomatous inflammation and M-CSF-receptor mutation.
JO  - Blood
VL  - 126
IS  - 24
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Fibrinogen, coagulation factor VII (FVII), factor VIII (FVIII), and its carrier von Willebrand factor (vWF) play key roles in hemostasis. Previously identified common variants explain only a small fraction of the trait heritabilities and additional variation may be explained by associations with rarer variants with larger effects. The aim of this study was to identify low-frequency (minor allele frequency [MAF] ≥0.01 and <0.05) and rare (MAF <0.01) variants that influence plasma concentrations of these 4 hemostatic factors by meta-analyzing exome chip data from up to 76,000 participants of 4 ancestries. We identified 12 novel associations of low-frequency (n=2) and rare (n=10) variants across the fibrinogen, FVII, FVIII, and vWF traits that were independent of previously identified associations. Novel loci were found within previously reported genes and had effect sizes much larger than and independent of previously identified common variants. In addition, associations at KCNT1, HID1, and KATNB1 identify new candidate genes related to hemostasis for follow-up replication and functional genomic analysis. Newly identified low-frequency and rare-variant associations accounted for modest amounts of trait variance and therefore are unlikely to increase predicted trait heritability but provide new information to understanding individual variation in hemostasis pathways.
AU  - Huffman, J.E.*
AU  - de Vries, P.S.*
AU  - Morrison, A.C.*
AU  - Sabater-Lleal, M.*
AU  - Kacprowski, T.*
AU  - Auer, P.L.*
AU  - Brody, J.A.*
AU  - Chasman, D.I.*
AU  - Chen, M.H.*
AU  - Guo, X.*
AU  - Lin, L.-A.*
AU  - Marioni, R.E.*
AU  - Müller-Nurasyid, M.
AU  - Yanek, L.R.*
AU  - Pankratz, N.*
AU  - Grove, M.L.*
AU  - de Maat, M.P.M.*
AU  - Cushman, M.*
AU  - Wiggins, K.L.*
AU  - Qi, L.*
AU  - Sennblad, B.*
AU  - Harris, S.E.*
AU  - Polasek, O.*
AU  - Riess, H.*
AU  - Rivadeneira, F.*
AU  - Rose, L.M.*
AU  - Goel, A.*
AU  - Taylor, K.D.*
AU  - Teumer, A.*
AU  - Uitterlinden, A.G.*
AU  - Vaidya, D.*
AU  - Yao, J.*
AU  - Tang, W.*
AU  - Levy, D.*
AU  - Waldenberger, M.
AU  - Becker, D.M.*
AU  - Folsom, A.R.*
AU  - Giulianini, F.*
AU  - Greinacher, A.*
AU  - Hofman, A.*
AU  - Huang, C.C.*
AU  - Kooperberg, C.*
AU  - Silveira, A.*
AU  - Starr, J.M.*
AU  - Strauch, K.
AU  - Strawbridge, R.J.*
AU  - Wright, A.F.*
AU  - McKnight, B.*
AU  - Franco, O.H.*
AU  - Zakai, N.*
AU  - Mathias, R.A.*
AU  - Psaty, B.M.*
AU  - Ridker, P.M.*
AU  - Tofler, G.H.*
AU  - Völker, U.*
AU  - Watkins, H.*
AU  - Fornage, M.*
AU  - Hamsten, A.*
AU  - Deary, I.J.*
AU  - Boerwinkle, E.*
AU  - Koenig, W.*
AU  - Rotter, J.I.*
AU  - Hayward, C.*
AU  - Dehghan, A.*
AU  - Reiner, A.P.*
AU  - O'Donnell, C.J.*
AU  - Smith, N.L.*
C1  - 45423
C2  - 37331
SP  - e19-e29
TI  - Rare and low-frequency variants and their association with plasma levels of fibrinogen, FVII, FVIII, and vWF.
JO  - Blood
VL  - 126
IS  - 11
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Background: In acute myeloid leukemia (AML), detection of minimal residual disease (MRD) by flow cytometry is an adverse prognostic factor besides pre-treatment risk classifications, including cytogenetic and molecular aberrations. High dimensional multiparameter flow cytometry (MPFC) offers improved sensitivity and specificity, however manual analysis is increasingly challenging. In this study, we explore the value of the recently proposed viSNE algorithm to quantify MRD levels in patients with AML achieving complete remission (CR) after intensive induction chemotherapy.
      
      	Methods: Bone marrow samples from patients with AML (excluding patients with acute promyelocytic leukemia) were analyzed by 8-10 MPFC using a NAVIOS flow cytometer (Beckman Coulter, Brea, CA, USA). Only patients achieving a CR or CR with incomplete blood count recovery (CRi) post-induction were included in this analysis. Manual gating of MRD flow data was performed as described previously (Köhnke et al., Leukemia 2014) using a cutoff for MRD positivity of 0.1%. The viSNE algorithm was performed as described previously (Amir et al., Nat. Biotech. 2013) and MRD positivity was defined as the presence of a distinct cluster of >100 cells which consisted of >90% patient cells. Kaplan-Meier estimator and log-rank test as well as Cox's proportional hazards regression model were used to analyze survival data.
      
      	Results: Post-induction flow cytometry and clinical data of 38 patients with AML achieving CR (n=34) or CRi (n=4) were available for analysis (median age 53 years; de-novo AML n=32, tAML n=1, sAML n=5). Most patients belonged to the intermediate cytogenetic risk group (MRC favorable n=5, intermediate n=22, adverse n=11).
      
      	19/38 patients were MRD positive post-induction by manual gating. 12/19 patients deemed MRD positive relapsed, whereas 3/19 patients deemed MRD negative relapsed. Therefore, MRD positivity by manual gating correlated with reduced relapse free survival (median RFS for MRD positive patients: 7.5 months vs. median not reached for MRD negative patients, log-rank test p=0.017). For overall survival (OS), no significant impact of MRD positivity could be detected so far (p=0.3), however follow-up was short (median follow-up 9.3 months). MRD positivity by manual gating remained an independent risk factor for RFS (HR 4.8, p=0.021) when compared to genetic risk and age.
      
      	MRD positivity by viSNE clustering was seen in 19/38 patients. 10/19 patients deemed MRD positive by viSNE relapsed, while 5/19 deemed MRD negative by viSNE experienced a relapse. This resulted in a trend towards shorter RFS for MRD positivity by viSNE (median RFS 9.9 months vs. 19.0 months for MRD negative patients, p=0.185). Among the patients deemed MRD positive by viSNE who did not relapse, i.e. false-positive patients, follow-up was very short (<3.5 months) in 4/9 cases and final judgment whether these patients are truly negative should be withheld. However, differentiation between healthy monocytes and potential MRD cells by viSNE seems to be especially challenging and warrants optimization.
      
      	Nevertheless, within the group of patients deemed MRD negative by manual gating, viSNE was able to detect all patients who experienced a subsequent relapse. To maximize sensitivity for MRD detection, we therefore combined the results of both methods and defined MRD positivity as positivity by manual gating and/or viSNE clustering. Using this combined strategy, 28/38 patients were defined as MRD positive and 10/38 as MRD negative. Within the MRD positive group, 15/28 patients relapsed, whereas none of the patients in the MRD negative group relapsed (Figure 1, median RFS for MRD positive patients: 7.7 months, log-rank test p=0.028). Of note, within the MRD negative group only 2 patients (both with intermediate genetic risk) went on to receive a allogeneic stem cell transplantation whereas 8 remained in remission with chemotherapy alone.
      
      	Conclusion: In summary, viSNE clustering used in combination with manual gating improves the sensitivity of MRD detection by flow cytometry. Importantly, viSNE is able to detect MRD positivity within the manual MRD negative patient population. This is the first report of MRD detection in AML using this method. However, improvements in the algorithm as well as further studies are needed to validate the prognostic value of viSNE clustering in AML.
AU  - Köhnke, T.
AU  - Rechkemmer, S.*
AU  - Bücklein, V.L.
AU  - Pfannes, K.*
AU  - Fiegl, M.*
AU  - Spiekermann, K.*
AU  - Hiddemann, W.*
AU  - Subklewe, M.
C1  - 48031
C2  - 39855
CY  - Washington
TI  - Improved detection of minimal residual disease by flow cytometry in AML by combining manual gating and visne clustering.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - In our previous work, we showed elimination of primary acute myeloid leukemia (AML) cells by CD33/CD3 BiTE® antibody construct (AMG 330) mediated cytotoxicity (Krupka et al, Blood 2014). The goal of the present study was to identify innate and adaptive resistance mechanisms to AMG 330 mediated lysis of AML cells. Immune checkpoint molecules have been shown to be a highly relevant escape mechanism of malignant cells to evade innate and adaptive immunity. Previously, it was shown that AML cells upregulate the expression of inhibitory ligands in response to proinflammatory cytokines (Krönig et al, 2014). As AMG 330 mediated T-cell activation induces high levels of the proinflammatory cytokines IFNγ and TNFα, we assessed the constitutive and inducible expression profile of different immune checkpoint molecules on AML cell lines and primary AML cells, including PD-L1, HVEM, ILT3 and SLAMF7 by flow cytometry. No constitutive expression was observed for PD-L1 at time of primary diagnosis in 83.7% of the cases (103/123). In contrast, constitutive expression of HVEM and ILT3 was detected in 73.7% (42/57) and 91.9% (68/74) of patient samples, respectively. Adaptive resistance was evaluated by incubating AML cell lines and primary AML samples with IFNγ and TNFα. We observed an upregulation of PD-L1 and SLAMF7 on AML cell lines and on primary AML patient samples whereas HVEM and ILT3 did not show a significant change in expression level. To test the functional relevance of the immune checkpoint molecules upon AMG 330 mediated lysis, we used an ex vivo long term culture system that enabled us to analyse the dynamic process of receptor-ligand interaction over time. Blockade of the PD-1/PD-L1 interaction resulted in a significantly increase in AMG 330 mediated lysis of primary AML cells (n=9, p=0.03). Currently, blockade of the inducible molecule SLAMF7 in AMG 330 mediated cytotoxicity is being tested. Blocking of HVEM or ILT3 did not result in a significant increase in T cell activation and concomitant lysis of AML cells suggesting a less relevant role of HVEM and ILT3 in resistance to AMG 330 mediated cytotoxicity. The latter might also be influenced by the cytokine microenvironment which favours immune resistance of AML cells.  Using a bead based multiplex assay we screened the bone marrow (BM) plasma from 16 AML patients and 3 healthy donors (HD) for the presence of 33 cytokines. The cytokine profile differed between AML patients and healthy donors (HDs). The plasma levels of IL-8, IP-10 and CXCL-16 were higher in the AML samples compared to those of HDs (p=0.0041, 0.0248 and 0.0289, respectively). In contrast, EGF, FLT3-ligand, RANTES and IL-4 were significantly lower in AML samples compared to HDs (p=0.0227, 0.0145, 0.0041 and 0.0041, respectively). However, we did observe a high inter-patient variability of cytokine composition in AML. To explore the functional relevance of the BM plasma on AMG 330 mediated cytotoxicity, cocultures of AML cell lines and HD T cells were set up using different sources of plasma including fetal calf serum (FCS) and patient derived BM plasma. Interestingly, AMG 330 mediated cytotoxicity was significantly reduced using patient derived BM plasma (n=5) compared to cultures containing FCS (n=4) (mean % lysis FCS 97.4 vs PT 70.6). This was accompanied by a considerable impairment in T-cell proliferation (mean % proliferation FKS 44.7% vs PT 26.6%). Currently, we are investigating which soluble factors are responsible for the immunosuppressive effects and if we can increase lysis efficacy and T-cell proliferation through specific blocking of them. In summary we have identified possible resistance mechanisms of AML cells to AMG 330 mediated cytotoxicity. Dynamic receptor-ligand interactions between target and effector cells as well as soluble factors contribute to AMG 330 mediated lysis of primary AML cells. We hypothesize that AMG 330 mediated cytotoxicity can be augmented through combinatorial approaches including PD-1 blockade. The significance of our findings will first be validated in an in vivo mouse model and prospectively translated into human studies.
AU  - Krupka, C.
AU  - Köhnke, T.
AU  - Kufer, P.*
AU  - Kischel, R.*
AU  - Lichtenegger, F.S.
AU  - Brauneck, F.
AU  - Rohling, S.*
AU  - Altmann, T.
AU  - Hiddemann, W.*
AU  - Subklewe, M.
C1  - 48030
C2  - 39856
CY  - Washington
TI  - Identifying immune resistance mechanisms to CD33/CD3 BiTE® antibody construct (AMG 330) mediated cytotoxicity.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - To expedite the translation of biologic discoveries into novel therapeutics, there is a pressing need for panels of in vivo models that capture the molecular complexity of human disease. While traditional cell lines and genetically engineered mouse models are useful tools, they are insufficient to assess the broad diversity of human tumors within a context that recapitulates in situ biology. Patient-derived xenografts (PDXs), generated by transplanting primary human tumor cells into immune-deficient NOD.Cg-Prkdcscid/Il2rgtm1Wjl/SzJ (NSG) mice, surmount some of the limitations of these traditional platforms and have been increasingly utilized as tools for preclinical investigation. However, the infrastructure required to generate, bank, and characterize PDX models limits their availability to only a few investigators. To address this issue, we established a repository of PDX models of leukemia and lymphoma, which we have named the Public Repository of Xenografts (PRoXe). At the time of this writing, PRoXe contains 213 independent lines that have been passaged through mice once (P0), 123 of which have been repassaged in a second generation (P1) or further repassaged. The repository encompasses AML, B- and T-ALL, and B- and T-cell non-Hodgkin lymphoma (NHL) across a range of cytogenetic- and molecularly-defined subtypes (Table 1). PRoXe is extensively annotated with patient-level information, including demographics, phase of treatment, prior therapies, tumor immunophenotye, cytogenetics, and molecular diagnostics. PDX lines available for distribution are characterized by immunophenotyping, whole transcriptome sequencing (RNAseq), and targeted exon sequencing of ~300 genes. To confirm fidelity of engrafted tumors to their corresponding clinical samples, lymphomas were morphologically assessed in P0 mice by H&E and, when pathologic adjudication was required, by immunohistochemistry. Xenografted leukemias were compared to their original tumors immunophenotypically. Unsupervised hierarchical clustering was performed on 132 of these lines based on transcriptome sequencing data and demonstrated 94% concordance between classification of the PDX lines by RNA expression and by the annotated clinical-pathologic diagnoses. Discordant cases highlighted unusual variants, such as B-ALL with aberrant expression of myeloid markers and a follicular lymphoma that underwent blastic transformation in the mouse. Multiple lines have been luciferized and confirmed to home to bone marrow, spleen, and liver. Existing lines from PRoXe have already been shared with more than ten academic laboratories and multiple industrial partners. All of the data referenced here are freely available through a customized web-based search application at http://proxe.org, and lines can be requested for in vitro or in vivo experiments. We are actively expanding the size of PRoXe to allow for large pre-clinical studies that are powered to detect differences across genetically defined subsets. Thus, we are happy to host additional lines from outside investigators on PRoXe and thereby expand the availability of these valuable reagents. Finally, we have made the source code for PRoXe (in R Shiny) open-access, so that other investigators can establish their own portals.
AU  - Murakami, M.A.*
AU  - Christodoulou, A.N.*
AU  - Christie, A.L.*
AU  - DeSouza, T.*
AU  - Louissaint, A.*
AU  - Vojinovic, U.*
AU  - Koch, R.*
AU  - Li, L.S.*
AU  - Kallgren, S.P.*
AU  - Rao, P.*
AU  - Koester, J.*
AU  - Vadhi, R.*
AU  - Duberow, E.*
AU  - Morgan, E.A.*
AU  - Wang, H.*
AU  - Ahmed, S.S.*
AU  - Majewski, K.L.*
AU  - Konopleva, M.*
AU  - Tamburini, J.*
AU  - Gutierrez, A.*
AU  - Kelliher, M.*
AU  - Etchin, J.*
AU  - Jeremias, I.
AU  - Weng, A.P.*
AU  - Kung, A.L.*
AU  - Lane, A.A.*
AU  - Carnache-Ottou, F.*
AU  - Izraeli, S.*
AU  - Jacobsen, E.S.*
AU  - Galinsky, I.*
AU  - Stone, R.M.*
AU  - Harris, M.H.*
AU  - Dorfman, D.M.*
AU  - Aster, J.C.*
AU  - Long, H.*
AU  - Silverman, L.B.*
AU  - Weinstock, D.M.*
C1  - 48027
C2  - 39859
CY  - Washington
TI  - Proxe: A public repository of xenografts to facilitate studies of biology and expedite preclinical drug development in leukemia and lymphoma.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Reiter, K.
AU  - Polzer, H.
AU  - Krupka, C.
AU  - Vick, B.
AU  - Maiser, A.*
AU  - Subklewe, M.
AU  - Jeremias, I.
AU  - Leonhardt, H.*
AU  - Spiekermann, K.
AU  - Greif, P.A.
C1  - 48067
C2  - 39889
CY  - Washington
TI  - Tyrosin kinase inhibition restores the membrane localization of FLT3-ITD.
JO  - Blood
VL  - 126
IS  - 23
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The treatment of non-Hodgkin lymphomas has benefited enormously from the introduction of monoclonal antibody-based therapies. However, the efficacy of these treatments varies with lymphoma subtypes and typically decreases with subsequent relapses. Here, we report on antigen-armed antibodies (AgAbs) as a potential treatment for B-cell lymphoma. AgAbs include antigens from ubiquitous pathogens, such as Epstein-Barr virus (EBV), that persist in their host and elicit strong lifelong T cell responses. They act as vectors by introducing antigen directly into tumor cells to induce an antigen-specific CD4+ T cell response against these cells. We have fused antibodies targeting human B-cell surface receptors (CD19-22) to immunodominant T cell antigens from EBV proteins, including EBNA1, EBNA3B and EBNA3C. Exposure of EBV-transformed B-cells and of BL cells to AgAbs led to antigen presentation, T cell recognition and target cell killing. The efficiency of AgAb action paralleled the abundance of the targeted molecules on lymphoma cells as well as their HLA class II expression levels. AgAbs can also induce the activation and proliferation of EBV-specific memory CD4+ T cells ex vivo. These studies show the potential of AgAbs as an effective therapeutic strategy against B-cell lymphomas.
AU  - Yu, X.*
AU  - Ilecka, M.*
AU  - Bartlett, E.J.*
AU  - Schneidt, V.*
AU  - Bhat, R.*
AU  - Mautner, J.
AU  - Feederle, R.
AU  - Delecluse, H.J.*
C1  - 43382
C2  - 36361
CY  - Washington
SP  - 1601-1610
TI  - Antigen-armed antibodies targeting B lymphoma cells effectively activate antigen-specific CD4+ T cells.
JO  - Blood
VL  - 125
IS  - 10
PB  - Amer Soc Hematology
PY  - 2015
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Bigalke, I.*
AU  - Honnashagen, K.*
AU  - Lundby, M.*
AU  - Kasten, J.
AU  - Inderberg, E.M.S.*
AU  - Skoge, L.*
AU  - Saboe-Larssen, S.*
AU  - Schendel, D.J.*
AU  - Kvalheim, G.*
C1  - 43916
C2  - 36643
CY  - Washington
TI  - Vaccination with a new generation of fast dendritic cells transfected with mRNA from hTERT, survivin and autologous tumor mount strong immune responses and prolong survival.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Systemic bacterial infection induces a hematopoietic response program termed 'emergency granulopoiesis' that is characterized by increased de novo bone marrow (BM) neutrophil production. How loss of local immune control and bacterial dissemination is sensed and subsequently translated into the switch from steady-state to emergency granulopoiesis is, however, unknown. Using tissue-specific myeloid differentiation primary response gene 88 (Myd88)-deficient mice and in vivo lipopolysaccharide (LPS) administration to model severe bacterial infection, we here show that endothelial cells (ECs) but not hematopoietic cells, hepatocytes, pericytes or bone marrow stromal cells, are essential cells for this process. Indeed, ECs from multiple tissues including BM express high levels of Tlr4 and Myd88, and are the primary source of granulocyte colony-stimulating factor (G-CSF), the key granulopoietic cytokine, following LPS challenge or infection with E. coli. Endothelial cell-intrinsic MYD88-signaling and subsequent G-CSF production by ECs is required for myeloid progenitor lineage skewing towards granulocyte-macrophage progenitors (GMPs), increased colony-forming unit granulocyte (CFU-G) activity in BM, and accelerated BM neutrophil generation following LPS stimulation. Thus, ECs catalyze the detection of systemic infection into demand-adapted granulopoiesis.
AU  - Boettcher, S.*
AU  - Gerosa, R.*
AU  - Radpour, R.*
AU  - Bauer, J.
AU  - Ampenberger, F.*
AU  - Heikenwälder, M.
AU  - Kopf, M.*
AU  - Manz, M.G.*
C1  - 31731
C2  - 34735
CY  - Washington
SP  - 1393-1403
TI  - Endothelial cells translate pathogen signals into G-CSF-driven emergency granulopoiesis.
JO  - Blood
VL  - 124
IS  - 9
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Bruns, H.*
AU  - Buettner, M.*
AU  - Fabri, M.*
AU  - Mougiakakos, D.*
AU  - Bittenbring, J.T.*
AU  - Pasemann, S.*
AU  - Neumann, F.J.*
AU  - Maurberger, A.*
AU  - Kempkes, B.
AU  - Mackensen, A.*
AU  - Gerbitz, A.*
C1  - 44028
C2  - 36707
CY  - Washington
TI  - Vitamin D-dependent induction of cathelicidin in human macrophages results in cytotoxicity against high grade B-cell lymphoma.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Calvo-Vidal, N.*
AU  - Patel, J.*
AU  - Krumsiek, J.*
AU  - Dupont, T.*
AU  - Goldstein, R.S.*
AU  - Yang, S.N.*
AU  - Melnick, A.*
AU  - Chiosis, G.*
AU  - Cerchietti, L.*
C1  - 44029
C2  - 36711
CY  - Washington
TI  - Hsp90 at the hub of metabolic homeostasis in malignant B cells.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Dutta, S.
AU  - Krause, A.*
AU  - Vosberg, S.
AU  - Herold, T.*
AU  - Ksienzyk, B.*
AU  - Quintanilla-Fend, L.*
AU  - Tizazu, B.
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Greif, P.A.
AU  - Schneider, M.*
AU  - Dahlhof, M.*
AU  - Spiekermann, K.
AU  - Zimber-Strobl, U.
AU  - Wolf, E.*
AU  - Bohlander, S.K.*
C1  - 43913
C2  - 36646
CY  - Washington
TI  - Analysis of the tissue-specific expression requirements and identification of cooperating mutations for leukemogenesis in an inducible CALM/AF10 knock-in mouse model.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Hartmann, L.
AU  - Vosberg, S.
AU  - Metzeler, K.H.
AU  - Schumacher, D.*
AU  - Pastore, F.
AU  - Bräundl, K.
AU  - Zellmeier, E.*
AU  - Ksienzyk, B.*
AU  - Konstandin, N.P.*
AU  - Schneider, S.*
AU  - Graf, A.*
AU  - Blum, H.*
AU  - Neumann, M.*
AU  - Baldus, C.D.*
AU  - Bohlander, S.K.*
AU  - Wolf, S.*
AU  - Wiemann, S.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
AU  - Greif, P.A.
C1  - 44033
C2  - 36705
CY  - Washington
TI  - Genetic evolution of Cytogenetically Normal Acute Myeloid Leukemia (CN-AML) during therapy and relapse: An exome sequencing study of 47 cases.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Isolated trisomy 13 (AML+13) is a rare chromosomal abnormality in acute myeloid leukemia (AML), and its prognostic relevance is poorly characterized. We analyzed the clinical course of 34 AML+13 patients enrolled in the German AMLCG-1999 and SAL trials and studied their biological characteristics by exome sequencing, targeted sequencing of candidate genes and gene expression profiling. Relapse-free (RFS) and overall survival (OS) of AML+13 patients were inferior compared to other ELN Intermediate-II patients (n=855) (median RFS, 7.8 vs 14.1 months, p=0.006; median OS 9.3 vs. 14.8 months, p=0.004). Besides the known high frequency of RUNX1 mutations (75%), we identified mutations in spliceosome components in 88%, including SRSF2 codon 95 mutations in 81%, of AML+13 patients. Moreover, recurring mutations were detected in ASXL1 (44%) and BCOR (25%). Two patients carried mutations in CEBPZ, suggesting that CEBPZ is a novel recurrently mutated gene in AML. Gene expression analysis revealed a homogenous expression profile including upregulation of FOXO1 and FLT3 and downregulation of SPRY2. This is the most comprehensive clinical and biological characterization of AML+13 to date, and reveals a striking clustering of lesions in a few genes, defining AML+13 as a genetically homogenous leukemia subgroup with alterations in a few critical cellular pathways. These studies were registered at clinicaltrials.gov, identifiers: AMLCG-1999: NCT00266136; AML96: NCT00180115; AML2003: NCT00180102; and AML60+: NCT00893373.
AU  - Herold, T.
AU  - Metzeler, K.H.
AU  - Vosberg, S.
AU  - Hartmann, L.
AU  - Röllig, C.*
AU  - Stölzel, F.*
AU  - Schneider, S.*
AU  - Hubmann, M.
AU  - Zellmeier, E.*
AU  - Ksienzyk, B.*
AU  - Jurinovic, V.*
AU  - Pasalic, Z.*
AU  - Kakadia, P.M.*
AU  - Dufour, A.*
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Sauerland, M.C.*
AU  - Büchner, T.*
AU  - Berdel, W.E.*
AU  - Woermann, B.J.*
AU  - Bornhäuser, M.*
AU  - Ehninger, G.*
AU  - Mansmann, U.*
AU  - Hiddemann, W.
AU  - Bohlander, S.K.*
AU  - Spiekermann, K.
AU  - Greif, P.A.
C1  - 31609
C2  - 34632
CY  - Washington
SP  - 1304-1311
TI  - Isolated trisomy 13 defines a genetically homogenous AML subgroup with high frequency of mutations in spliceosome genes and poor prognosis.
JO  - Blood
VL  - 124
IS  - 8
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Hoellein, A.*
AU  - Schoeffmann, S.*
AU  - Mohammad, F.*
AU  - Cleveland, J.L.*
AU  - Gloeckner, C.J.
AU  - Ueffing, M.
AU  - Rudelius, M.*
AU  - Peschel, C.*
AU  - Keller, U.*
C1  - 43915
C2  - 36644
CY  - Washington
TI  - Myc commands an aurora kinase - sumoylation circuit required for B cell lymphoma growth and survival.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Hutter, G.
AU  - Zimmermann, Y.*
AU  - Zoellner, A.-K.*
AU  - Irrgang, P.*
AU  - Weigert, O.*
AU  - Hiddemann, W.*
AU  - Dreyling, M.*
C1  - 44031
C2  - 36709
CY  - Washington
TI  - Combination of PI3K and PDPK1 inhibitors is highly effective in mantle cell lymphoma.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Jeremias, I.
AU  - Vick, B.
AU  - Rothenberg, M.*
AU  - Sandhöfer, N.*
AU  - Carlet, M.
AU  - Krupka, C.*
AU  - Trumpp, A.*
AU  - Corbacioglu, S.*
AU  - Ebinger, M.*
AU  - Andre, M.C.*
AU  - Hiddemann, W.*
AU  - Schneider, S.*
AU  - Subklewe, M.*
AU  - Metzeler, K.H.*
AU  - Spiekermann, K.*
C1  - 43918
C2  - 36641
CY  - Washington
TI  - Bioluminescence in vivo imaging improves the model of individual patients' AML cells growing in mice for sensitive and reliable preclinical treatment trials on various genetic subgroups.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Jost, P.J.*
AU  - Yabal, M.*
AU  - Adler, H.
AU  - Knies, N.*
AU  - Gross, C.*
AU  - Damgaard, R.B.*
AU  - Kanegane, H.*
AU  - Peschel, C.*
AU  - Strasser, A.*
AU  - Gross, O.*
AU  - Ruland, J.*
AU  - Gyrd-Hansen, M.*
C1  - 43914
C2  - 36645
CY  - Washington
TI  - A mouse model for XLP-2 disease uncovers a critical function for IL-1beta and TNF in driving hyper-inflammation.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Antibody-based immunotherapy represents a promising strategy to target and eliminate chemoresistant leukemic cells. Here, we evaluated the CD33/CD3-bispecific BiTE® antibody (AMG 330) for its suitability as therapeutic agent in AML. We first assessed CD33 expression levels by flow cytometry and found expression in >99% of patient samples (n=621). CD33 was highest expressed in AMLs with NPM1 mutations (p<0.001) and lower in AMLs with complex karyotypes and t(8;21) translocations (p<0.001). Furthermore, leukemic stem cells within the CD34(+)/CD38(-) compartment displayed CD33 at higher levels than healthy donor stem cells (p=0.047). In MS-5 feeder cell-based long-term cultures that supported the growth of primary AML blasts for up to 36 days, AMG 330 efficiently recruited and expanded residual CD3(+)/CD45RA(-)/CCR7(+) memory T-cells within the patient sample. Even at low effector to target ratios, the recruited T-cells lysed autologous blasts completely in the majority of samples and substantially in the remaining samples in a time- dependent manner. This study provides the first correlation of CD33 expression levels with AML genotype in a comprehensive analysis of adult patients. Targeting CD33 ex-vivo using AMG 330 in primary AML samples led to T-cell recruitment and expansion and remarkable antibody-mediated cytotoxicity suggesting efficient therapeutic potential in-vivo.
AU  - Krupka, C.
AU  - Kufer, P.*
AU  - Kischel, R.*
AU  - Zugmaier, G.*
AU  - Bögeholz, J.
AU  - Köhnke, T.
AU  - Lichtenegger, F.S.
AU  - Schneider, S.*
AU  - Metzeler, K.H.*
AU  - Fiegl, M.*
AU  - Spiekermann, K.
AU  - Baeuerle, P.A.*
AU  - Hiddemann, W.
AU  - Riethmüller, G.*
AU  - Subklewe, M.
C1  - 28602
C2  - 33474
SP  - 356-365
TI  - CD33 target validation and sustained depletion of AML blasts in long-term cultures by the bispecific T-cell-engaging antibody AMG 330.
JO  - Blood
VL  - 123
IS  - 3
PB  - Amer. Soc. Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Li, L.S.*
AU  - Wu, S.*
AU  - Kopp, N.*
AU  - Montero, J.*
AU  - Ryan, J.*
AU  - Chapuy, B.*
AU  - van der Zwet, J.C.*
AU  - Layer, J.*
AU  - Weigert, O.
AU  - Christie, A.L.*
AU  - Christodoulou, A.N.*
AU  - Liu, H.*
AU  - Yoda, A.*
AU  - Hofmann, F.*
AU  - Baffert, F.*
AU  - Vangrevelinghe, E.*
AU  - Gaul, C.*
AU  - Radimerski, T.*
AU  - Letai, A.G.*
AU  - Weinstock, D.M.*
C1  - 44030
C2  - 36710
CY  - Washington
TI  - Type II JAK2 inhibitor NVP-CHZ868 is active in vivo against JAK2-dependent B-Cell Acute Lymphoblastic Leukemias (B-ALLs).
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT) are threatened by potentially lethal viral manifestations like cytomegalovirus (CMV) reactivation. Because the success of today's virostatic treatment is limited by side effects and resistance development, adoptive transfer of virus-specific memory T cells derived from the stem cell donor has been proposed as an alternative therapeutic strategy. In this context, dose minimization of adoptively transferred T cells might be warranted for the avoidance of graft-versus-host disease (GVHD), in particular in prophylactic settings after T-cell-depleting allo-HSCT protocols. To establish a lower limit for successful adoptive T-cell therapy, we conducted low-dose CD8(+) T-cell transfers in the well-established murine Listeria monocytogenes (L.m.) infection model. Major histocompatibility complex-Streptamer-enriched antigen-specific CD62L(hi) but not CD62L(io) CD8(+) memory T cells proliferated, differentiated, and protected against L.m. infections after prophylactic application. Even progenies derived from a single CD62L(hi) L.m.-specific CD8(+) T cell could be protective against bacterial challenge. In analogy, low-dose transfers of Streptamer-enriched human CMV-specific CD8(+) T cells into allo-HSCT recipients led to strong pathogen-specific T-cell expansion in a compassionate-use setting. In summary, low-dose adoptive T-cell transfer (ACT) could be a promising strategy, particularly for prophylactic treatment of infectious complications after allo-HSCT.
AU  - Stemberger, C.*
AU  - Graef, P.*
AU  - Odendahl, M.*
AU  - Albrecht, J.
AU  - Doessinger, G.*
AU  - Anderl, F.*
AU  - Buchholz, V.R.*
AU  - Gasteiger, G.
AU  - Schiemann, M.
AU  - Grigoleit, G.U.*
AU  - Schuster, F.R.*
AU  - Borkhardt, A.*
AU  - Versluys, B.*
AU  - Tonn, T.*
AU  - Seifried, E.*
AU  - Einsele, H.*
AU  - Germeroth, L.*
AU  - Busch, D.H.
AU  - Neuenhahn, M.
C1  - 32648
C2  - 35219
CY  - Washington
SP  - 628-637
TI  - Lowest numbers of primary CD8+ T cells can reconstitute protective immunity upon adoptive immunotherapy.
JO  - Blood
VL  - 124
IS  - 4
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The outcome of patients with acute myeloid leukemia who are older than 60 years has remained poor because of unfavorable disease characteristics and patient-related factors. The randomized German-Austrian AML Study Group 06-04 protocol was designed on the basis of in vitro synergistic effects of valproic acid (VPA) and all-trans retinoic acid with chemotherapy. Between 2004 and 2006, 186 patients were randomly assigned to receive 2 induction cycles with idarubicin, cytarabine, and all-trans retinoic acid either with VPA or without (STANDARD). In all patients, consolidation therapy was intended. Complete remission rates after induction tended to be lower in VPA compared with STANDARD (40% vs 52%; P 5 .14) as a result of a higher early death rate (26% vs 14%; P 5 .06). The main toxicities attributed to VPA were delayed hematologic recovery and grade 3/4 infections, observed predominantly during the second induction cycle. After restricting VPA to the first induction cycle and reducing the dose of idarubicin, these toxicities dropped to rates observed in STANDARD. After a median follow-up time of 84 months, event-free and overall survivalwere not different between the 2 groups (P 5 .95 andP5.57, respectively). However, relapse-free-survival was significantly superior in VPAcompared with STANDARD(24.4% vs 6.4% at 5 years; P 5 .02). Explorative subset analyses revealed that AML with mutated Nucleophosmin 1 (NPM1) may particularly benefit from VPA. This trial was registered at www.clinicaltrials.gov as #NCT00151255.
AU  - Tassara, M.*
AU  - Döhner, K.*
AU  - Brossart, P.*
AU  - Held, G.*
AU  - Götze, K.S.*
AU  - Horst, H.A.*
AU  - Ringhoffer, M.*
AU  - Köhne, C.H.*
AU  - Kremers, S.*
AU  - Raghavachar, A.N.*
AU  - Wulf, G.G.*
AU  - Kirchen, H.G.*
AU  - Nachbaur, D.M.*
AU  - Derigs, H.G.*
AU  - Wattad, M.*
AU  - Koller, E.A.*
AU  - Brugger, W.*
AU  - Matzdorff, A.C.*
AU  - Greil, R.F.*
AU  - Heil, G.*
AU  - Paschka, P.*
AU  - Gaidzik, V.I.*
AU  - Göttlicher, M.
AU  - Döhner, H.*
AU  - Schlenk, R.F.*
C1  - 31763
C2  - 34747
SP  - 4027-4036
TI  - Valproic acid in combination with all-trans retinoic acid and intensive therapy for acute myeloid leukemia in older patients.
JO  - Blood
VL  - 123
IS  - 26
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - von der Heide, E.K.*
AU  - Vosberg, S.
AU  - Neumann, M.*
AU  - Mochmann, L.H.*
AU  - James, A.R.*
AU  - Tanchez, J.O.*
AU  - Schlee, C.*
AU  - Isaakidis, K.*
AU  - Luther, M.*
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Hofmann, W.*
AU  - Greif, P.A.
AU  - Baldus, C.D.*
C1  - 44032
C2  - 36706
CY  - Washington
TI  - Molecular alterations in bone marrow mesenchymal stroma cells of AML patients.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Vosberg, S.
AU  - Herold, T.*
AU  - Metzeler, K.H.*
AU  - Schneider, S.*
AU  - Ksienzyk, B.*
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Spiekermann, K.
AU  - Bohlander, S.K.*
AU  - Mansmann, U.*
AU  - Greif, P.A.
C1  - 43917
C2  - 36642
CY  - Washington
TI  - Copy Number Alteration (CNA) analysis in targeted sequencing data from Acute Myeloid Leukemia (AML) patients with chromosome 9q deletion.
JO  - Blood
VL  - 124
IS  - 21
PB  - Amer Soc Hematology
PY  - 2014
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - In this issue of Blood, Nogai et al have identified the atypical nuclear factor–κB (NF-κB) modulator IκBζ as a key factor that drives oncogenic NF-κB activity in an activated B-cell subtype of diffuse large B-cell lymphoma (ABC DLBCL).
AU  - Krappmann, D.
C1  - 27788
C2  - 32813
SP  - 2146-2147
TI  - Shaping oncogenic NF-κB activity in the nucleus.
JO  - Blood
VL  - 122
IS  - 13
PB  - Amer. Soc. Hematology
PY  - 2013
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Early T-cell precursor (ETP) acute lymphoblastic leukemia (ALL) is a high-risk subgroup of T-lineage ALL characterized by specific stem cell and myeloid features. In adult ETP-ALL, no comprehensive studies on the genetic background have been performed to elucidate molecular lesions of this distinct subgroup. We performed whole-exome sequencing of 5 paired ETP-ALL samples. In addition to mutations in genes known to be involved in leukemogenesis (ETV6, NOTCH1, JAK1, and NF1), we identified novel recurrent mutations in FAT1 (25%), FAT3 (20%), DNM2 (35%), and genes associated with epigenetic regulation (MLL2, BMI1, and DNMT3A). Importantly, we verified the high rate of DNMT3A mutations (16%) in a larger cohort of adult patients with ETP-ALL (10/68). Mutations in epigenetic regulators support clinical trials, including epigenetic-orientated therapies, for this high-risk subgroup. Interestingly, more than 60% of adult patients with ETP-ALL harbor at least a single genetic lesion in DNMT3A, FLT3, or NOTCH1 that may allow use of targeted therapies.
AU  - Neumann, M.*
AU  - Heesch, S.*
AU  - Schlee, C.*
AU  - Schwartz, S.*
AU  - Gökbuget, N.*
AU  - Hoelzer, D.*
AU  - Konstandin, N.P.
AU  - Ksienzyk, B.
AU  - Vosberg, S.
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Raff, T.*
AU  - Brüggemann, M.*
AU  - Hofmann, W.-K.*
AU  - Hecht, J.*
AU  - Bohlander, S.K.
AU  - Greif, P.A.
AU  - Baldus, C.D.*
C1  - 27029
C2  - 32492
SP  - 4749-4752
TI  - Whole-exome sequencing in adult ETP-ALL reveals a high rate of DNMT3A mutations.
JO  - Blood
VL  - 121
IS  - 23
PB  - Amer. Soc. Hematology
PY  - 2013
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The t(8;21) and inv(16)/t(16;16) rearrangements affecting the core-binding factors, RUNX1 and CBFB, respectively, are found in 15-20% of adult de novo AML cases and are associated with a favourable prognosis. Since the expression of the fusion genes CBFB/MYH11 or RUNX1/RUNX1T1 alone is not sufficient to cause leukemia, we performed exome sequencing of an AML sample with an inv(16) to identify mutations, which may collaborate with the CBFB/MYH11 fusion during leukemogenesis. We discovered an N676K mutation in the ATP-binding domain (TKD1) of the fms-related tyrosine kinase 3 (FLT3) gene. In a cohort of 84 de novo AML patients with a CBFB/MYH11 rearrangement and in 36 patients with a RUNX1/RUNX1T1 rearrangement, the FLT3 N676K mutation was identified in 5 and 1 patients, respectively (5/84, 6%; 1/36, 3%). The FLT3-N676K mutant alone leads to factor-independent growth in Ba/F3 cells and, together with a concurrent FLT3-ITD, confers resistance to the FLT3 PTK inhibitors PKC412 and AC220. Gene expression analysis of AML patients with CBFB/MYH11 rearrangement and FLT3 N676K mutation showed a trend towards a specific expression profile. Ours is the first report of recurring FLT3 N676 mutations in CBF leukemias and suggests a defined subgroup of CBF leukemias. Registered at www.clinicaltrials.gov: AMLCG-1999 trial (NCT00266136).
AU  - Opatz, S.*
AU  - Polzer, H.
AU  - Herold, T.
AU  - Konstandin, N.P.*
AU  - Ksienzyk, B.*
AU  - Zellmeier, E.*
AU  - Vosberg, S.
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Hopfner, K.P.*
AU  - Kakadia, P.M.*
AU  - Schneider, S.*
AU  - Dufour, A.*
AU  - Braess, J.*
AU  - Sauerland, M.C.*
AU  - Berdel, W.E.*
AU  - Büchner, T.*
AU  - Woermann, B.J.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
AU  - Bohlander, S.K.
AU  - Greif, P.A.
C1  - 26380
C2  - 32203
SP  - 1761-1769
TI  - Exome sequencing identifies recurring FLT3 N676K mutations in core-binding factor leukemia.
JO  - Blood
VL  - 122
IS  - 10
PB  - Amer. Soc. Hematology
PY  - 2013
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Cancer stem cells represent the most important target cells for antitumor therapy. TRAIL (TNF-related apoptosis inducing ligand) is a potential anticancer agent that induces apoptosis in a wide variety of tumor cells, but its ability to target cancer stem cells is currently unknown. Here we investigated whether TRAIL targets leukemia-initiating cells. Limiting dilution transplantation assays were performed on xenografts from pediatric patients with precursor B-cell acute lymphoblastic leukemia (pre-B ALL) in NSG mice. In vitro treatment of xenograft cells with TRAIL significantly reduced and delayed their engraftment and procrastinated animal death from leukemia. Systemic TRAIL treatment of mice injected with patient-derived pre-B ALL xenograft cells abrogated leukemia in 3 of 5 mice in 1 sample. In conclusion, our data suggest that TRAIL targets leukemia-initiating cells derived from pre-B ALL xenografts in vitro and in vivo, and hence constitutes an attractive candidate drug for treatment of ALL.
AU  - Alves, C.C.
AU  - Terziyska, N.
AU  - Grunert, M.
AU  - Gündisch, S.
AU  - Graubner, U.*
AU  - Quintanilla-Martinez, L.*
AU  - Jeremias, I.
C1  - 7610
C2  - 29868
SP  - 4224-4227
TI  - Leukemia-initiating cells of patient-derived acute lymphoblastic leukemia xenografts are sensitive toward TRAIL.
JO  - Blood
VL  - 119
IS  - 18
PB  - Amer. Soc. Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Cytogenetically normal acute myeloid leukemia (CN-AML) with biallelic CEBPA gene mutations (biCEPBA) represents a distinct disease entity with a favorable clinical outcome. So far, it is not known whether other genetic alterations cooperate with biCEBPA mutations during leukemogenesis. To identify additional mutations, we performed whole exome sequencing of 5 biCEBPA patients and detected somatic GATA2 zinc finger 1 (ZF1) mutations in 2 of 5 cases. Both GATA2 and CEBPA are transcription factors crucial for hematopoietic development. Inherited or acquired mutations in both genes have been associated with leukemogenesis. Further mutational screening detected novel GATA2 ZF1 mutations in 13 of 33 biCEBPA-positive CN-AML patients (13/33, 39.4%). No GATA2 mutations were found in 38 CN-AML patients with a monoallelic CEBPA mutation and in 89 CN-AML patients with wild-type CEBPA status. The presence of additional GATA2 mutations (n = 10) did not significantly influence the clinical outcome of 26 biCEBPA-positive patients. In reporter gene assays, all tested GATA2 ZF1 mutants showed reduced capacity to enhance CEBPA-mediated activation of transcription, suggesting that the GATA2 ZF1 mutations may collaborate with biCEPBA mutations to deregulate target genes during malignant transformation. We thus provide evidence for a genetically distinct subgroup of CN-AML. The German AML cooperative group trials 1999 and 2008 are registered with the identifiers NCT00266136 and NCT01382147 at www.clinicaltrials.gov. (Blood. 2012; 120(2): 395-403)
AU  - Greif, P.A.
AU  - Dufour, A.*
AU  - Konstandin, N.P.
AU  - Ksienzyk, B.*
AU  - Zellmeier, E.
AU  - Tizazu, B.
AU  - Sturm, J.
AU  - Benthaus, T.*
AU  - Herold, T.*
AU  - Yaghmaie, M.
AU  - Dorge, P.*
AU  - Hopfner, K.P.*
AU  - Hauser, A.*
AU  - Graf, A.*
AU  - Krebs, S.*
AU  - Blum, H.*
AU  - Kakadia, P.M.*
AU  - Schneider, S.*
AU  - Hoster, E.*
AU  - Schneider, F.*
AU  - Stanulla, M.*
AU  - Braess, J.*
AU  - Sauerland, M.C.*
AU  - Berdel, W.E.*
AU  - Büchner, T.*
AU  - Woermann, B.J.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
AU  - Bohlander, S.K.
C1  - 8441
C2  - 30148
SP  - 395-403
TI  - GATA2 zinc finger 1 mutations associated with biallelic CEBPA mutations define a unique genetic entity of acute myeloid leukemia.
JO  - Blood
VL  - 120
IS  - 2
PB  - Amer. Soc. Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - We conducted a genome-wide association study to identify novel associations between genetic variants and circulating plasminogen activator inhibitor-1 (PAI-1) concentration, and examined functional implications of variants and genes that were discovered. A discovery meta-analysis was performed in 19 599 subjects, followed by replication analysis of genome-wide significant (P < 5 × 10(-8)) single nucleotide polymorphisms (SNPs) in 10 796 independent samples. We further examined associations with type 2 diabetes and coronary artery disease, assessed the functional significance of the SNPs for gene expression in human tissues, and conducted RNA-silencing experiments for one novel association. We confirmed the association of the 4G/5G proxy SNP rs2227631 in the promoter region of SERPINE1 (7q22.1) and discovered genome-wide significant associations at 3 additional loci: chromosome 7q22.1 close to SERPINE1 (rs6976053, discovery P = 3.4 × 10(-10)); chromosome 11p15.2 within ARNTL (rs6486122, discovery P = 3.0 × 10(-8)); and chromosome 3p25.2 within PPARG (rs11128603, discovery P = 2.9 × 10(-8)). Replication was achieved for the 7q22.1 and 11p15.2 loci. There was nominal association with type 2 diabetes and coronary artery disease at ARNTL (P < .05). Functional studies identified MUC3 as a candidate gene for the second association signal on 7q22.1. In summary, SNPs in SERPINE1 and ARNTL and an SNP associated with the expression of MUC3 were robustly associated with circulating levels of PAI-1.
AU  - Huang, J.*
AU  - Sabater-Lleal, M.*
AU  - Asselbergs, F.W.*
AU  - Tregouet, D.*
AU  - Shin, S.Y.*
AU  - Ding, J.*
AU  - Baumert, J.J.
AU  - Oudot-Mellakh, T.*
AU  - Folkersen, L.*
AU  - Johnson, A.D.*
AU  - Smith, N.L.*
AU  - Williams, S.M.*
AU  - Ikram, M.A.*
AU  - Kleber, M.E.*
AU  - Becker, D.M.*
AU  - Truong, V.*
AU  - Mychaleckyj, J.C.*
AU  - Tang, W.*
AU  - Yang, Q.*
AU  - Sennblad, B.*
AU  - Moore, J.H.*
AU  - Williams, F.M.*
AU  - Dehghan, A.*
AU  - Silbernagel, G.*
AU  - Schrijvers, E.M.*
AU  - Smith, S.*
AU  - Karakas, M.*
AU  - Tofler, G.H.*
AU  - Silveira, A.*
AU  - Navis, G.J.*
AU  - Lohman, K.*
AU  - Chen, M.H.*
AU  - Peters, A.
AU  - Goel, A.*
AU  - Hopewell, J.C.*
AU  - Chambers, J.C.*
AU  - Saleheen, D.*
AU  - Lundmark, P.*
AU  - Psaty, B.M.*
AU  - Strawbridge, R.J.*
AU  - Boehm, B.O.*
AU  - Carter, A.M.*
AU  - Meisinger, C.
AU  - Peden, J.F.*
AU  - Bis, J.C.*
AU  - McKnight, B.*
AU  - Ohrvik, J.*
AU  - Taylor, K.*
AU  - Franzosi, M.G.*
AU  - Seedorf, U.*
AU  - Collins, R.*
AU  - Franco-Cereceda, A.*
AU  - Syvanen, A.C.*
AU  - Goodall, A.H.*
AU  - Yanek, L.R.*
AU  - Cushman, M.*
AU  - Müller-Nurasyid, M.
AU  - Folsom, A.R.*
AU  - Basu, S.*
AU  - Matijevic, N.*
AU  - van Gilst, W.H.*
AU  - Kooner, J.S.*
AU  - Hofman, A.*
AU  - Danesh, J.*
AU  - Clarke, R.*
AU  - Meigs, J.B.*
AU  - DIAGRAM Consortium (Petersen, A.-K.
AU  - Gieger, C.
AU  - Klopp, N.
AU  - Thorand, B.
AU  - Grallert, H.
AU  - Wichmann, H.-E.
AU  - Illig, T.
AU  - Meitinger, T.
AU  - Huth, C.)
AU  - Kathiresan, S.*
AU  - Reilly, M.P.*
AU  - CARDIoGRAM Consortium (Peters, A.
AU  - Klopp, N.
AU  - Wichmann, H.-E.
AU  - Meisinger, C.
AU  - Meitinger, T.
AU  - Döring, A.
AU  - Illig, T.)
AU  - Harris, T.B.*
AU  - Winkelmann, B.R.*
AU  - Grant, P.J.*
AU  - Hillege, H.L.*
AU  - Watkins, H.*
AU  - C4D Consortium (*)
AU  - Spector, T.D.*
AU  - Becker, L.C.*
AU  - Tracy, R.P.*
AU  - Marz, W.*
AU  - Uitterlinden, A.G.*
AU  - Eriksson, P.*
AU  - Cambien, F.*
AU  - CARDIOGENICS Consortium (*)
AU  - Morange, P.E.*
AU  - Koenig, W.*
AU  - Soranzo, N.*
AU  - van der Harst, P.*
AU  - Liu, Y.*
AU  - O'Donnell, C.J.*
AU  - Hamsten, A.*
C1  - 11565
C2  - 30690
SP  - 4873-4881
TI  - Genome-wide association study for circulating levels of PAI-1 provides novel insights into its regulation.
JO  - Blood
VL  - 120
IS  - 24
PB  - American Society of Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Lymphoid enhancer-binding factor-1 (LEF1) is a key transcription factor of Wnt signaling. We recently showed that aberrant LEF1 expression induces acute myeloid leukemia (AML) in mice, and found high LEF1 expression in a subset of cytogenetically normal AML (CN-AML) patients. Whether LEF1 expression associates with clinical and molecular patient characteristics and treatment outcomes remained unknown. We therefore studied LEF1 expression in 210 adults with CN-AML treated on German AML Cooperative Group trials using microarrays. High LEF1 expression (LEF1(high)) associated with significantly better relapse-free survival (RFS; P < .001), overall survival (OS; P < .001), and event-free survival (EFS; P < .001). In multivariable analyses adjusting for established prognosticators, LEF1(high) status remained associated with prolonged RFS (P = .007), OS (P = .01), and EFS (P = .003). In an independent validation cohort of 196 CN-AML patients provided by the German-Austrian AML Study Group, LEF1(high) patients had significantly longer OS (P = .02) and EFS (P = .04). We validated the prognostic relevance of LEF1 expression by quantitative PCR, thereby providing a clinically applicable platform to incorporate this marker into future risk-stratification systems for CN-AML. Gene-expression profiling and immunophenotyping revealed up-regulation of lymphopoiesis-related genes and lymphoid cell-surface antigens in LEF1(high) patients. In summary, we provide evidence that high LEF1 expression is a novel favorable prognostic marker in CN-AML. (Blood. 2012; 120(10):2118-2126)
AU  - Metzeler, K.H.*
AU  - Heilmeier, B.*
AU  - Edmaier, K.E.*
AU  - Rawat, V.P.S.*
AU  - Dufour, A.*
AU  - Döhner, K.*
AU  - Feuring-Buske, M.*
AU  - Braess, J.
AU  - Spiekermann, K.
AU  - Büchner, T.*
AU  - Sauerland, M.C.*
AU  - Döhner, H.*
AU  - Hiddemann, W.
AU  - Bohlander, S.K.
AU  - Schlenk, R.F.*
AU  - Bullinger, L.*
AU  - Buske, C.*
C1  - 10629
C2  - 30317
SP  - 2118-2126
TI  - High expression of Lymphoid Enhancer-binding Factor-1 (LEF1) is a novel favorable prognostic factor in cytogenetically normal acute myeloid leukemia.
JO  - Blood
VL  - 120
IS  - 10
PB  - Amer. Soc. Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The ontogenic relationship between the common dendritic cell (DC) progenitor (CDP), the committed conventional DC precursor (pre-cDC), and cDC subpopulations in lymphoid and nonlymphoid tissues has been largely unraveled. In contrast, the sequential steps of plasmacytoid DC (pDC) development are less defined, and it is unknown at which developmental stage and location final commitment to the pDC lineage occurs. Here we show that CCR9(-) pDCs from murine BM which enter the circulation and peripheral tissues have a common DC precursor function in vivo in the steady state, in contrast to CCR9(-) pDCs which are terminally differentiated. On adoptive transfer, the fate of CCR9(-) pDC-like precursors is governed by the tissues they enter. In the BM and liver, most transferred CCR9(-) pDC-like precursors differentiate into CCR9(-) pDCs, whereas in peripheral lymphoid organs, lung, and intestine, they additionally give rise to cDCs. CCR9(-) pDC-like precursors which are distinct from pre-cDCs can be generated from the CDP. Thus, CCR9(-) pDC-like cells are novel CDP-derived circulating DC precursors with pDC and cDC potential. Their final differentiation into functionally distinct pDCs and cDCs depends on tissue-specific factors allowing adaptation to local requirements under homeostatic conditions. (Blood. 2012; 119(25): 6063-6071)
AU  - Schlitzer, A.*
AU  - Heiseke, A.F.*
AU  - Einwächter, H.*
AU  - Reindl, W.*
AU  - Schiemann, M.
AU  - Manta, C.P.*
AU  - See, P.*
AU  - Niess, J.H.*
AU  - Suter, T.*
AU  - Ginhoux, F.*
AU  - Krug, A.B.*
C1  - 8504
C2  - 30276
SP  - 6063-6071
TI  - Tissue-specific differentiation of a circulating CCR9- pDC-like common dendritic cell precursor.
JO  - Blood
VL  - 119
IS  - 25
PB  - Amer. Soc. Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The impact of a FLT3-internal tandem duplication (FLT3ITD) on prognosis of patients with acute myeloid leukemia (AML) is dependent on the ratio of mutated to wild-type allele. In 648 normal karyotype (NK) AML patients, we found a significant independent effect of the quantitative FLT3ITD mRNA level-measured as (FLT3ITD/wtFLT3)/(FLT3ITD/wtFLT3 + 1)-on outcome. Moreover, this effect was clearly seen in 329 patients with a mutated NPM1 gene (NPM1(+)), but not in 319 patients without a NPM1 mutation (wtNPM1). In a multivariate Cox regression model, the quantitative FLT3ITD mRNA level showed an independent prognostic impact on overall survival (OS) and relapse-free survival (RFS) only in the NPM1(+) subgroup (OS: hazard ratio, 5.9; [95% confidence interval [CI]: 3.1-11.2]; RFS: hazard ratio, 7.5 [95% CI: 3.4-16.5]). The FLT3ITD mRNA level contributes to relapse risk stratification and might help to guide postremission therapy in NPM1-mutated AML.
AU  - Schneider, F.*
AU  - Hoster, E.*
AU  - Unterhalt, M.*
AU  - Schneider, S.*
AU  - Dufour, A.*
AU  - Benthaus, T.*
AU  - Mellert, G.*
AU  - Zellmeier, E.*
AU  - Kakadia, P.M.*
AU  - Bohlander, S.K.
AU  - Feuring-Buske, M.*
AU  - Buske, C.*
AU  - Braess, J.*
AU  - Heinecke, A.*
AU  - Sauerland, M.C.*
AU  - Berdel, W.E.*
AU  - Büchner, T.*
AU  - Wörmann, B.J.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
C1  - 7686
C2  - 29890
SP  - 4383-4386
TI  - The FLT3ITD mRNA level has a high prognostic impact in NPM1 mutated, but not in NPM1 unmutated, AML with a normal karyotype.
JO  - Blood
VL  - 119
IS  - 19
PB  - Amer. Soc. Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The hyaluronan-mediated motility receptor (HMMR/Rhamm) is overexpressed in numerous tumor types, including acute lymphoid leukemia and acute myeloid leukemia (AML). Several studies have reported the existence of T-cell responses directed against HMMR in AML patients that are linked to better clinical outcome. Therefore, we explored the use of HMMR-specific TCRs for transgenic expression in lymphocytes and their in vivo impact on HMMR(+) solid tumors and disseminated leukemia. We obtained TCRs via an in vitro priming approach in combination with CD137-mediated enrichment. Recipient lymphocytes expressing transgenic TCR revealed the specific tumor recognition pattern seen with the original T cells. Adoptive transfer experiments using a humanized xenograft mouse model resulted in significantly retarded solid tumor outgrowth, which was enhanced using IL-15-conditioned, TCR-transgenic effector memory cells. These cells also showed an increased potency to retard the outgrowth of disseminated AML, and this was further improved using CD8-enriched effector memory cells. To define a safe clinical setting for HMMR-TCR gene therapy, we analyzed transgenic T-cell recognition of hematopoietic stem cells (HSCs) and found on-target killing of HLA-A2(+) HSCs. Our findings clearly limit the use of HMMR-TCR therapy to MHC- mismatched HSC transplantation, in which HLA-A2 differences can be used to restrict recognition to patient HSCs and leukemia.
AU  - Spranger, S.
AU  - Jeremias, I.
AU  - Wilde, S.
AU  - Leisegang, M.*
AU  - Stärck, L.*
AU  - Mosetter, B.
AU  - Uckert, W.*
AU  - Heemskerk, M.H.*
AU  - Schendel, D.J.
AU  - Frankenberger, B.
C1  - 7922
C2  - 29920
SP  - 3440-3449
TI  - TCR-transgenic lymphocytes specific for HMMR/Rhamm limit tumor outgrowth in vivo.
JO  - Blood
VL  - 119
IS  - 15
PB  - American Society of Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Natural killer (NK) cells play a critical role in early host defense to infected and transformed cells. Here we show that mice deficient in Eri1, a conserved 3'-to-5' exoribonuclease that represses RNA interference, have a cell-intrinsic defect in NK cell development and maturation. Eri1(-/-) NK cells displayed delayed acquisition of Ly49 receptors in the bone marrow and a selective reduction in Ly49D and Ly49H activating receptors in the periphery. Eri1 was required for immune-mediated control of mouse cytomegalovirus (MCMV) infection. Ly49H(+) NK cells deficient in Eri1 failed to expand efficiently during MCMV infection, and virus-specific responses were also diminished among Eri1(-/-) T cells. We identified miRNAs as the major endogenous small RNA target of Eri1 in mouse lymphocytes. Both NK and T cells deficient in Eri1 displayed a global, sequence-independent increase in miRNA abundance. Ectopic Eri1 expression rescued defective miRNA expression in mature Eri1(-/-) T cells. Thus, mouse Eri1 regulates miRNA homeostasis in lymphocytes and is required for normal NK cell development and anti-viral immunity.
AU  - Thomas, M.F.*
AU  - Abdul-Wajid, S.*
AU  - Panduro, M.*
AU  - Babiarz, J.E.*
AU  - Rajaram, M.*
AU  - Woodruff, P.*
AU  - Lanier, L.L.*
AU  - Heissmeyer, V.
AU  - Ansel, K.M.*
C1  - 7552
C2  - 29785
SP  - 130-142
TI  - Eri1 regulates microRNA homeostasis and mouse lymphocyte development and anti-viral function.
JO  - Blood
VL  - 120
IS  - 1
PB  - American Society of Hematology
PY  - 2012
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - We used a retroviral integration screen to search for novel genes that regulate HSC function. One of the genes that conferred HSC dominance when overexpressed due to an adjacent retroviral insertion was Musashi 2 (Msi2), an RNA-binding protein that can act as a translational inhibitor. A gene-trap mouse model that inactivates the gene shows that Msi2 is more highly expressed in long-term (LT) and short-term (ST) HSCs, as well as in lymphoid myeloid primed progenitors (LMPPs), but much less in intermediate progenitors and mature cells. Mice lacking Msi2 are fully viable for up to a year or more, but exhibit severe defects in primitive precursors, most significantly a reduction in the number of ST-HSCs and LMPPs and a decrease in leukocyte numbers, effects that are exacerbated with age. Cell-cycle and gene-expression analyses suggest that the main hematopoietic defect in Msi2-defective mice is the decreased proliferation capacity of ST-HSCs and LMPPs. In addition, HSCs lacking Msi2 are severely impaired in competitive repopulation experiments, being overgrown by wild-type cells even when mutant cells were provided in excess. Our data indicate that Msi2 maintains the stem cell compartment mainly by regulating the proliferation of primitive progenitors downstream of LT-HSCs.
AU  - de Andrés-Aguayo, L.*
AU  - Varas, F.*
AU  - Kallin, E.M.*
AU  - Infante, J.F.*
AU  - Wurst, W.
AU  - Floß, T.
AU  - Graf, T.*
C1  - 6568
C2  - 28879
CY  - Washington, DC
SP  - 554-564
TI  - Musashi 2 is a regulator of the HSC compartment identified by a retroviral insertion screen and knockout mice.
JO  - Blood
VL  - 118
IS  - 3
PB  - American Society of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Application of anthracyclines and Vinca alkaloids on the same day represents a hallmark of polychemotherapy protocols for hematopoietic malignancies. Here we show, for the first time, that both drugs might act most efficiently if they are applied on different days. Proof-of-concept studies in 18 cell lines revealed that anthracyclines inhibited cell death by Vinca alkaloids in 83% of cell lines. Importantly, in a preclinical mouse model, doxorubicin reduced the anti-tumor effect of vincristine. Both drugs acted in a sequence-dependent manner and the strongest anti-tumor effect was obtained if both drugs were applied on different days. Most notably for clinical relevance, in 34% of 35 fresh primary childhood leukemia cells tested in vitro, doxorubicin reduced the anti-tumor effect of vincristine. As underlying mechanism, doxorubicin activated p53, p53 induced cell-cycle arrest, and cell-cycle arrest disabled inactivation of antiapoptotic Bcl-2 family members by vincristine; therefore, vincristine was unable to activate downstream apoptosis signaling. As molecular proof, antagonism was rescued by knockdown of p53, whereas knockdown of cyclin A inhibited vincristine-induced apoptosis. Our data suggest evaluating anthracyclines and Vinca alkaloids on different days in future trials. Selecting drug combinations based on mechanistic understanding represents a novel conceptional strategy for potent polychemotherapy protocols.
AU  - Ehrhardt, H.
AU  - Schrembs, D.
AU  - Moritz, C.
AU  - Wachter, F.
AU  - Haldar, S.*
AU  - Graubner, U.*
AU  - Nathrath, M.
AU  - Jeremias, I.
C1  - 6767
C2  - 29246
SP  - 6123-6131
TI  - Optimized anti-tumor effects of anthracyclines plus Vinca alkaloids using a novel, mechanism-based application schedule.
JO  - Blood
VL  - 118
IS  - 23
PB  - American Society of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - no abstract
AU  - Hammerschmidt, W.
C1  - 6485
C2  - 28762
CY  - Washington, USA
SP  - 1777-1778
TI  - What keeps the power on in lymphomas? (Comment on Vereide and Sudgen, page 1977)
JO  - Blood
VL  - 117
IS  - 6
PB  - American Society of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - B cell-specific gene ablation of Notch2 results in the loss of the Marginal Zone (MZ) B cell lineage. To analyze the effects of constitutive Notch2 signaling in B cells, we have generated a transgenic mouse strain allowing the conditional expression of a constitutively active, intracellular form of Notch2 (Notch2IC). Expression of Notch2IC at the earliest developmental stages of the B cell lineage completely abolished B cell generation and led to the development of ectopic T cells in the bone marrow, showing that Notch2IC is acting redundantly with Notch1IC in driving ectopic T cell differentiation. In B cells clearly committed to the B cell lineage induction of Notch2IC drove all cells towards the MZ B cell compartment at the expense of Follicular B cells. Notch2IC-expressing B cells reflected the phenotype of wild type MZ B cells with respect to their localization in the MZ, the expression of characteristic surface markers, their enhanced proliferation after stimulation and increased basal activity of Akt, Erk and Jnk. Notch2IC-driven MZ B cell generation in the spleen was achieved even in the absence of CD19. Our results implicate that a constitutive Notch2 signal in T1 B cells is sufficient to drive MZ B cell differentiation.
AU  - Hampel, F.
AU  - Ehrenberg, S.
AU  - Hojer, C.
AU  - Draeseke, A.
AU  - Marschall-Schröter, G.
AU  - Kühn, R.
AU  - Mack, B.*
AU  - Gires, O.
AU  - Vahl, C.J.*
AU  - Schmidt-Supprian, M.*
AU  - Strobl, L.J.
AU  - Zimber-Strobl, U.
C1  - 6627
C2  - 29006
CY  - Washington, DC, USA
SP  - 6321-6331
TI  - CD19-independent instruction of murine marginal zone B-cell development by constitutive Notch2 signaling.
JO  - Blood
VL  - 118
IS  - 14
PB  - American Society of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Pleiotrophin (Ptn) is strongly expressed by stromal cells which maintain HSCs. However, in vivo, Ptn deficiency does not alter steady-state hematopoiesis. However, knockdown of Ptn (Ptn(KD)) in stromal cells increases production of hematopoietic progenitors as well as HSC activity in cocultures, suggesting that Ptn may have a role in HSC activation. Indeed, transplantations of wild-type (Ptn(+/+)) HSCs into Ptn(-/-) mice show increased donor cell production in serial transplantations and dominant myeloid regeneration caused by Ptn-dependent regulation of HSC repopulation behavior. This regulation of Lin(-)Kit(+)Sca1(+) function is associated with increased proliferation and, on a molecular level, with up-regulated expression of cyclin D1 (Ccnd1) and C/EBPα (Cepba), but reduced of PPARγ. The known HSC regulator β-catenin is, however, not altered in the absence of Ptn. In conclusion, our results point to different Ptn-mediated regulatory mechanisms in normal hemostasis and in hematopoietic regeneration and in maintaining the balance of myeloid and lymphoid regeneration. Moreover, our results support the idea that microenvironmental Ptn regulates hematopoietic regeneration through β-catenin-independent regulation of Ccnd1 and Cebpa.
AU  - Istvanffy, R.*
AU  - Kröger, M.*
AU  - Eckl, C.*
AU  - Gitzelmann, S.*
AU  - Vilne, B.*
AU  - Bock, F.*
AU  - Graf, S.*
AU  - Schiemann, M.
AU  - Keller, U.B.*
AU  - Peschel, C.*
AU  - Oostendorp, R.A.*
C1  - 6926
C2  - 29434
SP  - 2712-2722
TI  - Stromal pleiotrophin regulates repopulation behavior of hematopoietic stem cells.
JO  - Blood
VL  - 118
IS  - 10
PB  - Amer. Soc. Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Adoptive transfer (AT) of T cells forced to express tumor-reactive T-cell receptor (TCR) genes is an attractive strategy to direct autologous T-cell immunity against tumor-associated antigens. However, clinical effectiveness has been hampered by limited in vivo persistence. We investigated whether the use of major histocompatibility complex-mismatched T cells would prolong the in vivo persistence of tumor-reactive TCR gene expressing T cells by continuous antigen-driven proliferation via the endogenous potentially alloreactive receptor. Donor-derived CD8(+) T cells engineered to express a TCR against a leukemia-associated antigen mediated strong graft-versus-leukemia (GVL) effects with reduced graft-versus-host disease (GVHD) severity when given early after transplantation. AT later after transplantation resulted in a complete loss of GVL. Loss of function was associated with reduced expansion of TCR-transduced T cells as assessed by CDR3 spectratyping analysis and PD-1 up-regulation on T cells in leukemia-bearing recipients. PD-L1 blockade in allogeneic transplant recipients largely restored the GVL efficacy without triggering GVHD, whereas no significant antileukemia effects of PD-L1 blockade were observed in syngeneic controls. These data suggest a clinical approach in which the AT of gene-modified allogeneic T cells early after transplantation can provide a potent GVL effect without GVHD, whereas later AT is effective only with concurrent PD-L1 blockade.
AU  - Koestner, W.*
AU  - Hapke, M.*
AU  - Herbst, J.*
AU  - Klein, C.*
AU  - Welte, K.*
AU  - Fruehauf, J.*
AU  - Flatley, A.
AU  - Vignali, D.A.*
AU  - Hardtke-Wolenski, M.*
AU  - Jaeckel, E.*
AU  - Blazar, B.R.*
AU  - Sauer, M.G.*
C1  - 4920
C2  - 28133
SP  - 1030-1041
TI  - PD-L1 blockade effectively restores strong graft-versus-leukemia effects without graft-versus-host disease after delayed adoptive transfer of T-cell receptor gene-engineered allogeneic CD8⁺ T cells.
JO  - Blood
VL  - 117
IS  - 3
PB  - American Society of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Fas Ligand (FasL) not only induces apoptosis in Fas receptor-bearing target cells, it is also able to transmit signals into the FasL-expressing cell via its intracellular domain (ICD). Recently, we described a Notch-like proteolytic processing of FasL that leads to the release of the FasL intracellular domain (ICD) into the cytoplasm and subsequent translocation into the nucleus where it may influence gene transcription. To study the molecular mechanism underlying such reverse FasL signaling in detail and to analyze its physiological importance in vivo, we established a knockout/knockin mouse model in which wildtype FasL was replaced with a deletion mutant lacking the ICD. Our results demonstrate that FasL ICD signaling impairs activation-induced proliferation in B and T cells by diminishing phosphorylation of PLCγ, PKC and ERK1/2. We also demonstrate that the FasL ICD interacts with the transcription factor Lymphoid-enhancer binding factor-1 (Lef-1) and inhibits Lef-1-dependent transcription. In vivo, plasma cell numbers, generation of germinal center B cells and, consequently, production of antigen-specific IgM antibodies in response to immunization with T cell-dependent (TD) or T cell-independent (TI) antigen are negatively affected in presence of the FasL ICD, suggesting that FasL reverse signaling participates in negative fine-tuning of certain immune responses.
AU  - Lückerath, K.*
AU  - Kirkin, V.*
AU  - Melzer, I.M.*
AU  - Thalheimer, F.B.*
AU  - Siele, D.*
AU  - Milani, W.*
AU  - Adler, T.*
AU  - Aguilar-Pimentel, J.A.*
AU  - Horsch, M.
AU  - Michel, G.
AU  - Beckers, J.
AU  - Busch, D.H.*
AU  - Ollert, M.*
AU  - Gailus-Durner, V.
AU  - Fuchs, H.
AU  - Hrabě de Angelis, M.
AU  - Staal, F.J.*
AU  - Rajalingam, K.*
AU  - Hueber, A.O.*
AU  - Strobl, L.J.
AU  - Zimber-Strobl, U.
AU  - Zörnig, M.*
C1  - 5368
C2  - 27961
SP  - 519-529
TI  - Immune modulation by Fas ligand reverse signaling: Lymphocyte proliferation is attenuated by the intracellular Fas ligand domain.
JO  - Blood
VL  - 117
IS  - 2
PB  - American Soc. of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Whereas the final differentiation of conventional dendritic cells (CDCs) from committed precursors occurs locally in secondary lymphoid or peripheral tissues, plasmacytoid dendritic cells (PDCs) are thought to fully develop in the bone marrow from common DC progenitors before migrating to the periphery. In our study, we define, for the first time, a subpopulation of CCR9(-) major histocompatibility complex class II(low) PDCs in murine bone marrow, which express E2-2 and are immediate precursors of CCR9(+) fully differentiated PDCs. However, CCR9(-) PDCs have the plasticity to acquire the phenotype and function of CD11b(+) CD8α(-) major histocompatibility complex class II(high) CDC-like cells under the influence of soluble factors produced by intestinal epithelial cells or recombinant GM-CSF. This deviation from the PDC lineage commitment is regulated on the level of transcription factors reflected by down-regulation of E2-2 and up-regulation of ID2, PU.1, and BATF3. Thus, CCR9(-) PDCs are immediate PDC precursors that can be reprogrammed to differentiate into CDC-like cells with higher antigen-presenting and cytokine-producing capacity under the influence of the local tissue microenvironment.
AU  - Schlitzer, A.*
AU  - Loschko, J.*
AU  - Mair, K.*
AU  - Vogelmann, R.*
AU  - Henkel, L.*
AU  - Einwächter, H.*
AU  - Schiemann, M.
AU  - Niess, J.H.*
AU  - Reindl, W.*
AU  - Krug, A.
C1  - 6388
C2  - 28598
SP  - 6562-6570
TI  - Identification of CCR9- murine plasmacytoid DC precursors with plasticity to differentiate into conventional DCs.
JO  - Blood
VL  - 117
IS  - 24
PB  - Amer Soc Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Splenic marginal zone (MZ) B cells are a lineage distinct from follicular and peritoneal B1 B cells. They are located next to the marginal sinus where blood is released. Here they pick up antigens and shuttle the load onto follicular dendritic cells inside the follicle. On activation, MZ B cells rapidly differentiate into plasmablasts secreting antibodies, thereby mediating humoral immune responses against blood-borne type 2 T-independent antigens. As Krüppel-like factors are implicated in cell differentiation/function in various tissues, we studied the function of basic Krüppel-like factor (BKLF/KLF3) in B cells. Whereas B-cell development in the bone marrow of KLF3-transgenic mice was unaffected, MZ B-cell numbers in spleen were increased considerably. As revealed in chimeric mice, this occurred cell autonomously, increasing both MZ and peritoneal B1 B-cell subsets. Comparing KLF3-transgenic and nontransgenic follicular B cells by RNA-microarray revealed that KLF3 regulates a subset of genes that was similarly up-regulated/down-regulated on normal MZ B-cell differentiation. Indeed, KLF3 expression overcame the lack of MZ B cells caused by different genetic alterations, such as CD19-deficiency or blockade of B-cell activating factor-receptor signaling, indicating that KLF3 may complement alternative nuclear factor-κB signaling. Thus, KLF3 is a driving force toward MZ B-cell maturation.
AU  - Turchinovich, G.*
AU  - Vu, T.T.*
AU  - Frommer, F.*
AU  - Kranich, J.*
AU  - Schmid, S.*
AU  - Alles, M.*
AU  - Loubert, J.B.*
AU  - Goulet, J.P.*
AU  - Zimber-Strobl, U.
AU  - Schneider, P.*
AU  - Bachl, J.*
AU  - Pearson, R.*
AU  - Crossley, M.*
AU  - Agenès, F.*
AU  - Kirberg, J.*
C1  - 6487
C2  - 28764
CY  - Washington, USA
SP  - 3780-3792
TI  - Programming of marginal zone B-cell fate by basic Krüppel-like factor (BKLF/KLF3).
JO  - Blood
VL  - 117
IS  - 14
PB  - American Society of Hematology
PY  - 2011
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - A large proportion of patients with mutations in the Wiskott-Aldrich syndrome (WAS) protein gene exhibit the milder phenotype termed X-linked thrombocytopenia (XLT). Whereas stem cell transplantation at an early age is the treatment of choice for patients with WAS, therapeutic options for patients with XLT are controversial. In a retrospective multicenter study we defined the clinical phenotype of XLT and determined the probability of severe disease-related complications in patients older than 2 years with documented WAS gene mutations and mild-to-moderate eczema or mild, infrequent infections. Enrolled were 173 patients (median age, 11.5 years) from 12 countries spanning 2830 patient-years. Serious bleeding episodes occurred in 13.9%, life-threatening infections in 6.9%, autoimmunity in 12.1%, and malignancy in 5.2% of patients. Overall and event-free survival probabilities were not significantly influenced by the type of mutation or intravenous immunoglobulin or antibiotic prophylaxis. Splenectomy resulted in increased risk of severe infections. This analysis of the clinical outcome and molecular basis of patients with XLT shows excellent long-term survival but also a high probability of severe disease-related complications. These observations will allow better decision making when considering treatment options for individual patients with XLT.
AU  - Albert, M.H.*
AU  - Bittner, T.C.*
AU  - Nonoyama, S.*
AU  - Notarangelo, L.D.*
AU  - Burns, S.*
AU  - Imai, K.*
AU  - Espanol, T.*
AU  - Fasth, A.*
AU  - Pellier, I.*
AU  - Strauss, G.*
AU  - Morio, T.*
AU  - Gathmann, B.*
AU  - Noordzij, J.G.*
AU  - Fillat, C.*
AU  - Hoenig, M.*
AU  - Nathrath, M.
AU  - Meindl, A.*
AU  - Pagel, P.*
AU  - Wintergerst, U.*
AU  - Fischer, A.*
AU  - Thrasher, A.J.*
AU  - Belohradsky, B.H.*
AU  - Ochs, H.D.*
C1  - 2036
C2  - 27848
SP  - 3231-3238
TI  - X-linked thrombocytopenia (XLT) due to WAS mutations: Clinical characteristics, long-term outcome, and treatment options.
JO  - Blood
VL  - 115
IS  - 16
PB  - Amer. Soc. Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Blood of both humans and mice contains 2 main monocyte subsets. Here, we investigated the extent of their similarity using a microarray approach. Approximately 270 genes in humans and 550 genes in mice were differentially expressed between subsets by 2-fold or more. More than 130 of these gene expression differences were conserved between mouse and human monocyte subsets. We confirmed numerous of these differences at the cell surface protein level. Despite overall conservation, some molecules were conversely expressed between the 2 species' subsets, including CD36, CD9, and TREM-1. Other differences included a prominent peroxisome proliferator-activated receptor gamma (PPARgamma) signature in mouse monocytes, which is absent in humans, and strikingly opposed patterns of receptors involved in uptake of apoptotic cells and other phagocytic cargo between human and mouse monocyte subsets. Thus, whereas human and mouse monocyte subsets are far more broadly conserved than currently recognized, important differences between the species deserve consideration when models of human disease are studied in mice.
AU  - Ingersoll, M.A.*
AU  - Spanbroek,R.*
AU  - Lottaz, C.*
AU  - Gautier, E.L.*
AU  - Frankenberger, M.
AU  - Hoffmann, R.*
AU  - Lang, R.*
AU  - Haniffa, M.*
AU  - Collin, M.*
AU  - Tacke, F.*
AU  - Habenicht, A.J.R.*
AU  - Ziegler-Heitbrock, L.
AU  - Randolph, G.J.*
C1  - 4726
C2  - 28198
CY  - Washington
SP  - 10-19
TI  - Comparison of gene expression profiles between human and mouse monocyte subsets.
JO  - Blood
VL  - 115
IS  - 3
PB  - Amer. Soc. Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The 46/1 JAK2 haplotype predisposes to V617F-positive myeloproliferative neoplasms, but the underlying mechanism is obscure. We analyzed essential thrombocythemia patients entered into the PT-1 studies and, as expected, found that 46/1 was overrepresented in V617F-positive cases (n = 404) versus controls (n = 1492, P = 3.9 x 10(-11)). The 46/1 haplotype was also overrepresented in cases without V617F (n = 347, P = .009), with an excess seen for both MPL exon 10 mutated and V617F, MPL exon 10 nonmutated cases. Analysis of further MPL-positive, V617F-negative cases confirmed an excess of 46/1 (n = 176, P = .002), but no association between MPL mutations and MPL haplotype was seen. An excess of 46/1 was also seen in JAK2 exon 12 mutated cases (n = 69, P = .002), and these mutations preferentially arose on the 46/1 chromosome (P = .029). No association between 46/1 and clinical or laboratory features was seen in the PT-1 cohort either with or without V617F. The excess of 46/1 in JAK2 exon 12 cases is compatible with both the "hypermutability" and "fertile ground" hypotheses, but the excess in MPL-mutated cases argues against the former. No difference in sequence, splicing, or expression of JAK2 was found on 46/1 compared with other haplotypes, suggesting that any functional difference of JAK2 on 46/1, if it exists, must be relatively subtle.
AU  - Jones, A.V.*
AU  - Campbell, P.J.*
AU  - Beer, P.A.*
AU  - Schnittger, S.*
AU  - Vannucchi, A.M.*
AU  - Zoi, K.*
AU  - Percy, M.J.*
AU  - McMullin, M.F.*
AU  - Scott, L.M.*
AU  - Tapper, W.*
AU  - Silver, R.T.*
AU  - Oscier, D.*
AU  - Harrison, C.N.*
AU  - Grallert, H.
AU  - Kisialiou, A.*
AU  - Strike, P.*
AU  - Chase, A.J.*
AU  - Green, A.R.*
AU  - Cross, N.C.P.*
C1  - 955
C2  - 27393
SP  - 4517-4523
TI  - The JAK2 46/1 haplotype predisposes to MPL-mutated myeloproliferative neoplasms.
JO  - Blood
VL  - 115
IS  - 22
PB  - American Soc. of Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - T-cell development depends on recruitment of bone marrow-derived precursor cells to the thymus via a multistep adhesion cascade involving the chemokine receptor CCR9. However, CCR9 deficiency does not result in complete abrogation of progenitor entry into the adult thymus. Therefore, we tested the hypothesis that additional chemokine/chemokine receptor systems might play a role in this process. To this end, we generated mice deficient in both CCR9 and CCR7. Deficiency in both chemokine receptors resulted in severely reduced numbers of early T-cell progenitors and in near-complete abrogation of thymus reconstitution. Progenitors in bone marrow and peripheral blood remained largely unaffected in CCR7(-/-)CCR9(-/-) mice, and direct intrathymic transfer of precursors from CCR7(-/-)CCR9(-/-) mice as well as single-mutant mice showed that intrathymic differentiation of these precursors remained functional. Thus, our data reveal a previously unrecognized role of CCR7 in progenitor seeding of the adult thymus, which is largely masked by compensatory effects of CCR9 signals. In turn, CCR7 signals can partially compensate for CCR9 signals, thus explaining the rather mild phenotype of CCR9(-/-) mice with respect to progenitor seeding.
AU  - Krueger, A.*
AU  - Willenzon, S.*
AU  - Lyszkiewicz, M.*
AU  - Kremmer, E.
AU  - Forster, R.*
C1  - 5920
C2  - 27492
SP  - 1906-1912
TI  - CC chemokine receptor 7 and 9 double-deficient hematopoietic progenitors are severely impaired in seeding the adult thymus.
JO  - Blood
VL  - 115
IS  - 10
PB  - American Society Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Posttransplantation lymphoproliferative disease (PTLD) associated with Epstein-Barr virus (EBV) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation. PTLD is efficiently prevented by adoptive transfer of EBV-specific T cells from the donor. To make EBV-specific T cells available in urgent clinical situations, we developed a rapid protocol for their isolation by overnight stimulation of donor blood cells with peptides derived from 11 EBV antigens, interferon-gamma surface capture, and immunomagnetic separation. Six patients with PTLD received 1 transfusion of EBV-specific T cells. No response was seen in 3 patients who had late-stage disease with multiorgan dysfunction at the time of T-cell transfer. In 3 patients who received T cells at an earlier stage of disease, we observed complete and stable remission of PTLD. Two patients have remained free from EBV-associated disease for more than 2 years. CD8(+) T cells specific for EBV early antigens rapidly expanded after T-cell transfer, temporarily constituted greater than 20% of all peripheral blood lymphocytes, and were maintained throughout the observation period. Thus, a rapid and sustained reconstitution of a protective EBV-specific T-cell memory occurred after the infusion of small numbers of directly isolated EBV-specific T cells.
AU  - Moosmann, A.
AU  - Bigalke, I.
AU  - Tischer, J.
AU  - Schirrmann, L.
AU  - Kasten, J.
AU  - Tippmer, S.
AU  - Leeping, M.
AU  - Prevalsek, D.
AU  - Jaeger, G.*
AU  - Ledderose, G.
AU  - Mautner, J.
AU  - Hammerschmidt, W.
AU  - Schendel, D.J.
AU  - Kolb, H.-J.
C1  - 5655
C2  - 27734
SP  - 2960-2970
TI  - Effective and long-term control of EBV PTLD after transfer of peptide-selected T cells.
JO  - Blood
VL  - 115
IS  - 14
PB  - American Society of Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Transcription factor CCAAT enhancer binding protein alpha (C/EBPalpha) is essential for granulopoiesis and its function is deregulated in leukemia. Inhibition of E2F1, the master regulator of cell-cycle progression, by C/EBPalpha is pivotal for granulopoiesis. Recent studies show microRNA-223 (miR-223), a transcriptional target of C/EBPalpha, as a critical player during granulopoiesis. In this report, we demonstrate that during granulopoiesis microRNA-223 targets E2F1. E2F1 protein was up-regulated in miR-223 null mice. We show that miR-223 blocks cell-cycle progression in myeloid cells. miR-223 is down-regulated in different subtypes of acute myeloid leukemia (AML). We further show that E2F1 binds to the miR-223 promoter in AML blast cells and inhibits miR-223 transcription, suggesting that E2F1 is a transcriptional repressor of the miR-223 gene in AML. Our study supports a molecular network involving miR-223, C/EBPalpha, and E2F1 as major components of the granulocyte differentiation program, which is deregulated in AML.
AU  - Pulikkan, J.A.*
AU  - Dengler, V.*
AU  - Peramangalam, P.S.*
AU  - Peer Zada, A.A.*
AU  - Müller-Tidow, C.*
AU  - Bohlander, S.K.*
AU  - Tenen, D.G.*
AU  - Behre, G.*
C1  - 4503
C2  - 27806
SP  - 1768-1778
TI  - Cell-cycle regulator E2F1 and microRNA-223 comprise an autoregulatory negative feedback loop in acute myeloid leukemia.
JO  - Blood
VL  - 115
IS  - 9
PB  - American Society of Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The transcription factor CCAAT Enhancer Binding Protein alpha (C/EBPα) is crucial for granulopoiesis and is deregulated by various mechanisms in acute myeloid leukemia (AML). Mutations in the CEBPA gene are reported in 10% of human patients with AML. Even though the C/EBPα-mutants are known to display distinct biological function during leukemogenesis, the molecular basis for this subtype of AML remains elusive. We have recently showed the significance of deregulation of C/EBPα regulated microRNA (miR) in AML. In this study, we report that miR-34a is a novel target of C/EBPα in granulopoiesis. During granulopoiesis miR-34a targets E2F3 and blocks myeloid cell proliferation. Analysis of AML samples with CEBPA mutations revealed a lower expression of miR-34a and elevated levels of E2F3 as well as E2F1, a transcriptional target of E2F3. Manipulation of miR-34a re-programmes granulocytic differentiation of AML blast cells with CEBPA mutations. These results define miR-34a as a novel therapeutic target in AML with CEBPA mutations.
AU  - Pulikkan, J.A.*
AU  - Peramangalam, P.S.*
AU  - Dengler, V.*
AU  - Ho, P.A.*
AU  - Preudhomme, C.*
AU  - Meshinchi, S.*
AU  - Christopeit, M.*
AU  - Nibourel, O.*
AU  - Müller-Tidow, C.*
AU  - Bohlander, S.K.
AU  - Tenen, D.G.*
AU  - Behre, G.*
C1  - 6062
C2  - 27808
SP  - 5638-5649
TI  - C/EBPα-regulated microRNA-34a targets E2F3 during granulopoiesis and is downregulated in AML with CEBPA mutations.
JO  - Blood
VL  - 116
IS  - 25
PB  - American Society of Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - To identify the genetic basis of circulating concentrations of monocyte chemoattractant protein-1 (MCP-1), we conducted genome-wide association analyses for MCP-1 in 3 independent cohorts (n = 9598). The strongest association was for serum MCP-1 with a nonsynonymous polymorphism, rs12075 (Asp42Gly) in DARC, the gene for Duffy antigen receptor for chemokines, a known vascular reservoir of proinflammatory cytokines (minor allele frequency, 45.6%; P < 1.0 * 10(-323)). This association was supported by family-based genetic linkage at a locus encompassing the DARC gene (genome-wide P = 8.0 * 10(-13)). Asp42Gly accounted for approximately 20% of the variability in serum MCP-1 concentrations and also was associated with serum concentrations of interleukin-8 and RANTES. While exploring a lack of association between this polymorphism and EDTA plasma MCP-1 concentrations (P = .82), we determined that both clotting and exogenous heparan sulfate (unfractionated heparin) released substantial amounts of MCP-1 from Darc. Quantitative immunoflow cytometry failed to identify meaningful Asp42Gly-associated differences in Darc expression, suggesting that a functional change is responsible for the differential cytokine binding. We conclude that Asp42Gly is a major regulator of erythrocyte Darc-mediated cytokine binding and thereby the circulating concentrations of several proinflammatory cytokines. We have also identified for the first time 2 mechanisms for the release of reservoir chemokines with possible clinical implications.
AU  - Schnabel, R.B.*
AU  - Baumert, J.J.
AU  - Barbalic, M.*
AU  - Dupuis, J.*
AU  - Ellinor, P.T.*
AU  - Durda, P.*
AU  - Dehghan, A.*
AU  - Bis, J.C.*
AU  - Illig, T.
AU  - Morrison, A.C.*
AU  - Jenny, N.S.*
AU  - Keaney, J.F.*
AU  - Gieger, C.
AU  - Tilley, C.*
AU  - Yamamoto, J.F.*
AU  - Khuseyinova, N.*
AU  - Heiss, G.*
AU  - Doyle, M.*
AU  - Blankenberg, S.*
AU  - Herder, C.*
AU  - Walston, J.D.*
AU  - Zhu, Y.Y.*
AU  - Vasan, R.S.*
AU  - Klopp, N.
AU  - Boerwinkle, E.*
AU  - Larson, M.G.*
AU  - Psaty, B.M.*
AU  - Peters, A.
AU  - Ballantyne, C.M.*
AU  - Witteman, J.C.M.*
AU  - Hoogeveen, R.C.*
AU  - Benjamin, E.J.*
AU  - Koenig, W.*
AU  - Tracy, R.P.*
C1  - 1740
C2  - 27386
SP  - 5289-5299
TI  - Duffy antigen receptor for chemokines (Darc) polymorphism regulates circulating concentrations of monocyte chemoattractant protein-1 and other inflammatory mediators.
JO  - Blood
VL  - 115
IS  - 26
PB  - American Soc. of Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Platelets play a key role in hemostasis and various diseases including arterial thrombosis. Glycoprotein VI (GPVI) mediates adhesion to collagen structures exposed at sites of vascular injury and subsequent platelet activation. We determined the effects of specific activation of GPVI on the human platelet proteome. Isolated human platelets were stimulated with an activating monoclonal antibody specific for GPVI. Platelet proteins were analyzed by 2-dimensional difference gel electrophoresis (2D-DIGE) and mass spectrometry. We identified 8 differentially abundant proteins associated with cell signaling, metabolism, organization and rearrangement of the cytoskeleton, and membrane trafficking. Differentially abundant proteins included aldose reductase (AR), beta-centractin, charged multivesicular body protein 3, Src substrate cortactin, ERp57, and pleckstrin. Importantly, GPVI-modulated protein abundance was functionally relevant. Correspondingly, AR enzyme activity significantly increased upon GPVI activation and inhibition of AR resulted in reduced platelet aggregation. Furthermore, ERp57 was released upon ligation of platelet GPVI and increased the activity of tissue factor, a major initiator of blood coagulation. In summary, GPVI activation results in differential changes in abundance of platelet proteins, including AR and ERp57, which support platelet aggregation and platelet-dependent coagulation. These results provide further insight into the mechanisms that underlie platelet activation through the GPVI receptor and may help to identify novel pharmacologic targets.
AU  - Schulz, C.*
AU  - Leuschen, N.V.*
AU  - Fröhlich, T.*
AU  - Lorenz, M.*
AU  - Pfeiler, S.*
AU  - Gleissner, C.A.*
AU  - Kremmer, E.
AU  - Kessler, M.*
AU  - Khandoga, A.G.*
AU  - Engelmann, B.*
AU  - Ley, K.*
AU  - Massberg, S.*
AU  - Arnold, G.J.*
C1  - 5418
C2  - 27487
SP  - 4102-4110
TI  - Identification of novel downstream targets of platelet glycoprotein VI activation by differential proteome analysis: Implications for thrombus formation.
JO  - Blood
VL  - 115
IS  - 20
PB  - American Society of Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Monocytes and cells of the dendritic cell lineage circulate in blood and eventually migrate into tissue where they further mature and serve various functions, most notably in immune defense. Over recent years these cells have been characterized in detail with the use of cell surface markers and flow cytometry, and subpopulations have been described. The present document proposes a nomenclature for these cells and defines 3 types of monocytes (classical, intermediate, and nonclassical monocytes) and 3 types of dendritic cells (plasmacytoid and 2 types of myeloid dendritic cells) in human and in mouse blood. This classification has been approved by the Nomenclature Committee of the International Union of Immunological Societies, and we are convinced that it will facilitate communication among experts and in the wider scientific community.
AU  - Ziegler-Heitbrock, L.
AU  - Ancuta, P.*
AU  - Crowe, S.*
AU  - Dalod, M.*
AU  - Grau, V.*
AU  - Hart, D.N.*
AU  - Leenen, P.J.M.*
AU  - Liu, Y.J.*
AU  - MacPherson, G.*
AU  - Randolph, G.J.*
AU  - Scherberich, J.*
AU  - Schmitz, J.*
AU  - Shortman, K.*
AU  - Sozzani, S.*
AU  - Strobl, H.*
AU  - Zembala, M.*
AU  - Austyn, J.M.*
AU  - Lutz, M.B.*
C1  - 3731
C2  - 28007
CY  - Washington
SP  - e74-e80
TI  - Nomenclature of monocytes and dendritic cells in blood.
JO  - Blood
VL  - 116
IS  - 16
PB  - Amer. Soc. Hematology
PY  - 2010
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Defects in apoptosis contribute to poor outcome in pediatric acute lymphoblastic leukemia (ALL), calling for novel strategies that counter apoptosis resistance. Here, we demonstrate for the first time that small molecule inhibitors of the antiapoptotic protein XIAP cooperate with TRAIL to induce apoptosis in childhood acute leukemia cells. XIAP inhibitors at subtoxic concentrations, but not a structurally related control compound, synergize with TRAIL to trigger apoptosis and to inhibit clonogenic survival of acute leukemia cells, whereas they do not affect viability of normal peripheral blood lymphocytes, suggesting some tumor selectivity. Analysis of signaling pathways reveals that XIAP inhibitors enhance TRAIL-induced activation of caspases, loss of mitochondrial membrane potential, and cytochrome c release in a caspase-dependent manner, indicating that they promote a caspase-dependent feedback mitochondrial amplification loop. Of note, XIAP inhibitors even overcome Bcl-2-mediated resistance to TRAIL by enhancing Bcl-2 cleavage and Bak conformational change. Importantly, XIAP inhibitors kill leukemic blasts from children with ALL ex vivo and cooperate with TRAIL to induce apoptosis. In vivo, they significantly reduce leukemic burden in a mouse model of pediatric ALL engrafted in non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice. Thus, XIAP inhibitors present a promising novel approach for apoptosis-based therapy of childhood ALL.
AU  - Fakler, M.*
AU  - Loeder, S.*
AU  - Vogler, M.*
AU  - Schneider, K.
AU  - Jeremias, I.
AU  - Debatin, K.-M.*
AU  - Fulda, S.*
C1  - 572
C2  - 26883
SP  - 1710-1722
TI  - Small molecule XIAP inhibitors cooperate with TRAIL to induce apoptosis in childhood acute leukemia cells and overcome Bcl-2-mediated resistance.
JO  - Blood
VL  - 113
IS  - 8
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Neutrophils play a vital role in the immune defense, which is evident by the severity of neutropenia causing life-threatening infections. Granulocyte macrophage-colony stimulating factor (GM-CSF) controls homeostatic and emergency development of granulocytes. However, little is known about the contribution of the downstream mediating transcription factors signal transducer and activator of transcription 5A and 5B (STAT5A/B). To elucidate the function of this pathway, we generated mice with complete deletion of both Stat5a/b genes in hematopoietic cells. In homeostasis, peripheral neutrophils were markedly decreased in these animals. Moreover, during emergency situations, such as myelosuppression, Stat5a/b mutant mice failed to produce enhanced levels of neutrophils and were unable to respond to GM-CSF. Both the GM-CSF permitted survival of mature neutrophils and the generation of granulocytes from granulocyte-macrophage progenitors (GMPs) were markedly reduced in Stat5a/b mutants. GMPs showed impaired colony-formation ability with reduced number and size of colonies on GM-CSF stimulation. Moreover, continuous cell fate analyses by time-lapse microscopy and single cell tracking revealed that Stat5a/b-null GMPs showed both delayed cell-cycle progression and increased cell death. Finally, transcriptome analysis indicated that STAT5A/B directs GM-CSF signaling through the regulation of proliferation and survival genes.
AU  - Kimura, A.*
AU  - Rieger, M.
AU  - Simone, J.M.*
AU  - Chen, W.P.*
AU  - Wickre, M.C.*
AU  - Zhu, B.M.*
AU  - Hoppe, P.S.
AU  - O'Shea, J.J.*
AU  - Schroeder, T.
AU  - Hennighausen, L.*
C1  - 901
C2  - 26726
SP  - 4721-4728
TI  - The transcription factors STAT5A/B regulate GM-CSF-mediated granulopoiesis.
JO  - Blood
VL  - 114
IS  - 21
PB  - American Society of Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Signaling through tumor necrosis factor receptor 1 (TNFR1) controls bacterial infections and the induction of inflammatory Th1 cell-mediated autoimmune diseases. By dissecting Th1 cell-mediated delayed-type hypersensitivity responses (DTHRs) into single steps, we localized a central defect to the missing TNFR1 expression by endothelial cells (Ecs). Adoptive transfer and mast cell knockin experiments into Kit(W)/Kit(W-v), TNF-/-, and TNFR1(-/-) mice showed that the signaling defect exclusively affects mast cell-EC interactions but not T cells or antigen-presenting cells. As a consequence, TNFR1(-/-) mice had strongly reduced mRNA and protein expression of P-selectin, E-selectin, ICAM-1, and VCAM-1 during DTHR elicitation. In consequence, intravital fluorescence microscopy revealed up to 80% reduction of leukocyte rolling and firm adhesion in TNFR1(-/-) mice. As substitution of TNF-/- mice with TNF-producing mast cells fully restored DTHR in these mice, signaling of mast cell-derived TNF through TNFR1-expressing Ecs is essential for the recruitment of leukocytes into sites of inflammation.
AU  - Kneilling, M.*
AU  - Mailhammer, R.
AU  - Hültner, L.
AU  - Schönberger, T.*
AU  - Fuchs, K.*
AU  - Schaller, M.*
AU  - Bukala, D.*
AU  - Massberg, S.*
AU  - Sander, C.A.*
AU  - Braumüller, H.*
AU  - Eichner, M.*
AU  - Maier, K.L.
AU  - Hallmann, R.*
AU  - Pichler, B.J.*
AU  - Haubner, R.*
AU  - Gawaz, M.
AU  - Pfeffer, K.*
AU  - Biedermann, T.*
AU  - Röcken, M.*
C1  - 894
C2  - 26337
CY  - Washington
SP  - 1696-1706
TI  - Direct crosstalk between mast cell-TNF and TNFR1-expressing endothelia mediates local tissue inflammation.
JO  - Blood
VL  - 114
IS  - 8
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The canonical mode of transcriptional activation by both the Epstein-Barr viral protein, Epstein-Barr virus-encoded nuclear antigen 2 (EBNA2), and an activated Notch receptor (Notch-IC) requires their recruitment to RBPJ, suggesting that EBNA2 uses the Notch pathway to achieve B-cell immortalization. To gain further insight into the biologic equivalence between Notch-IC and EBNA2, we performed a genome-wide expression analysis, revealing that Notch-IC and EBNA2 exhibit profound differences in the regulation of target genes. Whereas Notch-IC is more potent in regulating genes associated with differentiation and development, EBNA2 is more potent in inducing viral and cellular genes involved in proliferation, survival, and chemotaxis. Because both EBNA2 and Notch-IC induced the expression of cell cycle associated genes, we analyzed whether Notch1-IC or Notch2-IC can replace EBNA2 in B-cell immortalization. Although Notch-IC could drive quiescent B cells into the cell cycle, B-cell immortalization was not maintained, partially due to an increased apoptosis rate in Notch-IC expressing cells. Expression analysis revealed that both EBNA2 and Notch-IC induced the expression of proapoptotic genes, but only in EBNA2-expressing cells were antiapoptotic genes strongly up-regulated. These findings suggest that Notch signaling in B cells and B-cell lymphomas is only compatible with proliferation if pathways leading to antiapototic signals are active.
AU  - Kohlhof, H.
AU  - Hampel, F.
AU  - Hoffmann, R.*
AU  - Burtscher, H.*
AU  - Weidle, U.H.*
AU  - Hölzel, M.*
AU  - Eick, D.
AU  - Zimber-Strobl, U.
AU  - Strobl, L.J.
C1  - 2086
C2  - 26546
SP  - 5506-5515
TI  - Notch1, Notch2, and Epstein-Barr virus-encoded nuclear antigen 2 signaling differentially affects proliferation and survival of Epstein-Barr virus-infected B cells.
JO  - Blood
VL  - 113
IS  - 22
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Chromosomal translocations generating fusion proteins are frequently found in human leukemias. The fusion proteins play an important role in leukemogenesis by subverting the function of one or both partner proteins. The leukemogenic CALM-AF10 fusion protein is capable of interacting with the histone H3 lysine 79 ( H3K79)-specific methyltransferase hDOT1L through the fused AF10 moiety. This interaction leads to local H3K79 hypermethylation on Hoxa5 loci, which up-regulates the expression of Hoxa5 and contributes to leukemogenesis. However, the long latency of leukemogenesis of CALM-AF10 transgenic mice suggests that the direct effects of fusion oncogene are not sufficient for the induction of leukemia. In this study, we show that the CALM-AF10 fusion protein can also greatly reduce global H3K79 methylation in both human and murine leukemic cells by disrupting the AF10-mediated association of hDOT1L with chromatin. Cells with reduced H3K79 methylation are more sensitive to gamma-irradiation and display increased chromosomal instability. Consistently, leukemia patients harboring CALM-AF10 fusion have more secondary chromosomal aberrations. These findings suggest that chromosomal instability associated with global epigenetic alteration contributes to malignant transformation in certain leukemias, and that leukemias with this type of epigenetic alteration might benefit from treatment regimens containing DNA-damaging agents. This study is registered with www.clinicaltrials.gov as NCT00266136. (Blood. 2009; 114: 651-658)
AU  - Lin, Y.H.*
AU  - Kakadiya, P.M.
AU  - Chen, Y.
AU  - Li, Y.Q.*
AU  - Deshpande, A.J.
AU  - Buske, C.
AU  - Zhang, K.L.*
AU  - Zhang, Y.*
AU  - Xu, G.L.*
AU  - Bohlander, S.K.
C1  - 513
C2  - 26649
SP  - 651-658
TI  - Global reduction of the epigenetic H3K79 methylation mark and increased chromosomal instability in CALM-AF10-positive leukemias.
JO  - Blood
VL  - 114
IS  - 3
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Mutations in the NPM1 gene represent the most frequent genetic alterations in patients with acute myeloid leukemia (AML) and are associated with a favorable outcome. In 690 normal karyotype (NK) AML patients the complete remission rates (CRs) and the percentage of patients with adequate in vivo blast cell reduction 1 week after the end of the first induction cycle were significantly higher in NPM1(+) (75% and 80%, respectively) than in NPM1(-) (57% and 57%, respectively) patients, but were unaffected by the FLT3-ITD status. Multivariate analyses revealed the presence of a NPM1 mutation as an independent positive prognostic factor for the achievement of an adequate day-16 blast clearance and a CR. In conclusion, NPM1(+) blast cells show a high in vivo sensitivity toward induction chemotherapy irrespective of the FLT3-ITD mutation status. These findings provide insight into the pathophysiology and help to understand the favorable clinical outcome of patients with NPM1(+) AML.
AU  - Schneider, F.*
AU  - Hoster, E.*
AU  - Unterhalt, M.*
AU  - Schneider, S.*
AU  - Dufour, A.*
AU  - Benthaus, T.*
AU  - Mellert, G.*
AU  - Zellmeier, E.*
AU  - Bohlander, S.K.
AU  - Feuring-Buske, M.
AU  - Buske, C.
AU  - Braess, J.*
AU  - Fritsch, S.*
AU  - Heinecke, A.*
AU  - Sauerland, M.C.*
AU  - Berdel, W.E.*
AU  - Buechner, T.*
AU  - Woermann, B.J.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
C1  - 2508
C2  - 26884
SP  - 5250-5253
TI  - NPM1 but not FLT3-ITD mutations predict early blast cell clearance and CR rate in patients with normal karyotype AML (NK-AML) or high-risk myelodysplastic syndrome (MDS).
JO  - Blood
VL  - 113
IS  - 21
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Mean platelet volume (MPV) and platelet count (PLT) are highly heritable and tightly regulated traits. We performed a genome-wide association study for MPV and identified one SNP, rs342293, as having highly significant and reproducible association with MPV (per-G allele effect 0.016 +/- 0.001 log fL; P < 1.08 x 10(-24)) and PLT (per-G effect -4.55 +/- 0.80 10(9)/L; P < 7.19 x 10(-8)) in 8586 healthy subjects. Whole-genome expression analysis in the 1-MB region showed a significant association with platelet transcript levels for PIK3CG (n = 35; P = .047). The G allele at rs342293 was also associated with decreased binding of annexin V to platelets activated with collagen-related peptide (n = 84; P = .003). The region 7q22.3 identifies the first QTL influencing platelet volume, counts, and function in healthy subjects. Notably, the association signal maps to a chromosome region implicated in myeloid malignancies, indicating this site as an important regulatory site for hematopoiesis. The identification of loci regulating MPV by this and other studies will increase our insight in the processes of megakaryopoiesis and proplatelet formation, and it may aid the identification of genes that are somatically mutated in essential thrombocytosis.
AU  - Soranzo, N.*
AU  - Rendon, A.*
AU  - Gieger, C.
AU  - Jones, C.I.*
AU  - Watkins, N.A.*
AU  - Menzel, S.*
AU  - Döring, A.
AU  - Stephens, J.*
AU  - Prokisch, H.
AU  - Erber, W.*
AU  - Potter, S.C.*
AU  - Bray, S.L.*
AU  - Burns, P.*
AU  - Jolley, J.*
AU  - Falchi, M.*
AU  - Kühnel, B.
AU  - Erdmann, J.*
AU  - Schunkert, H.*
AU  - Samani, N.J.*
AU  - Illig, T.
AU  - Garner, S.F.*
AU  - Rankin, A.*
AU  - Meisinger, C.
AU  - Bradley, J.R.*
AU  - Thein, S.L.*
AU  - Goodall, A.H.*
AU  - Spector, T.D.*
AU  - Deloukas, P.*
AU  - Ouwehand, W.H.
C1  - 2145
C2  - 26131
SP  - 3831-3837
TI  - A novel variant on chromosome 7q22.3 associated with mean platelet volume, counts, and function.
JO  - Blood
VL  - 113
IS  - 16
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Adoptive transfer of T cells expressing transgenic T-cell receptors (TCRs) with antitumor function is a hopeful new therapy for patients with advanced tumors; however, there is a critical bottleneck in identifying high-affinity TCR specificities needed to treat different malignancies. We have developed a strategy using autologous dendritic cells cotransfected with RNA encoding an allogeneic major histocompatibility complex mole-cule and a tumor-associated antigen to obtain allo-restricted peptide-specific T cells having superior capacity to recognize tumor cells and higher functional avidity. This approach provides maximum flexibility because any major histocompatibility complex molecule and any tumor-associated antigen can be combined in the dendritic cells used for priming of autologous T cells. TCRs of allorestricted T cells, when expressed as transgenes in activated peripheral blood lymphocytes, transferred superior function compared with self-restricted TCR. This approach allows high-avidity T cells and TCR specific for tumor-associated self-peptides to be easily obtained for direct adoptive T-cell therapy or for isolation of therapeutic transgenic TCR sequences.
AU  - Wilde, S.
AU  - Sommermeyer, D.*
AU  - Frankenberger, B.
AU  - Schiemann, M.
AU  - Milosevic, S.
AU  - Spranger, S.
AU  - Pohla, H.
AU  - Uckert, W.*
AU  - Busch, D.H.
AU  - Schendel, D.J.
C1  - 826
C2  - 26606
CY  - WASHINGTON
SP  - 2131-2139
TI  - Dendritic cells pulsed with RNA encoding allogeneic MHC and antigen induce T cells with superior antitumor activity and higher TCR functional avidity.
JO  - Blood
VL  - 114
IS  - 10
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Experimental tumor vaccination and adoptive T-cell therapies show that interferon-gamma (IFN-gamma)-producing CD4(+) T helper cells (Th1) can be highly effective in tumor prevention and therapy. Unexpectedly, first vaccine trials in humans revealed that tumor immune therapy may not only be protective, but, on the contrary, even promote tumor progression. Here, we analyzed T-cell immune responses to the epithelial cell adhesion molecule (EpCAM), one of the most common tumor-associated antigens (TAA) serving as immune target in colon cancer patients. Th-cell priming against EpCAM inevitably resulted in interleukin-4 (IL-4)-dominated Th2 responses, even under most stringent Th1-inducing conditions. These EpCAM-reactive Th2 cells rather promoted growth of EpCAM-expressing tumors. To analyze the role of IL-4 in tumor immune evasion, we generated EpCAM-reactive Th1 cells from IL-4.ko mice. These Th1 cells provided tumor-specific protection and established highly protective Th1 memory responses, even in naive BALB/c mice. Inhibition of tumor growth by Th1 cells resulted in intratumoral expression of cytokines of the IL-12 family and of IFN-gamma. Preventing activation-associated death of Th1 cells further increased intratumoral IFN-gamma expression and improved therapeutic efficacy. Thus, human TAA may promote tumor immune evasion by strongly favoring Th2 development.
AU  - Ziegler, A.*
AU  - Heidenreich, R.*
AU  - Braumüller, H.*
AU  - Wolburg, H.*
AU  - Weidemann, S.*
AU  - Mocikat, R.
AU  - Röcken, M.*
C1  - 371
C2  - 26148
SP  - 3494-3502
TI  - EpCAM, a human tumor-associated antigen promotes Th2 development and tumor immune evasion.
JO  - Blood
VL  - 113
IS  - 15
PB  - Amer Soc Hematology
PY  - 2009
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Herens, C.*
AU  - Lambert, F.*
AU  - Quintanilla-Martinez, L.
AU  - Bisig, B.*
AU  - Deusings, C.*
AU  - de Leval, L.*
C1  - 4160
C2  - 25124
SP  - 1745-1746
TI  - Cyclin D1-negative mantle cell lymphoma with cryptic t(12;14)(p13;q32) and cyclin D2 overexpression.
JO  - Blood
VL  - 111
IS  - 3
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) is the third most frequent virus-associated human malignancy. How this tumor escapes immune recognition despite the expression of several viral antigens has remained poorly understood. Our previous in vitro studies have shown that NPC cells release exosomes containing high amounts of galectin-9, a ligand of the membrane receptor Tim-3, which is able to induce apoptosis in mature Th1 lymphocytes. Here, we sought to determine whether galectin-9-carrying exosomes were produced in NPC patients and whether such exosomes might play a role in the immune evasion of NPC cells. We report that galectin-9-containing exosomes are selectively detected in plasma samples from NPC patients and mice xenografted with NPC tumors. The incorporation into exosomes protects galectin-9 against proteolytic cleavage but retains its Tim-3-binding capacity. Importantly, NPC exosomes induce massive apoptosis in EBV-specific CD4+ cells used as a model of target T cells. This effect is inhibited by both anti-Tim-3 and anti-galectin-9 blocking antibodies. These results indicate that blocking galectin-9/Tim-3 interaction in vivo might alleviate the Th1 suppressive effect of NPC exosomes and sustain anti-tumoral T cell responses, and thereby improve clinical efficacy of immunotherapeutic approaches against NPC.
AU  - Klibi, J.*
AU  - Niki, T.*
AU  - Riedel, A.*
AU  - Pioche-Durieu, C.*
AU  - Souquere, S.*
AU  - Rubinstein, E.*
AU  - Le, Moulec, S.*
AU  - Guigay, J.*
AU  - Hirashima, M.*
AU  - Guemira, F.*
AU  - Adhikary, D.
AU  - Mautner, J.
AU  - Busson, P.*
C1  - 5009
C2  - 25923
SP  - 1957-1966
TI  - Blood diffusion and Th1-suppressive effects of galectin-9-containing exosomes released by Epstein-Barr virus-infected nasopharyngeal carcinoma cells.
JO  - Blood
VL  - 113
IS  - 9
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The fact that you can vaccinate a child at 5 years of age and find lymphoid B cells and antibodies specific for this vaccination 70 years later remains an immunologic enigma. It has never been determined how these long-lived memory B cells are maintained and whether they are protected by storage in a special niche. We report that, whereas blood and spleen compartments present similar frequencies of IgG(+) cells, antismallpox memory B cells are specifically enriched in the spleen where they account for 0.24% of all IgG(+) cells (ie, 10-20 million cells) more than 30 years after vaccination. They represent, in contrast, only 0.07% of circulating IgG(+) B cells in blood (ie, 50-100,000 cells). An analysis of patients either splenectomized or rituximab-treated confirmed that the spleen is a major reservoir for long-lived memory B cells. No significant correlation was observed between the abundance of these cells in blood and serum titers of antivaccinia virus antibodies in this study, including in the contrasted cases of B cell-depleting treatments. Altogether, these data provide evidence that in humans, the two arms of B-cell memory--long-lived memory B cells and plasma cells--have specific anatomic distributions--spleen and bone marrow--and homeostatic regulation.
AU  - Mamani-Matsuda, M.*
AU  - Cosma, A.
AU  - Weller, S.*
AU  - Faili, A.*
AU  - Staib, C.
AU  - Garçon, L.*
AU  - Hermine, O.*
AU  - Beyne-Rauzy, O.*
AU  - Fieschi, C.*
AU  - Pers, J.O.*
AU  - Arakelyan, N.*
AU  - Varet, B.*
AU  - Sauvanet, A.*
AU  - Berger, A.*
AU  - Paye, F.*
AU  - Andrieu, J.M.*
AU  - Michel, M.*
AU  - Godeau, B.*
AU  - Buffet, P.*
AU  - Reynaud, C.A.*
AU  - Weill, J.C.*
C1  - 704
C2  - 25317
SP  - 4653-4659
TI  - The human spleen is a major reservoir for long-lived vaccinia virus-specific memory B cells.
JO  - Blood
VL  - 111
IS  - 9
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Patients with cytogenetically normal acute myeloid leukemia (CN-AML) show heterogeneous treatment outcomes. We used gene-expression profiling to develop a gene signature that predicts overall survival (OS) in CN-AML. Based on data from 163 patients treated in the German AMLCG 1999 trial and analyzed on oligonucleotide microarrays, we used supervised principal component analysis to identify 86 probe sets (representing 66 different genes), which correlated with OS, and defined a prognostic score based on this signature. When applied to an independent cohort of 79 CN-AML patients, this continuous score remained a significant predictor for OS (hazard ratio [HR], 1.85; P = .002), event-free survival (HR = 1.73; P = .001), and relapse-free survival (HR = 1.76; P = .025). It kept its prognostic value in multivariate analyses adjusting for age, FLT3 ITD, and NPM1 status. In a validation cohort of 64 CN-AML patients treated on CALGB study 9621, the score also predicted OS (HR = 4.11; P < .001), event-free survival (HR = 2.90; P < .001), and relapse-free survival (HR = 3.14, P < .001) and retained its significance in a multivariate model for OS. In summary, we present a novel gene-expression signature that offers additional prognostic information for patients with CN-AML.
AU  - Metzeler, K.H.*
AU  - Hummel, M.*
AU  - Bloomfield, C.D.*
AU  - Spiekermann, K.
AU  - Braess, J.*
AU  - Sauerland, M.C.*
AU  - Heinecke, A.*
AU  - Radmacher, M.*
AU  - Marcucci, G.*
AU  - Whitman, SP.*
AU  - Maharry, K.*
AU  - Paschka, P.*
AU  - Larson, R.A.*
AU  - Berdel, W.E.*
AU  - Büchner, T.*
AU  - Wörmann, B.*
AU  - Mansmann, U.*
AU  - Hiddemann, W.
AU  - Bohlander, S.K.
AU  - Buske, C.
AU  - Cancer and Leukemia Group B (*)
AU  - German AML Cooperative Group (*)
C1  - 3456
C2  - 25788
SP  - 4193-4201
TI  - An 86-probe-set gene-expression signature predicts survival in cytogenetically normal acute myeloid leukemia.
JO  - Blood
VL  - 112
IS  - 10
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The Epstein-Barr virus (EBV) protein LMP1 is considered to be a functional homologue of the CD40 receptor. However, in contrast to the latter, LMP1 is a constitutively active signaling molecule. To compare B cell-specific LMP1 and CD40 signaling in an unambiguous manner, we generated transgenic mice conditionally expressing a CD40/LMP1 fusion protein, which retained the LMP1 cytoplasmic tail but has lost the constitutive activity of LMP1 and needs to be activated by the CD40 ligand. We show that LMP1 signaling can completely substitute CD40 signaling in B cells, leading to normal B-cell development, activation, and immune responses including class-switch recombination, germinal center formation, and somatic hypermutation. In addition, the LMP1-signaling domain has a unique property in that it can induce class-switch recombination to IgG1 independent of cytokines. Thus, our data indicate that LMP1 has evolved to imitate T-helper cell function allowing activation, proliferation, and differentiation of EBV-infected B cells independent of T cells.
AU  - Rastelli, J.
AU  - Hömig-Hölzel, C.
AU  - Seagal, J.*
AU  - Müller, W.*
AU  - Hermann, A.C.
AU  - Rajewsky, K.*
AU  - Zimber-Strobl, U.
C1  - 2199
C2  - 25244
SP  - 1448-1455
TI  - LMP1 signaling can replace CD40 signaling in B cells in vivo and has unique features of inducing class-switch recombination to IgG1.
JO  - Blood
VL  - 111
IS  - 3
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The mechanisms underlying deregulation of HOX gene expression in AML are poorly understood. The ParaHox gene CDX2 was shown to act as positive upstream regulator of several HOX genes. In this study, constitutive expression of Cdx2 caused perturbation of leukemogenic Hox genes such as Hoxa10 and Hoxb8 in murine hematopoietic progenitors. Deletion of the N-terminal domain of Cdx2 abrogated its ability to perturb Hox gene expression and to cause acute myeloid leukemia (AML) in mice. In contrast inactivation of the putative Pbx interacting site of Cdx2 did not change the leukemogenic potential of the gene. In an analysis of 115 patients with AML, expression levels of CDX2 were closely correlated with deregulated HOX gene expression. Patients with normal karyotype showed a 14-fold higher expression of CDX2 and deregulated HOX gene expression compared with patients with chromosomal translocations such as t(8:21) or t(15;17). All patients with AML with normal karyotype tested were negative for CDX1 and CDX4 expression. These data link the leukemogenic potential of Cdx2 to its ability to dysregulate Hox genes. They furthermore correlate the level of CDX2 expression with HOX gene expression in human AML and support a potential role of CDX2 in the development of human AML with aberrant Hox gene expression.
AU  - Rawat, V.P.S.
AU  - Thoene, S.
AU  - Naidu, V.M.
AU  - Arseni, N.
AU  - Heilmeier, B.*
AU  - Metzeler, K.*
AU  - Petropoulos, K.
AU  - Deshpande, A.J.
AU  - Quintanilla-Martinez, L.
AU  - Bohlander, S.K.
AU  - Spiekermann, K.
AU  - Hiddemann, W.
AU  - Feuring-Buske, M.
AU  - Buske, C.
C1  - 797
C2  - 25130
SP  - 309-319
TI  - Overexpression of CDX2 perturbs HOX gene expression in murine progenitors depending on its N-terminal domain and is closely correlated with deregulated HOX gene expression in human acute myeloid leukemia.
JO  - Blood
VL  - 111
IS  - 1
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Human herpesvirus-8 (HHV-8), also known as Kaposi sarcoma-associated herpesvirus (KSHV), is etiologically linked to primary effusion lymphoma (PEL). At least 10 KSHV-encoded proteins with potential roles in KSHV-associated neoplasia have been identified. However, with few exceptions, these putative oncogenes were analyzed in heterologous systems only using overexpression of single genes. Thus, the pathogenetic relevance of most of these putative oncogenes remains essentially unclear. We used RNA interference (RNAi) to knock down the expression of several KSHV genes in cultured PEL cells carrying the KSHV genome. The viral interferon-regulatory factor-3 (vIRF-3) was found to be required for proliferation and survival of cultured PEL cells. Knock-down of vIRF-3 expression by various RNAi approaches unequivocally resulted in reduced proliferation and increased activity of caspase-3 and/or caspase-7. Thus, vIRF-3 can be seen as a bona fide oncogene of KSHV-associated lymphoma. Surprisingly, although the related Epstein-Barr virus (EBV) is usually sufficient to immortalize human B lymphocytes, silencing of vIRF-3 reduced the viability of both EBV(-) and EBV(+) PEL cells. This suggests that KSHV is the driving force in the pathogenesis of PEL.
AU  - Wies, E.*
AU  - Mori, Y.*
AU  - Hahn, A.*
AU  - Kremmer, E.
AU  - Stürzl, M.*
AU  - Fleckenstein, B.*
AU  - Neipel, F.*
C1  - 732
C2  - 25128
SP  - 320-327
TI  - The viral interferon-regulatory factor-3 is required for the survival of KSHV-infected primary effusion lymphoma cells.
JO  - Blood
VL  - 111
IS  - 1
PB  - American Society of Hematology
PY  - 2008
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - RhoH is a small GTPase expressed only in the hematopoietic system. With the use of mice with targeted disruption of the RhoH gene, we demonstrated that RhoH is crucial for thymocyte maturation during DN3 to DN4 transition and during positive selection. Furthermore, the differentiation and expansion of DN3 and DN4 thymocytes in vitro were severely impaired. These defects corresponded to defective TCR signaling. Although RhoH is not required for TCR-induced activation of ZAP70 and ZAP70-mediated activation of p38, it is crucial for the tyrosine phosphorylation of LAT, PLCgamma1, and Vav1 and for the activation of Erk and calcium influx. These data suggest that RhoH is important for pre-TCR and TCR signaling because it allows the efficient interaction of ZAP70 with the LAT signalosome, thus regulating thymocyte development.
AU  - Dorn, T.*
AU  - Kuhn, U.*
AU  - Bungartz, G.*
AU  - Stiller, S.*
AU  - Bauer, M.*
AU  - Ellwart, J.W.
AU  - Peters, T.*
AU  - Scharffetter-Kochanek, K.*
AU  - Semmrich, M.*
AU  - Laschinger, M.*
AU  - Holzmann, B.*
AU  - Klinkert, W.E.*
AU  - Straten, P.T.*
AU  - Køllgaard, T.*
AU  - Sixt, M.*
AU  - Brakebusch, C.*
C1  - 2972
C2  - 24825
SP  - 2346-2355
TI  - RhoH is important for positive thymocyte selection and T-cell receptor signaling.
JO  - Blood
VL  - 109
IS  - 6
PB  - American Society of Hematology
PY  - 2007
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Many cells latently infected with Epstein-Barr virus (EBV), including certain virus-associated tumors, express latent membrane protein 2A (LMP2A), suggesting an important role for this protein in viral latency and oncogenesis. LMP2A mimics B-cell receptor signaling but can also act as a decoy receptor blocking B-cell receptor (BCR) activation. Studies of peripheral B cells have not resolved this apparent contradiction because LMP2A seems to be dispensable for EBV-induced transformation of these B cells in vitro. We show here that LMP2A is essential for growth transformation of germinal center B cells, which do not express the genuine BCR because of deleterious somatic hypermutations in their immunoglobulin genes. BCR-positive (BCR(+)) and BCR-negative (BCR(-)) B cells are readily transformed with a recombinant EBV encoding a conditional, floxed LMP2A allele, but the survival and continued proliferation of both BCR(+) and BCR(-) B cells is strictly dependent on LMP2A. These findings indicate that LMP2A has potent, distinct antiapoptotic and/or transforming characteristics and point to its role as an indispensable BCR mimic in certain B cells from which human B-cell tumors such as Hodgkin lymphoma originate.
AU  - Mancao, C.
AU  - Hammerschmidt, W.
C1  - 4764
C2  - 25000
SP  - 3715-3721
TI  - Epstein-Barr virus latent membrane protein 2A is a B-cell receptor mimic and essential for B-cell survival.
JO  - Blood
VL  - 110
IS  - 10
PB  - HighWire Press
PY  - 2007
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - In Ph(+) chronic myeloid leukemia (CML), the constitutively active Bcr-Abl kinase leads to the up-regulation and activation of multiple genes, which may subsequently result in the expression of leukemia-associated antigens. In this study, we investigated the immunogenicity of Bcr-Abl-regulated antigens by stimulating CD8(+) T lymphocytes with autologous dendritic cells transfected with RNA coding for Bcr-Abl wild-type or a kinase-deficient mutant. Significant HLA class I-restricted T-cell responses were detected against antigens regulated by the Bcr-Abl kinase, but not toward the Bcr-Abl protein itself. The T-cell repertoire of a patient with CML in major molecular remission due to imatinib mesylate was also dominated by T cells directed against Bcr-Abl-regulated antigens. These results encourage the development of immunotherapeutic approaches against Bcr-Abl-regulated antigens for the treatment of CML patients with residual disease following therapy with Bcr-Abl kinase inhibitors.
AU  - Scheich, F.*
AU  - Duyster, J.*
AU  - Peschel, C.*
AU  - Bernhard, H.
C1  - 3473
C2  - 25032
SP  - 2556-2560
TI  - The immunogenicity of Bcr-Abl expressing dendritic cells is dependent on the Bcr-Abl kinase activity and dominated by Bcr-Abl regulated antigens.
JO  - Blood
VL  - 110
IS  - 7
PB  - American Society of Hematology
PY  - 2007
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Cell-based immunotherapy in settings of allogeneic stem cell transplantation or donor leukocyte infusion has curative potential, especially in hematologic malignancies. However, this approach is severely restricted due to graft-versus-host disease (GvHD). This limitation may be overcome if target antigens are molecularly defined and effector cells are specifically selected. We chose formin-related protein in leukocytes 1 (FMNL1) as a target antigen after intensive investigation of its expression profile at the mRNA and protein levels. Here, we confirm restricted expression in peripheral blood mononuclear cells (PBMCs) from healthy donors but also observe overexpression in different leukemias and aberrant expression in transformed cell lines derived from solid tumors. We isolated allorestricted T-cell clones expressing a single defined TCR recognizing a particular HLA-A2–presented peptide derived from FMNL1. This T-cell clone showed potent antitumor activity against lymphoma and renal cell carcinoma cell lines, Epstein-Barr virus (EBV)–transformed B cells, and primary tumor samples derived from patients with chronic lymphocytic leukemia (CLL), whereas nontransformed cells with the exception of activated B cells were only marginally recognized. Allorestricted TCRs with specificity for naturally presented FMNL1-derived epitopes may represent promising reagents for the development of adoptive therapies in lymphoma and other malignant diseases.
AU  - Schuster, I.G.
AU  - Busch, D.H.
AU  - Eppinger, E.
AU  - Kremmer, E.
AU  - Milosevic, S.
AU  - Hennard, C.
AU  - Kuttler, C.
AU  - Ellwart, J.W.
AU  - Frankenberger, B.
AU  - Nößner, E.
AU  - Salat, C.*
AU  - Bogner, C.*
AU  - Borkhardt, A.*
AU  - Kolb, H.-J.
AU  - Krackhardt, A.M.
C1  - 5835
C2  - 24696
SP  - 2931-2939
TI  - Allorestricted T cells with specificity for the FMNL1-derived peptide PP2 have potent antitumor activity against hematologic and other malignancies.
JO  - Blood
VL  - 110
IS  - 8
PB  - American Society of Hematology
PY  - 2007
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - In this issue of Blood, Bros and colleagues report thoroughly how glucocorticoid (GC) treatment of a murine dendritic cell (DC) line during maturation induces a stable dendritic cell state with tolerogenic characteristics, similar to that described with GC-treated bone marrow–derived dendritic cells.
AU  - Tschoep, K.E.*
AU  - Nößner, E.
C1  - 3546
C2  - 25633
SP  - 3616
TI  - Understand tolerogenic dendritic cells.
JO  - Blood
VL  - 109
IS  - 9
PB  - American Society of Hematology
PY  - 2007
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - FLT3–internal tandem duplications (FLT3-ITDs) comprise a heterogeneous group of mutations in patients with acute leukemias that are prognostically important. To characterize the mechanism of transformation by FLT3-ITDs, we sequenced the juxtamembrane region (JM) of FLT3 from 284 patients with acute leukemias. The length of FLT3-ITDs varied from 2 to 42 amino acids (AAs) with a median of 17 AAs. The analysis of duplicated AAs showed that in the majority of patients, the duplications localize between AAs 591 to 599 (YVDFREYEY). Arginine 595 (R595) within this region is duplicated in 77% of patients. Single duplication of R595 in FLT3 conferred factor-independent growth to Ba/F3 cells and activated STAT5. Moreover, deletion or substitution of the duplicated R595 in 2 FLT3-ITD constructs as well as the deletion of wild-type R595 in FLT3-ITD substantially reduced the transforming potential and STAT5 activation, pointing to a critical role of the positive charge of R595 in stabilizing the active confirmation of FLT3-ITDs. Deletion of R595 in FLT3-WT nearly abrogated the ligand-dependent activation of FLT3-WT. Our data provide important insights into the molecular mechanism of transformation by FLT3-ITDs and show that duplication of R595 is important for the leukemic potential of FLT3-ITDs.
AU  - Vempati, S.
AU  - Reindl, C.
AU  - Kaza, S.K.
AU  - Kern, R.
AU  - Malamoussi, T.
AU  - Mellert, G.*
AU  - Schnittger, S.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
C1  - 4019
C2  - 24704
SP  - 686-694
TI  - Arginine 595 is duplicated in patients with acute leukemias carrying internal tandem duplications of FLT3 and modulates its transforming potential.
JO  - Blood
VL  - 110
IS  - 2
PB  - American Society of Hematology
PY  - 2007
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Activation-induced cytidine deaminase (AID) is necessary for immunoglobulin somatic hypermutation (SHM) and class switch recombination (CSR) in T-dependent immune response in germinal centers (GCs). The structural similarity of AID with RNA-editing enzymes and its largely cytoplasmic location have fueled controversial views of its mode of interaction with DNA. We show that AID, a mature B-cell–restricted cytoplasmic antigen, is relocated into the nucleus in 2.5% of CDKN1B–, CCNB1– GC cells. The GC dark zone and the outer zone (OZ), but not the light zone, contain nuclear and cytoplasmic AID+ blasts. AID+ cells in the OZ are in contact with T cells and CD23– follicular dendritic cells. In addition, AID is expressed in extrafollicular large proliferating B cells, 14% of which have nuclear AID. GC and extrafollicular AID+ cells express E47 but not the inhibiting BHLH protein Id2. Outside the GC, AID+ B cells are in contact with T cells and show partial evidence of CD40 plus bcr stimulation-dependent signature (CCL22, JunB, cMYC, CD30) but lack early and late plasma cell markers. The distribution of nuclear AID is consistent with the topography of SHM and CSR inside the GC and in extrafollicular activated B cells.
AU  - Cattoretti, G.*
AU  - Büttner, M.*
AU  - Shaknovich, R.*
AU  - Kremmer, E.
AU  - Alobeid, B.*
AU  - Niedobitek, G.*
C1  - 4933
C2  - 23894
SP  - 3967-3975
TI  - Nuclear and cytoplasmic AID in extrafollicular and germinal center B cells.
JO  - Blood
VL  - 107
IS  - 10
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - El-Maarri, O.*
AU  - Singer, H.*
AU  - Klein, C.*
AU  - Watzka, M.*
AU  - Herbiniaux, U.*
AU  - Brackmann, H.H.*
AU  - Schröder, J.*
AU  - Graw, J.
AU  - Müller, C.R.*
AU  - Schramm, W.*
AU  - Schwaab, R.*
AU  - Haaf, T.*
AU  - Hanfland, P.*
AU  - Oldenburg, J.*
C1  - 4301
C2  - 23913
SP  - 2759-2765
TI  - Lack of F8 mRNA: A novel mechanism leading to hemophilia A.
JO  - Blood
VL  - 107
IS  - 7
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Hofer, S.*
AU  - Pfeil, K.*
AU  - Niederegger, H.*
AU  - Ebner, S.*
AU  - Nguyen, V.A.*
AU  - Kremmer, E.
AU  - Auffinger, M.*
AU  - Neyer, S.*
AU  - Fürhapter, C.*
AU  - Heufler, C.*
C1  - 1803
C2  - 23478
SP  - 1003-1009
TI  - Dendritic cells regulate T-cell deattachment through the integrin-interavting protein CYTIP.
JO  - Blood
VL  - 107
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Kzhyshkowska, J.*
AU  - Mamidi, S.*
AU  - Gratchev, A.*
AU  - Kremmer, E.
AU  - Schuttermaier, C.*
AU  - Krusell, L.*
AU  - Haus, G.*
AU  - Utikal, J.*
AU  - Schledzewski, K.*
AU  - Scholtze, J.*
AU  - Goerdt, S.*
C1  - 829
C2  - 23890
SP  - 3221-3228
TI  - Novel stabilin-1 interacting chitinase-like protein (SI-CLP) is up-regulated in alternatively activated macrophages and secreted via lysosomal pathway.
JO  - Blood
VL  - 107
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - le Clorennec, C.*
AU  - Youlyouz-Marfak, I.*
AU  - Adriaenssens, E.*
AU  - Coll, J.*
AU  - Bornkamm, G.W.
AU  - Feuillard, J.*
C1  - 307
C2  - 23487
SP  - 2070-2078
TI  - EBV latency III immortalization program sensitizes B cells to induction of CD95-mediated apoptosis via LMP1: Role of NF-kB, STAT1 and p53.
JO  - Blood
VL  - 107
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Fc-receptor homolog 5 (FcRH5) is a recently identified B-cell membrane protein of unknown function. In Burkitt lymphoma cell lines with chromosome 1q21 abnormalities, FcRH5 expression is deregulated, implicating FcRH5 in lymphomagenesis. Epstein-Barr virus infects and immortalizes B cells, and is implicated in the etiology of several tumors of B-cell origin. Overexpression of genes located on 1q21-25 has been proposed as a surrogate for Epstein-Barr virus in Burkitt lymphoma. We now report that Epstein-Barr virus nuclear antigen 2 (EBNA2) markedly induces the expression of the FcRH5 gene, encoded on chromosome 1q21. Induction occurred in the absence of other viral proteins and did not require de novo protein synthesis. EBNA2 lacks a DNA-binding domain and can target responsive genes through the host DNA binding protein CBF1. We show that induction of FcRH5 by EBNA2 is strictly CBF1 dependent, as it was abolished in CBF1-deficient cells. Accordingly, EBNA2 targeted CBF1 binding sites present in the FcRH5 promoter in vivo, as detected by chromatin immunoprecipitation. These results identify FcRH5 as a novel, direct target of EBNA2 that may contribute to the development of Epstein-Barr virus-associated tumors.
AU  - Mohan, J.*
AU  - Dement-Brown, J.*
AU  - Maier, S.
AU  - Ise, T.*
AU  - Kempkes, B.
AU  - Tolnay, M.*
C1  - 3289
C2  - 23594
SP  - 4433-4439
TI  - Epstein-Barr virus nuclear antigen 2 induces FcRH5 expression through CBF1.
JO  - Blood
VL  - 107
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Quintanilla-Martinez, L.
AU  - Pittaluga, S.*
AU  - Miething, C.*
AU  - Klier, M.
AU  - Rudelius, M.*
AU  - Davies-Hill, T.*
AU  - Anastasov, N.
AU  - Martinez, A.*
AU  - Vivero, A.*
AU  - Duyster, J.*
AU  - Jaffe, E.S.*
AU  - Fend, F.*
AU  - Raffeld, M.*
C1  - 3558
C2  - 23824
SP  - 2029-2036
TI  - NPM-ALK-dependent expression of the transcription factor CCAAT/enhancer binding protein beta in ALK-positive anaplastic large cell lymphoma.
JO  - Blood
VL  - 108
IS  - 6
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Reindl, C.
AU  - Bagrintseva, K.
AU  - Vempati, S.
AU  - Schnittger, S.*
AU  - Ellwart, J.W.
AU  - Wenig, K.
AU  - Hopfner, K.-P.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
C1  - 4935
C2  - 23896
SP  - 3700-3707
TI  - Point mutations in the juxtamembrane domain of FLT define a new class of activating mutations in AML.
JO  - Blood
VL  - 107
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Rudelius, M.*
AU  - Pittaluga, S.*
AU  - Nishizuka, S.*
AU  - Pham, T.H.-T.*
AU  - Fend, F.*
AU  - Jaffe, E.S.*
AU  - Quintanilla-Martinez, L.
AU  - Raffeld, M.*
C1  - 5690
C2  - 23826
SP  - 1668-1676
TI  - Constitutive activation of Akt contributes to the pathogenesis and survival of mantle cell lymphoma.
JO  - Blood
VL  - 108
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Schnittger, S.*
AU  - Kohl, T.M.*
AU  - Haferlach, T.*
AU  - Kern, W.*
AU  - Hiddemann, W.
AU  - Spiekermann, K.
AU  - Schoch, C.*
C1  - 2909
C2  - 23983
SP  - 1791-1799
TI  - KIT-D816 mutations in AML1-ETO-positive AML are associated with impaired event-free overall survival.
JO  - Blood
VL  - 107
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Somatic hypermutation and class-switch recombination in germinal centers critically depend on activation-induced cytidine deaminase (AID). Deregulation of AID may lead to the aberrant activation or persistence of both genetic processes, thus contributing to the pathogenesis of B-cell lymphomas by mistargeted mutagenesis or recombination. The Epstein-Barr virus (EBV) establishes an asymptomatic latent infection in more than 90% of the human population, but it has also been linked to lymphomagenesis. A cooperative relationship of EBV and the germinal center reaction during the establishment of viral persistence has been postulated, but the contribution of EBV latent genes to the respective genetic events remains to be investigated in detail. In the present study, we show that activation of the EBV growth program has a clear inhibitory effect on AID expression, due to a negative effect of the master transcription factor of this program, EBNA2. This mechanism may counterbalance AID induction by the LMP1 protein, in order to prevent deleterious genetic changes during EBV-induced B-cell growth. EBNA2-mediated AID inhibition also provides a molecular explanation for the previously observed differences in somatic hypermutation activity in EBV-associated lymphoproliferative diseases, thus pointing to a crucial mechanism of EBV-mediated regulation of genomic integrity.
AU  - Tobollik, S.
AU  - Meyer, L.*
AU  - Buettner, M.*
AU  - Klemmer, S.*
AU  - Kempkes, B.
AU  - Kremmer, E.
AU  - Niedobitek, G.*
AU  - Jungnickel, B.
C1  - 3838
C2  - 24125
SP  - 3859-3864
TI  - Epstein-barr virus nuclear antigen 2 inhibits AID expression during EBV-driven B-cell growth.
JO  - Blood
VL  - 108
IS  - 12
PY  - 2006
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - It is generally accepted that priming of
      	antitumor CD8
 cytotoxic T lymphocytes
      	(CTLs) needs help that can be provided
      	by CD4
 T cells. We show that interactions
      	between dendritic cells (DCs) and
      	natural killer (NK) cells can bypass the T
      	helper arm in CTL induction. Bone marrow–
      	derived DCs caused rejection of the
      	A20 lymphoma and induced tumor-specific
      	long-term memory, although they
      	were not loaded with tumor-derived antigen.
      	Experiments using CD40 knock-out
      	mice and cell depletion showed that this
      	effect did not require CD4
 cells. Both
      	primary rejection and long-term CTL
      	memory were the result of NK cell activation
      	by DCs. NK cytotoxicity, which was
      	necessary for primary rejection, was dependent
      	on expression of natural killer
      	group 2 D (NKG2D) ligands on tumor
      	cells. Blocking of these ligands using
      	NKG2D tetramers abrogated tumor killing
      	in vitro and in vivo. The long-term response
      	was due to CTLs directed against
      	antigen(s) expressed on A20 and in vitro–
      	differentiated DCs. The mechanism leading
      	to CD4
 helper cell–independent CTL
      	responses was elucidated as a cascade
      	that was initiated by NK cell activation.
      	This pathway was dependent on interferon-
      	 expression and involved priming
      	endogenous DCs for interleukin-12 production.
      	Our data suggest a novel pathway
      	linking innate and adaptive immunity.
AU  - Adam, C.
AU  - King, S.
AU  - Allgeier, T.
AU  - Braumüller, H.*
AU  - Lüking, C.
AU  - Mysliwietz, J.
AU  - Kriegeskorte, A.K.
AU  - Busch, D.H.*
AU  - Röcken, M.*
AU  - Mocikat, R.
C1  - 666
C2  - 22725
SP  - 338-344
TI  - DC-NK cell cross talk as a novel CD4⁺ T-cell-independent pathway for antitumor CTL induction.
JO  - Blood
VL  - 106
IS  - 1
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - FLT3 (fms-like tyrosine kinase 3) is constitu-
      
      	tively activated in about 30% of patients with
      
      	acute myeloid leukemia (AML) and repre-
      
      	sents a disease-specific molecular marker.
      
      	Although FLT3-LM (length mutation) and
      
      	TKD (tyrosine kinase domain) mutations
      
      	have been considered to be mutually exclu-
      
      	sive, 1% to 2% of patients carry both muta-
      
      	tions. However, the functional and clinical
      
      	significance of this observation is unclear.
      
      	We demonstrate that FLT3-ITD-TKD dual mu-
      
      	tants induce drug resistance toward PTK
      
      	inhibitors and cytotoxic agents in in vitro
      
      	model systems. As molecular mechanisms
      
      	of resistance, we found that FLT3-ITD-TKD
      
      	mutants cause hyperactivation of STAT5
      
      	(signal transducer and activator of transcrip-
      
      	tion-5), leading to upregulation of Bcl-x(L)
      
      	and RAD51 and arrest in the G
      
      	2
      
      	M phase of
      
      	the cell cycle. Overexpression of Bcl-x(L)
      
      	was identified as the critical mediator of
      
      	drug resistance and recapitulates the PTK
      
      	inhibitor and daunorubicin-resistant pheno-
      
      	type in FLT3-ITD cells. The combination of
      
      	rapamycin, a selective mTOR inhibitor, and
      
      	FLT3 PTK inhibitors restored the drug sensi-
      
      	tivity in FLT3 dual mutant–expressing cells.
      
      	Our data provide the molecular basis for
      
      	understanding clinical FLT3 PTK inhibitor
      
      	resistance and point to therapeutical strate-
      
      	gies to overcome drug resistance in pa-
      
      	tients withAML.
AU  - Bagrintseva, K.
AU  - Geisenhof, S.
AU  - Kern, R.
AU  - Eichenlaub, S.
AU  - Reindl, C.
AU  - Ellwart, J.W.
AU  - Hiddemann, W.
AU  - Spiekermann, K.
C1  - 2274
C2  - 23229
SP  - 3679-3685
TI  - FLT3-ITD-TKD dual mutants associated with AML confer resistance to FLT3 PTK inhibitors and cytotoxic agents by overexpression of Bcl-x(L).
JO  - Blood
VL  - 105
IS  - 9
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The biosynthesis of aberrant immunoglobulin polypeptides by monoclonal plasma cells has been implicated in the pathogenesis of nonsecretory myeloma. Our studies of a patient with this disorder indeed have demonstrated the presence of abnormal κ light chains that resulted from a frameshift mutation in nucleotides encoding the constant region of the molecule. As a consequence of a 2-base deletion in codon 187 and loss of the normal stop codon, this portion of the κ chain was composed of 128 amino acids (rather than the expected 106), with a completely anomalous sequence after position 187 that included absence of the cysteines required for intrachain and interchain disulfide bonds. The unusual primary structure of this component was confirmed by mass spectrometric and amino acid sequence analyses of cytoplasmic protein extracts. Our studies provide the first evidence that human nonsecretory myeloma may result from an alteration in the light-chain constant region.
AU  - Coriu, D.*
AU  - Weaver, K. *
AU  - Schell, M. *
AU  - Eulitz, M.
AU  - Murphy, Ch.*
AU  - Weiss, D.T.*
AU  - Solomon, A.*
C1  - 4303
C2  - 23027
SP  - 829-831
TI  - A molecular basis for nonsecretory myeloma.
JO  - Blood
VL  - 104
IS  - 3
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Latent membrane protein 1 (LMP-1) of Epstein-Barr virus (EBV) promotes tumorigenesis by inhibiting apoptosis. We show that an important antiapoptotic activity of LMP-1 is the inhibition of Bcl2-associated protein X (Bax), a potent proapoptotic protein. BAX expression was regulated by LMP-1 activation of nuclear factor κB (NF-κB) via the C-terminal activation region 1 (CTAR-1) and CTAR-2. Interestingly, p65/p50 inhibited, whereas p50/p50 increased, BAX promoter activity as demonstrated by overexpression and selective inhibition of these NF-κB isoforms. Electrophoretic mobility shift analysis revealed that LMP-1 activates 2 of the 3 NF-κB binding sites (κB1-κB3) in the BAX promoter. LMP-1 induced binding of the NF-κB heterodimer p65/p50 to the κB2 site and of the p50/p50 homodimer to the κB3 site. Promoter mutation analysis revealed that the κB2 site is necessary for inhibition of BAX promoter activity and the κB3 site, for its activation. However, the activation of the BAX promoter by LMP-1 was observed only in the presence of specific inhibitors of p65/p50. In all other cases, LMP-1 inhibited BAX promoter activity. Most importantly, the antiapoptotic activity of LMP-1 was considerably decreased in cells deficient for BAX. These results indicate that the inhibition of Bax may be an important antiapoptotic activity of LMP-1.
AU  - Grimm, T.
AU  - Schneider, S.
AU  - Naschberger, E.
AU  - Huber, J.*
AU  - Guenzi, E.
AU  - Kieser, A.
AU  - Reitmeir, P.
AU  - Schulz, T.F.*
AU  - Morris, C.A.*
AU  - Stürzl, M.
C1  - 172
C2  - 22862
SP  - 3263-3269
TI  - EBV latent membrane protein-1 protects B cells from apoptosis by inhibition of BAX.
JO  - Blood
VL  - 105
IS  - 8
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - KIT exon 8 mutations are located in the extracellular portion of the receptor and are strongly associated with core-binding factor (CBF)-acute myeloid leukemia (AML). To characterize the functional role of these mutants, we analyzed the proproliferative and antiapoptotic potential of 3 KIT exon 8 mutations in interleukin 3 (IL-3)-dependent Ba/F3 cells. All KIT exon 8 mutants induced receptor hyperactivation in response to stem cell factor (SCF) stimulation in terms of proliferation and resistance toward apoptotic cell death. A representative KIT exon 8 mutant showed spontaneous receptor dimerization, phosphorylation of mitogen-activated protein kinase (MAPK), and conferred IL-3-independent growth to Ba/F3 cells. MAPK and phosphatidylinositol 3-kinase (PI3-kinase) activation was essential for the phenotype of this mutant. Additionally, imatinib inhibited proliferation of KIT exon 8 mutant-expressing Ba/F3 cells. Our data show that KIT exon 8 mutations represent gain-of-function mutations and might represent a new molecular target for treatment of CBF leukemias.
AU  - Kohl, T.M.
AU  - Schnittger, S.
AU  - Ellwart, J.W.
AU  - Hiddemann, W.*
AU  - Spiekermann, K.
C1  - 4592
C2  - 23227
SP  - 3391
TI  - KIT exon 8 mutations associated with core-binding factor (CBF)-acute myeloid leukemia (AML) cause hyperactivation of the receptor in response to stem cell factor.
JO  - Blood
VL  - 105
IS  - 8
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Granzyme B (GzmB), a serine protease of cytotoxic T lymphocytes and natural killer (NK) cells, induces apoptosis by caspase activation after crossing the plasma membrane of target cells. The mechanism of this translocation during killer cell attack, however, is not understood. Killer cells release GzmB and the membrane-disturbing perforin at the contact site after target recognition. Receptor-mediated import of glycosylated GzmB and release from endosomes were suggested, but the role of the cation-independent mannose 6-phosphate receptor was recently refuted. Using recombinant nonglycosylated GzmB, we observed binding of GzmB to cellular membranes in a cell type–dependent manner. The basis and functional impact of surface binding were clarified. GzmB binding was correlated with the surface density of heparan sulfate chains, was eliminated on treatment of target cells with heparinase III or sodium chlorate, and was completely blocked by an excess of catalytically inactive GzmB or GzmK. Although heparan sulfate–bound GzmB was taken up rapidly into intracellular lysosomal compartments, neither of the treatments had an inhibitory influence on apoptosis induced by externally added streptolysin O and GzmB or by natural killer cells. We conclude that membrane receptors for GzmB on target cells are not crucial for killer cell–mediated apoptosis.
AU  - Kurschus, F.C.*
AU  - Bruno, R.*
AU  - Fellows, E.*
AU  - Falk, C.S.
AU  - Jenne, D.*
C1  - 4550
C2  - 22602
SP  - 2049-2058
TI  - Membrane receptors are not required to deliver granzyme B during killer cell attack.
JO  - Blood
VL  - 105
IS  - 5
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Epstein-Barr virus (EBV) is associated with B-cell lymphomas such as Hodgkin lymphoma, Burkitt lymphoma, and post-transplantation lymphoma, which originate from clonal germinal center (GC) B cells. During the process of somatic hypermutation, GC B cells can acquire deleterious or nonsense mutations in the heavy and light immunoglobulin genes. Such mutations abrogate the cell surface expression of the B-cell receptor (BCR), which results in the elimination of these nonfunctional B cells by immediate apoptosis. EBV encodes several latent genes, among them latent membrane protein 1 (LMP1) and LMP2A, which are regularly expressed in EBV-positive Hodgkin lymphoma and posttransplantation lymphomas. Since LMP1 and LMP2A mimic the function of 2 key receptors on B cells, CD40 and BCR, respectively, we wanted to learn whether EBV infection can rescue proapoptotic GC B cells with crippling mutations in the heavy chain immunoglobulin locus from apoptosis. We show here that BCR-negative GC B cells readily enter the cell cycle upon infection with EBV in vitro and yield clonal lymphoblastoid cell lines that are incapable of expressing a functional BCR because the rearranged and formerly functional heavy chain immunoglobulin alleles carry deleterious mutations. Our findings imply an important role for EBV in the process of lymphomagenesis in certain cases of Hodgkin lymphoma and posttransplantation lymphomas.
AU  - Mancao, C.
AU  - Altmann, M.
AU  - Jungnickel, B.
AU  - Hammerschmidt, W.
C1  - 4645
C2  - 23210
SP  - 4339-4344
TI  - Rescue of "crippled" germinal center B cells from apoptosis by Epstein-Barr virus.
JO  - Blood
VL  - 106
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Fibromodulin (FMOD) was shown to be highly overexpressed in chronic lymphocytic leukemia (CLL) cells compared with normal B lymphocytes by gene expression profiling. Therefore FMOD might serve as potential tumor-associated antigen (TAA) in CLL, enabling expansion of FMOD-specific T cells. In CLL samples derived from 16 different patients, high expression of FMOD by real-time reverse transcriptase–polymerase chain reaction (RT-PCR) was detectable in contrast to normal B lymphocytes. We used unpulsed native CLL cells and CD40 ligand (CD40L)–stimulated CLL cells as antigen-presenting cells (APCs) to expand autologous T cells from 13 patients. The number of T cells during 4 weeks of in vitro culture increased 2- to 3.5-fold and the number of T cells recognizing FMOD peptides bound to HLA-A2 dimers increased 10-fold. The expanded T cells also were able to secrete interferon-γ (IFN-γ) upon recognition of the antigen demonstrated by IFN-γ ELISPOT assays. T cells not only recognized HLA-A2–binding FMOD peptides presented by transporter-associated with antigen-processing (TAP)–deficient T2 cells, but also FMOD overexpressing autologous CLL cells in an HLA-A2–restricted manner. In summary, FMOD was shown for the first time to be naturally processed and presented as TAA in primary CLL cells, enabling the expansion of autologous tumor-specific T cells.
AU  - Mayr, C.
AU  - Bund, D.
AU  - Schlee, M.
AU  - Moosmann, A.*
AU  - Kofler, D.M.
AU  - Hallek, M.
AU  - Wendtner, C.-M.
C1  - 3093
C2  - 22400
SP  - 1566-1573
TI  - Fibromodulin as a novel tumor-associated antigen (TAA)in chronic lymphocytic leukemia (CLL) which allows expansion of specific CD8+ autologous T lymphocytes.
JO  - Blood
VL  - 105
IS  - 4
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - In chronic lymphocytic leukemia (CLL), chromosomes usually evade detailed cytogenetic analyses because cells poorly respond to the traditionally used set of mitogens. We applied novel technologies, such as stimulation of CLL cells either with CD40 ligand or with a combination of CpG-oligodeoxynucleotides and IL-2, to increase the frequency of metaphase spreads for detailed chromosome analysis in 96 patients with CLL. This approach revealed that translocations occurred in 33 of 96 (34%) of our patients with CLL. The presence of translocations defined a new prognostic subgroup because these patients have significantly shorter median treatment-free survival (24 months vs 106 months; P < .001) and significantly inferior overall survival (OS; median, 94 months) than patients without translocations (346 months; P < .001). In multivariate analysis-including Binet stage, complex karyotype, CD38 expression, and 17p deletions-translocation proved to be the prognostic marker with the highest impact for an unfavorable clinical outcome (P < .001). In summary, we identified a new subgroup of patients with CLL defined by chromosomal trans-locations and poor prognosis. Our data may facilitate the identification of molecular events crucial for transforming activity in this disease and should have implications for risk-adapted clinical management of patients with CLL.
AU  - Mayr, C.
AU  - Speicher, M.R.
AU  - Kofler, D.M.*
AU  - Buhmann, R.*
AU  - Strehl, J.*
AU  - Busch, R.*
AU  - Hallek, M.
AU  - Wendtner, C.M.
C1  - 3735
C2  - 28301
SP  - 742-751
TI  - Chromosomal translocations are associated with poor prognosis in chronic lymphocytic leukemia.
JO  - Blood
VL  - 107
IS  - 2
PB  - American Society of Hematology
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Several features of chronic lymphocytic leukemia (CLL) suggest that immune-based strategies may have therapeutic potential. A promising approach is provided by the transduction of CLL cells with CD40 ligand (CD40L) by viral vectors to enhance their immunogenicity. We compared the antigen-presenting capacity of CD40L-transduced CLL cells with mock-transduced or CD40L-stimulated CLL cells (CD40-CLL). A significantly higher number of T cells could be expanded using CD40L-transduced CLL cells as antigen-presenting cells (APCs) compared with the control group (P = .008). Using 5 different CLL-associated tumor antigens, including fibromodulin, MDM2 (murine double minute 2), survivin, p53, and KW-13, we show in interferon-γ (IFN-γ) enzyme-linked immunospot (ELISPOT) assays after 35 days of in vitro culture that the number of antigen-specific autologous T cells was also significantly higher when CD40L-transduced CLL cells were used as APCs (P < .001). Thus, CD40L-transduced CLL cells are able to induce an antigen-specific T-cell response and might be superior to CD40-CLL cells for immune-based therapeutic strategies in CLL.
AU  - Mayr, C.
AU  - Kofler, D.M.*
AU  - Büning, D.
AU  - Bund, D.
AU  - Hallek, M.*
AU  - Wendtner, C.-M.*
C1  - 4643
C2  - 23313
SP  - 3223-3226
TI  - Transduction fo CLL cells by CD40 ligand enhances an antigen-specific immune recognition by autologous T cells.
JO  - Blood
VL  - 106
IS  - 9
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Epstein-Barr virus (EBV) latently infects and immortalizes B lymphocytes and causes lymphoproliferative malignancies. We show here that the EBV nuclear antigen EBNA2 induces expression of the 2 chains of the interleukin-18 receptor (IL-18R) in Burkitt lymphoma (BL) cell lines and in nontransformed B cells. Activation of IL-18R expression by EBNA2 is independent of its interaction with the transcriptional repressor RBPJκ. It occurs in the absence of any other viral protein but requires de novo synthesis of cellular proteins. IL-18R induction is a highly specific function of EBNA2, because neither other EBV latent proteins nor the cellular proteins c-myc or Notch can exert this effect. Using cDNA microarray expression profiling, we find that the IL-18 receptor expressed in EBV-infected BL cells has signaling capacity, because IL-18 significantly modified gene expression. We report that EBNA2 expression is associated with IL-18R expression in vivo in EBV-positive B-lymphomas from AIDS patients.
AU  - Pagés, F.*
AU  - Galon, J.*
AU  - Karaschuk, G.*
AU  - Dudziak, D.
AU  - Camus, M.*
AU  - Lazar, V.*
AU  - Camilleri-Broët, S.*
AU  - Lagorce-Pagés, C.*
AU  - Lebel-Binay, S.*
AU  - Laux, G.
AU  - Fridman, W.H.*
C1  - 2476
C2  - 22568
SP  - 1632-1639
TI  - Epstein-Barr virus nuclear antigen 2 induces interleukin-18 receptor expression in B cells.
JO  - Blood
VL  - 105
IS  - 4
PY  - 2005
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Activating mutations in the juxtamembrane domain (FLT3-length mutations, FLT3-LM) and in the protein tyrosine kinase domain (TKD) of FLT3 (FLT3-TKD) represent the most frequent genetic alterations in acute myeloid leukemia (AML) and define a molecular target for therapeutic interventions by protein tyrosine kinase (PTK) inhibitors. We could show that distinct activating FLT3-TKD mutations at position D835 mediate primary resistance to FLT3 PTK inhibitors in FLT3-transformed cell lines. In the presence of increasing concentrations of the FLT3 PTK inhibitor SU5614, we generated inhibitor resistant Ba/F3 FLT3-internal tandem duplication (ITD) cell lines (Ba/F3 FLT3-ITD-R1-R4) that were characterized by a 7- to 26-fold higher IC50 (concentration that inhibits 50%) to SU5614 compared with the parental ITD cells. The molecular characterization of ITD-R1-4 cells demonstrated that specific TKD mutations (D835N and Y842H) on the ITD background were acquired during selection with SU5614. Introduction of these dual ITD-TKD, but not single D835N or Y842H FLT3 mutants, in Ba/F3 cells restored the FLT3 inhibitor resistant phenotype. Our data show that preexisting or acquired mutations in the PTK domain of FLT3 can induce drug resistance to FLT3 PTK inhibitors in vitro. These findings provide a molecular basis for the evaluation of clinical resistance to FLT3 PTK inhibitors in patients with AML.
AU  - Bagrintseva, K.
AU  - Schwab, R.
AU  - Kohl, T.M.*
AU  - Schnittger, S.
AU  - Eichenlaub, S.
AU  - Ellwart, J.W.
AU  - Hiddemann, W.
AU  - Spiekermann, K.
C1  - 939
C2  - 21965
SP  - 2266-2275
TI  - Mutations in the tyrosine kinase domain of FLT3 define a new molecular mechanism of acquired drug resistance to PTK inhibitors in FLT3-ITD-transformed hematopoitic cells.
JO  - Blood
VL  - 103
IS  - 6
PY  - 2004
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Signal transducer and activator of transcription 1 (STAT1), a transcription factor known to participate in antiviral responses, acts as a tumor suppressor inhibiting cell growth and promoting apoptosis. To study the role of STAT1 in DNA damage–induced apoptosis in B lymphocytes, its active form, STAT1α, was specifically inhibited by the overexpression of STAT1β, the STAT1α truncated inhibitory isoform. An episomal vector with a tetracycline-inducible bidirectional promoter was created to induce the expression of 2 proteins, STAT1β and enhanced green fluorescence protein (EGFP). The same vector was used to overexpress STAT1α as a control. Expression of STAT1β inhibited the phosphorylation, the DNA-binding activity, and the transcriptional activity of STAT1α, as well as the expression of STAT1α target genes such as p21WAF1/CIP1, TAP1, IRF1, and PKR. Inhibiting STAT1α by STAT1β increased the growth rate of transfected cells and their resistance to fludarabine-induced apoptosis and cell cycle arrest. Overexpressing STAT1β reversed the negative regulation of Mdm2 expression observed after treatment with interferon-gamma (IFN-γ), which activates STAT1, or with fludarabine. Nuclear translocation of p53 after fludarabine treatment was decreased when STAT1β was overexpressed, and it was increased when STAT1α was induced. Oligonucleotide pull-down experiments showed a physical STAT1/p53 interaction. Our results show that imbalance between the antiproliferative/proapoptotic isoform STAT1α and the proliferative isoform STAT1β is likely to play a crucial role in the regulation of proliferation and apoptosis and that STAT1α may regulate p53 activity and sensitize B cells to fludarabine-induced apoptosis.
AU  - Baran-Marszak, F.*
AU  - Feuillard, J.*
AU  - Najjar, I.*
AU  - Le Clorennec, Ch.*
AU  - Bechét, J.-P.*
AU  - Dusanter-Fourt, I.*
AU  - Bornkamm, G.W.
AU  - Raphaël, M.*
AU  - Fagard, R.*
C1  - 3783
C2  - 21999
SP  - 2475-2483
TI  - Differential roles of STAT1alpha and STAT1beta in fludarabine-induced cell cycle arrest and apoptosis in human B cells.
JO  - Blood
VL  - 104
IS  - 8
PY  - 2004
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Platelet adhesion and activation at the vascular wall are the initial steps leading to arterial thrombosis and vascular occlusion. Prostacyclin and nitric oxide inhibit platelet adhesion, acting via cyclic adenosine monophosphate (cAMP)– and cyclic guanosine monophosphate (cGMP)–dependent protein kinases. A major downstream target for both cAMP- and cGMP-dependent protein kinases is the vasodilator-stimulated phosphoprotein (VASP). To test the significance of VASP for the regulation of platelet adhesion in vivo, we studied platelet–vessel wall interactions using VASP-deficient (VASP–/–) mice. Under physiologic conditions, platelet adhesion to endothelial cells was significantly enhanced in VASP null mutants when compared with wild-type mice (P < .05). Platelet recruitment in VASP null mice involved P-selectin and the fibrinogen receptor glycoprotein IIb-IIIa (GPIIb-IIIa). Under pathophysiologic conditions, the loss of VASP increased platelet adhesion to the postischemic intestinal microvasculature, to the atherosclerotic endothelium of ApoE-deficient mice, and to the subendothelial matrix following endothelial denudation (P < .05 vs wild type). Importantly, platelet adhesion in VASP null mutants was unresponsive to nitric oxide. These data show for the first time in vivo that VASP is involved in down-regulation of platelet adhesion to the vascular wall under both physiologic and pathophysiologic conditions.
AU  - Massberg, S.*
AU  - Grüner, S.*
AU  - Konrad, I.*
AU  - Arguinzonis, M.I.G.*
AU  - Eigenthaler, M.*
AU  - Hemler, K.*
AU  - Kersting, J.*
AU  - Schulz, C.*
AU  - Müller, I.*
AU  - Besta, F.*
AU  - Nieswandt, B.*
C1  - 2555
C2  - 22279
SP  - 136-142
TI  - Enhanced in vivo platelet adhesion in vasodilator-stimulated phosphoprotein (VASP)-deficient mice.
JO  - Blood
VL  - 103
IS  - 1
PY  - 2004
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Antineutrophil cytoplasmic autoantibodies (ANCAs) recognizing human proteinase 3 of neutrophil granules are a diagnostic hallmark of Wegener granulomatosis, an autoimmune systemic vasculitis with predilection for the respiratory tract and kidneys. In vitro experiments have implicated several mechanisms by which ANCAs may lead to tissue injury. However, little is known about the pathogenic significance of proteinase 3–specific antibodies in vivo. In vivo models for ANCA-mediated proinflammatory effects have not been forthcoming, primarily because ANCA epitopes on human proteinase 3 are not shared by the murine homolog. In this study we generated ANCAs against recombinant murine proteinase 3 in proteinase 3/neutrophil elastase–deficient mice that recognized the murine antigen on the surface of neutrophils. Local inflammation induced by intradermal injection of tumor necrosis factor α triggered a stronger subcutaneous panniculitis in the presence of passively transferred systemic proteinase 3–ANCAs than in the presence of mock immune serum. When we transferred mouse proteinase 3-ANCA serum to systemically lipopolysaccharide-primed wild-type mice, mice treated with proteinase 3-ANCAs did not develop significantly stronger signs of inflammation of the lungs or kidneys than the respective mock immune serum-treated animals. In conclusion, our in vivo study provides the first evidence for a pathogenic effect of proteinase 3–specific ANCAs at local sites of inflammation.
AU  - Pfister, H.*
AU  - Ollert, M.
AU  - Fröhlich, L.F.*
AU  - Quintanilla-Martinez, L.
AU  - Colby, T.V.*
AU  - Specks, U.*
AU  - Jenne, D.*
C1  - 2696
C2  - 22235
SP  - 1411-1418
TI  - Antineutrophil cytoplasmic autoantibodies against the murine homolog of proteinase 3 (Wegener autoantigen) are pathogenic in vivo.
JO  - Blood
VL  - 104
IS  - 5
PY  - 2004
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The t(11;14)(q13;q32) is the most com-
      
      	mon translocation in multiple myeloma
      
      	(MM), resulting in up-regulation of cyclin
      
      	D1. We used a segregation fluorescence
      
      	in situ hybridization (FISH) assay to de-
      
      	tect t(11;14) breakpoints in primary MM
      
      	cases and real-time reverse transcriptase–
      
      	polymerase chain reaction (RT-PCR) to
      
      	quantify cyclin D1 and MYEOV (myeloma
      
      	overexpressed) expression, another puta-
      
      	tive oncogene located on chromosome
      
      	11q13. High levels of cyclin D1 mRNA
      
      	(cyclin D1/TBP [TATA box
      
      	binding pro-
      
      	tein] ratio
      
      	>
      
      	95) were found exclusively in
      
      	the presence of a t(11;14) translocation
      
      	(11/48 cases;
      
      	P
      
      	<
      
      	.00001). In addition, a
      
      	subgroup of MM cases (15/48) with inter-
      
      	mediate to low cyclin D1 mRNA (cyclin
      
      	D1/TBP ratio between 2.3 and 20) was
      
      	identified. FISH analysis ruled out a t(11;
      
      	14) translocation and 11q13 amplification
      
      	in these cases; however, in 13 of 15
      
      	patients a chromosome 11 polysomy was
      
      	demonstrated (
      
      	P
      
      	<
      
      	.0001). These results
      
      	indicate an effect of gene dosage as an
      
      	alternative mechanism of cyclin D1 de-
      
      	regulation in MM. The absence of chromo-
      
      	some 11 abnormalities in 2 of 15 patients
      
      	with intermediate cyclin D1 expression
      
      	supports that there are presumably other
      
      	mechanism(s) of cyclin D1 deregulation
      
      	in MM patients. Our data indicate that
      
      	deregulation of MYEOV is not favored in
      
      	MM and further strengthens the role of
      
      	cyclin D1 overexpression in lymphoid ma-
      
      	lignancies with a t(11;14)(q13;q32) trans-
      
      	location.
AU  - Specht, K.
AU  - Haralambieva, E.*
AU  - Bink, K.
AU  - Kremer, M.
AU  - Mandl-Weber, S.*
AU  - Koch, I.*
AU  - Tomer, R.*
AU  - Höfler, H.
AU  - Schuuring, E.*
AU  - Kluin, P.M.*
AU  - Fend, F.*
C1  - 5066
C2  - 21937
SP  - 1120-1126
TI  - Different mechanisms of cyclin D1 overexpression in multiple myeloma revealed by fluorescence in situ hybridization and quantitative analysis of mRNA levels.
JO  - Blood
VL  - 104
IS  - 4
PY  - 2004
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Katzenberger, T.*
AU  - Lohr, A.*
AU  - Schwarz, S.*
AU  - Dreyling, M.
AU  - Schoof, J.*
AU  - Nickenig, C.*
AU  - Stilgenbauer, St.*
AU  - Kalla, J.*
AU  - Ott, M.M.*
AU  - Müller-Hermelink, H.K.*
AU  - Ott, G.*
C1  - 10327
C2  - 21600
SP  - 699-702
TI  - Genetic analysis of de novo CD5+ diffuse large B-cell lymphomas suggests an origin from a somatically mutated CD5+ progenitor B cell.
JO  - Blood
VL  - 101
PY  - 2003
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - There is a strong graft-versus-leukemia (GVL) effect of allogeneic stem cell trans- plantation (SCT) due to elimination of tumor cells by alloimmune effector lym- phocytes. When leukemia relapses after allogeneic SCT, donor lymphocyte trans- fusions (DLTs) can induce sustained re- missions in some patients. This review summarizes the current status on clinical use of DLT, the basis of GVL reactions, problems associated with this therapy, and new strategies to improve DLT. Sev- eral multicenter surveys demonstrated that the GVL effect of DLT is most effec- tive in chronic myelogenous leukemia (CML), whereas it is less pronounced in acute leukemia and myeloma. Cytokine stimulation to induce differentiation of myeloid progenitor cells or to up-regulate costimulatory molecules on tumor cells may improve the efficacy of DLT. Infec- tions and graft-versus-host disease (GVHD) are major complications of DLT. Control of GVHD may be improved using suicide gene–modified T cells for DLT, allowing T-cell elimination if severe GVHD develops. Hopefully, in the future, GVL effect can be separated from GVHD through adoptive transfer of selected T cells that recognize leukemia-specific an- tigens or minor histocompatibility anti- gens, which are expressed predominantly on hematopoietic cells, thereby preclud- ing attack of normal tissues. In patients with leukemia and lymphomas with fast progression, tumor growth may outpace development of effector T cells. Here it may be preferable to select stem cell transplant donors with HLA-mismatches that allow alloreactive natural killer cells, which appear early after transplantation, to retain their cytolytic function. New ap- proaches for adoptive immune therapy of leukemia, which promise a better progno- sis for these patients, are being devel- oped.
AU  - Kolb, H.-J.
AU  - Schmid, C.
AU  - Barrett, A.J.*
AU  - Schendel, D.J.
C1  - 10329
C2  - 21657
SP  - 767-776
TI  - Graft-versus-leukemia reactions in allogeneic chimeras.
JO  - Blood
VL  - 103
IS  - 3
PY  - 2003
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - p27 is a cyclin-dependent kinase inhibitor that plays a critical role in regulating G1/S progression, and whose activity is, in part, regulated through interactions with D-type cyclins. Mantle cell lymphoma (MCL) is characterized by the t(11;14) translocation resulting in deregulated cyclin D1. We previously showed that p27 expression in MCL, as assessed by immunohistochemistry (IHC), does not show the usual inverse relationship to proliferate seen in most other lymphomas that do not overexpress cyclin D1. This suggested that the normal expression or control of p27 activity on cell growth might be altered through potential interactions with cyclin D1. Using Western blot and coimmunoprecipitation studies, we assessed the interrelationship between cyclin D1 and p27 in several cyclin D1+ cell lines and primary MCL cases. Similar to our previous results by IHC, typical MCLs showed lower expression of p27 when compared to the more highly proliferative blastic cases or cell lines (mean arbitrary units: 58 versus 236 versus 120). Cyclin D1 was expressed at variable levels in both typical and blastic MCLs. p27 protein could be consistently coimmunoprecipitated with cyclin D1 from both cell lines and cases. Using techniques of exhaustive immunoprecipitation, we could demonstrate that most p27 protein was sequestered into complexes containing cyclin D1. We hypothesize that mantle cell lymphomagenesis results not only from direct consequences of inappropriate cyclin D1 expression, but also from the ability of overexpressed cyclin D1 to buffer physiologic changes in p27 levels, thereby rendering p27 ineffective as an inhibitor of cellular growth.
AU  - Quintanilla-Martinez, L.
AU  - Davies-Hill, T.*
AU  - Fend, F.
AU  - Calzada-Wack, J.*
AU  - Sorbara, L.*
AU  - Campo, E.*
AU  - Jaffe, E.S.*
AU  - Raffeld, M.*
C1  - 10328
C2  - 20935
SP  - 3181-3187
TI  - Sequestration of p27Kip1 protein by cyclin D1 in typical and blastic variants of mantle cell lymphoma (MCL) : Implications for pathogenesis.
JO  - Blood
VL  - 101
IS  - 8
PY  - 2003
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The clinical progression of chronic myeloid leukemia (CML) from chronic phase to blast crisis is characterized by the increasing failure of myeloid precursors to differentiate into mature granulocytes. This study was undertaken to investigate the influence of Bcr-Abl and of the small molecule Abl tyrosine–kinase inhibitor imatinib mesylate on granulocyte colony-stimulating factor (G-CSF)–induced neutrophilic differentiation. We show that differentiation of 32Dcl3 cells into mature granulocytes is accompanied by the increased expression of the antigens macrophage adhesion molecule–1 (Mac-1) and Gr-1, of the G-CSF receptor (G-CSFR), of myeloid transcription factors (CCAAT/enhancer-binding protein–α [C/EBPα], C/EBPε, and PU.1), and of the cyclin-dependent kinase inhibitor p27Kip1. In 32Dcl3 cells transfected with thebcr-abl gene (32DBcr-Abl), G-CSF did not trigger either granulocytic differentiation or the up-regulation of C/EBPα, C/EBPε, and the G-CSFR. This could be correlated to a defect in c-Myc down-regulation. In contrast, the up-regulation of PU.1 and p27Kip1 by G-CSF was not affected by Bcr-Abl. Importantly, incubation of 32DBcr-Ablwtcells with the kinase inhibitor imatinib mesylate prior to G-CSF stimulation completely neutralized the effects of Bcr-Abl on granulocytic differentiation and on C/EBPα and C/EBPε expression. Taken together, the results suggest that the Bcr-Abl kinase induces a reversible block of the granulocytic differentiation program in myeloid cells by disturbing regulation of hematopoietic transcription factors such as C/EBPα and C/EBPε.
AU  - Schuster, C.*
AU  - Forster, K.
AU  - Dierks, H.*
AU  - Elsässer, A.
AU  - Behre, G.
AU  - Simon, N.
AU  - Danhauser-Riedl, S.*
AU  - Hallek, M.
AU  - Warmuth, M.
C1  - 22298
C2  - 21102
SP  - 655-663
TI  - The effects of Bcr-Abl on C/EBP transcription-factor regulation and neutrophilic differentiation are reversed by the Abl kinase inhibitor imatinib mesylate.
JO  - Blood
VL  - 101
IS  - 2
PY  - 2003
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The leukemogenic tyrosine kinase Bcr-Abl contains a highly conserved inhibitor-binding pocket (IBP), which serves as a binding site for imatinib mesylate. Mutations at the IBP may lead to resistance of the Abl kinase against imatinib mesylate. To examine the mechanisms of imatinib mesylate binding and resistance in more detail, we created several point mutations at amino acid positions 315 and 380 of Abl, blocking the access to the IBP and rendering Bcr-Abl imatinib mesylate–resistant. Moreover, introduction of a mutation destabilizing the inactive conformation of Abl (Asp276Ser/Glu279Ser) also led to imatinib mesylate resistance, suggesting that the inhibitor required inactivation of the kinase prior to binding. These Bcr-Abl mutants were then used to evaluate the binding mode and specificity of 2 compounds, PP1 and CGP76030, originally characterized as Src kinase inhibitors. Both compounds inhibited Bcr-Abl in a concentration-dependent manner by overlapping binding modes. However, in contrast to imatinib mesylate, PP1 and CGP76030 blocked cell growth and survival in cells expressing various inhibitor-resistant Abl mutants. Studies on the potential signaling mechanisms demonstrated that in cells expressing inhibitor-resistant Bcr-Abl mutants, PP1 and CGP76030 inhibited the activity of Src family tyrosine kinases and Akt but not signal transducer and activator of transcription–5 (STAT5) and JUN kinase (Jnk). The results suggest that the use of Src kinase inhibitors is a potential strategy to prevent or overcome clonal evolution of imatinib mesylate resistance in Bcr-Abl+ leukemia.
AU  - Warmuth, M.
AU  - Simon, N.
AU  - Mitina, O.
AU  - Mathes, R.*
AU  - Fabbro, D.*
AU  - Manley, P.W.*
AU  - Buchdunger, E.*
AU  - Forster, K.
AU  - Moarefi, I.*
AU  - Hallek, M.
C1  - 22299
C2  - 21103
SP  - 664-672
TI  - Dual-specific Src and Abl kinase inhibitors, PP1 and CGP76030, inhibit growth and survival of cells expressing imatinib mesylate-resistant Bcr-Abl kinases.
JO  - Blood
VL  - 101
IS  - 2
PY  - 2003
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Trioma cell vaccination is a potent new immunologic approach for the therapy of malignant B-cell lymphoma. It is based on targeting tumor antigens to internalizing receptors on antigen-presenting cells (APCs). Tumor cells are fused to an APC-specific hybridoma, where they are converted to trioma cells that include potentially all lymphoma-derived antigens and that express the APC-binding arm. In this study, the mechanisms of, trioma-mediated tumor immunity in immunocompetent mice were dissected, and it was shown in this model system that humoral anti-idiotypic immunity is indeed detectable after idiotype-specific immunization but that it does not reflect the degree of tumor protection obtained in vivo. Immunization against the idiotype alone was not sufficient for efficient tumor rejection in vivo. Targeting tumor antigens to APCs Is only successful in terms of inducing tumor protection when designed as a polyvalent vaccination protocol.
AU  - Kronenberger, K.
AU  - Diekmann, A.*
AU  - Selmayr, M.*
AU  - Strehl, J.
AU  - Wahl, U.
AU  - Lindhofer, H.
AU  - Kraal, G.*
AU  - Mocikat, R.
C1  - 10330
C2  - 20166
SP  - 1327-1331
TI  - Impact of the lymphoma idiotype on in vivo tumor protection in a vaccination model based on targeting antigens to antigen-presenting cells.
JO  - Blood
VL  - 99
PB  - American Society of Hematology
PY  - 2002
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Lymphoblastoid cell lines (LCLs) are human B cells latently infected and immortalized by Epstein-Barr virus (EBV). Presenting viral antigens, they efficiently induce EBV-specific T-cell responses in vitro. Analogous ways to generate T-cell cultures specific for other antigens of interest are highly desirable. Previously, we constructed a mini-EBV plasmid that consists of less than half the EBV genome, is unable to cause virus production, but still immortalizes B cells in vitro. Mini-EBV–immortalized B-cell lines (mini-LCLs) are efficiently produced by infection of B cells with viruslike particles carrying only mini-EBV DNA. Mini-EBV plasmids can be engineered to express an additional gene in immortalized B cells. Here we present a mini-EBV coding for a potent CD8+ T-cell antigen, the matrix phosphoprotein pp65 of human cytomegalovirus (CMV). By means of this pp65 mini-EBV, pp65-expressing mini-LCLs could be readily established from healthy donors in a one-step procedure. We used these pp65 mini-LCLs to reactivate and expand effector T cells from autologous peripheral blood cells in vitro. When generated from cytomegalovirus (CMV)–seropositive donors, these effector T-cell cultures displayed strong pp65-specific HLA-restricted cytotoxicity. A large fraction of CD8+ T cells with pp65 epitope specificity was present in such cultures, as demonstrated by direct staining with HLA/peptide tetramers. We conclude that the pp65 mini-EBV is an attractive tool for CMV-specific adoptive immunotherapy. Mini-EBVs could also facilitate the generation of T cells specific for various other antigens of interest.
AU  - Moosmann, A.*
AU  - Khan, N.*
AU  - Cobbold, M.*
AU  - Zentz, C.
AU  - Delecluse, H.-J.*
AU  - Hollweck, G.
AU  - Hislop, A.D.*
AU  - Blake, N.W.*
AU  - Croom-Carter, D.*
AU  - Wollenberg, B.
AU  - Moss, P.A.H.*
AU  - Zeidler, R.*
AU  - Rickinson, A.B.*
AU  - Hammerschmidt, W.
C1  - 21981
C2  - 20507
SP  - 1755-1764
TI  - B cells immortalized by a mini-Epstein-Barr virus encoding a foreign antigen efficiently reactive specific cytotoxic T cells.
JO  - Blood
VL  - 100
PY  - 2002
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Activating length mutations in the juxtamembrane (JM) domain of the FLT3 gene (FLT3-LM) and mutations in the catalytic domain (FLT3D835/836) of this receptor tyrosine kinase represent the most frequent genetic alterations in acute myeloid leukemia (AML). Here, we describe a 6-bp insertion in the activation loop of FLT3 between codons 840 and 841 of FLT3 (FLT3-840GS) in 2 unrelated patients with AML. Screening for other activating mutations of FLT3, KIT, and NRASshowed no further genetic alterations in patients carrying the FLT3-840GS. In functional analyses we could show that this mutant is hyperphosphorylated on tyrosine and confers interleukin 3–independent growth to Ba/F3 cells, which can be inhibited by a specific FLT3 protein tyrosine kinase (PTK) inhibitor. Our results show for the first time that in addition to known mutations in the JM and the catalytic domain, further activating length mutations exist in theFLT3 gene.
AU  - Spiekermann, K.
AU  - Bagrintseva, K.
AU  - Schoch, C.*
AU  - Haferlach, T.*
AU  - Hiddemann, W.
AU  - Schnittger, S.*
C1  - 22230
C2  - 20954
SP  - 3423-3425
TI  - A new and recurrent activating length mutation in exon 20 of the FLT3 gene in acute myeloid leukemia.
JO  - Blood
VL  - 100
PY  - 2002
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - B cells of chronic lymphocytic leukemia (B-CLL) are resistant to transduction with most currently available vector systems. Using an optimized adenovirus-free packaging system, recombinant adeno-associated virus (rAAV) vectors coding for the enhanced green fluorescent protein (AAV/EGFP) and CD40 ligand (AAV/CD40L) were packaged and highly purified resulting in genomic titers up to 3 × 1011/mL. Cells obtained from 24 patients with B-CLL were infected with AAV/EGFP or AAV/CD40L at a multiplicity of infection (MOI) of 100 resulting in transgene expression in up to 97% of cells as detected by flow cytometry 48 hours after infection. Viral transduction could be specifically blocked by heparin. Transduction with AAV/CD40L resulted in up-regulation of the costimulatory molecule CD80 not only on infected CLL cells but also on noninfected bystander leukemia B cells, whereas this effect induced specific proliferation of HLA-matched allogeneic T cells. Vaccination strategies for patients with B-CLL using leukemia cells infected ex vivo by rAAV vectors now seems possible in the near future.
AU  - Wendtner, C.-M.
AU  - Kofler, D.M.
AU  - Theiss, H.D.
AU  - Kurzeder, C.
AU  - Buhmann, R.*
AU  - Schweighofer, C.*
AU  - Perabo, L.*
AU  - Danhauser-Riedl, S.*
AU  - Baumert, J.J.
AU  - Hiddemann, W.
AU  - Hallek, M.
AU  - Buning, H.*
C1  - 22014
C2  - 20565
SP  - 1655-1661
TI  - Efficient gene transfer of CD40 ligand into primary B-CLL cells using recombinant adeno- associated virus (rAAV) vectors.
JO  - Blood
VL  - 100
PY  - 2002
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Bacterial lipopolysaccharide (LPS, endotoxin) is a ubiquitous component of dust and air pollution and is suspected to contribute after inhalation to an activation of eosinophils in bronchial tissues of asthmatic patients, provoking inflammatory and allergic processes. We were therefore interested in the interaction of eosinophil granulocytes with LPS and have examined the activation of and uptake to human peripheral blood eosinophils by LPS. Eosinophils were stimulated by LPS and the endotoxic component lipid A and the release of tumor necrosis factor alpha (TNF-α) and of the eosinophil-specific granule protein eosinophil cationic protein (ECP) was estimated. The results show induction of TNF-α and ECP-release by LPS and lipid A in a dose-dependent manner. Anti-CD14 monoclonal antibody (moAb) (clone MEM-18) and the synthetic lipid A partial structure 406 blocked the release of TNF-α and ECP by LPS-stimulated eosinophils. Studies with radioactively labeled LPS showed dose-dependent uptake of3H-LPS to eosinophils. The 3H-LPS uptake was found to be specific because preincubation with unlabeled LPS, compound 406 and also anti-CD14 antibodies inhibited uptake of3H-LPS to eosinophil granulocytes. By flow cytometry using anti-CD14 moAb and by reverse transcriptase-polymerase chain reaction (RT-PCR) technique, CD14 expression was detectable. Furthermore, messenger RNA (mRNA) expression of Toll-like receptors (TLR) 2 and TLR 4 was detected, indicating the presence of these CD14 coreceptors. The results indicate that eosinophils can take up LPS and can be stimulated by LPS in a CD14-dependent manner. Hence, in addition to allergens, eosinophils interact with endotoxin, a process that possibly exacerbates ongoing inflammatory and allergic processes.
AU  - Plötz, S.G.*
AU  - Lentschat, A.*
AU  - Behrendt, H.
AU  - Plötz, W.*
AU  - Hamann, L.*
AU  - Ring, J.*
AU  - Rietschel, E.T.*
AU  - Flad, H.-D.*
AU  - Ulmer, A.*
C1  - 21859
C2  - 20063
SP  - 235--241
TI  - The interaction of human peripheral blood eosinophils with bacterial lipopolysaccharide is CD14 dependent.
JO  - Blood
VL  - 97
PY  - 2001
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Bispecific antibodies (bsAbs) can efficiently mediate tumor cell killing by redirecting preactivated or costimulated T cells to disseminated tumor cells, especially in a minimal residual disease situation. This study demonstrates that the trifunctional bsAb BiLu is able to kill tumor cells very efficiently without any additional costimulation of effector cells in vitro and in vivo. Remarkably, this bsAb also induces a long-lasting protective immunity against the targeted syngeneic mouse tumors (B16 melanoma and A20 B-cell lymphoma, respectively). A strong correlation was observed between the induction of a humoral immune response with tumor-reactive antibodies and the survival of mice. This humoral response was at least in part tumor specific as shown in the A20 model by the detection of induced anti-idiotype antibodies. Both the survival of mice and antitumor titers were significantly diminished when F(ab')2 fragments of the same bsAb were applied, demonstrating the importance of the Fc region in this process. With the use of T-cell depletion, a contribution of a cellular antitumor response could be demonstrated. These results reveal the necessity of the Fc region of the bsAb with its potent imrnunoglobulin subclass combination mouse immunoglobulin G2a (IgG2a) and rat IgG2b. The antigen-presenting system, seems to be crucial for achieving an efficient tumor cell killing and induction of long-lasting antitumor immunity. Hereby, the recruitment and activation of accessory cells by the intact bsAb is essential.
AU  - Ruf, P.
AU  - Lindhofer, H.G.
C1  - 33185
C2  - 35638
SP  - 2526-2534
TI  - Induction of a long-lasting antitumor immunity by a trifunctional bispecific antibody.
JO  - Blood
VL  - 98
IS  - 8
PY  - 2001
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Alterations in the vascular system and the onset of angioproliferative lesions such as Kaposi's sarcoma (KS) are common traits of human immunodeficiency virus-1 (HIV-1)-infected patients. To investigate possible factors involved in acquired immunodeficiency syndrome (AIDS)-associated vasculopathy and vascular malfunction, expression of vascular endothelial cell growth factor-A (VEGF-A) was analyzed in HUT 78 T lymphocytes upon infection with HIV-1. VEGF-A was found to be increased in supernatants from infected cells as compared with uninfected cells. In addition, VEGF-A mRNA expression and protein secretion were significantly increased in HUT 78 cells incubated with conditioned medium (CM) derived from HIV-1 chronically infected HUT 78 cells (HIV-TCM) as compared with CM from uninfected cells (TCM). Increase of VEGF-A production in T cells was promoted by inflammatory cytokines (IC) present in HIV-TCM, including tumor necrosis factor alpha (TNFalpha), interferon gamma (IFNgamma), interleukin-1beta (IL-1beta), and IL-6. These IC that have been shown to be increased in sera of HIV-1-infected patients and to be increased by HIV-1 infection or cell activation in these individuals as well as HIV-TCM also increased VEGF-A expression in primary T lymphocytes. Consistent with this, VEGF-A concentrations were found to be higher in sera of HIV-1-infected patients with (mean, 357.1 +/- 197.9 pg/mL) and without KS (mean, 256.7 +/- 137.5 pg/mL) as compared with uninfected individuals (mean, 188.6 +/- 91.7 pg/mL). These data suggest that increased secretion of VEGF-A by T lymphocytes of HIV-1-infected individuals may induce vascular leakage and stimulate proliferation of vascular endothelial cells, which are hallmarks of AIDS-associated vasculopathy and especially of KS development.
AU  - Ascherl, G.*
AU  - Hohenadl, C.
AU  - Schatz, O.*
AU  - Shumay, E.*
AU  - Bogner, J.*
AU  - Eckhart, L.*
AU  - Tschachler, E.
AU  - Monini, P.*
AU  - Ensoli, B.*
AU  - Stürzl, M.
C1  - 21382
C2  - 19500
SP  - 4232-4241
TI  - Infection with human immunodeficiency virus-1 increases expression of vascular endothelial cell growth factor in T-cells: Implications for acquired immunodeficiency syndrome-associated vasculopathy.
JO  - Blood
VL  - 93
IS  - 12
PY  - 1999
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Heparan sulfate (HS) proteoglycans of bone marrow (BM) stromal cells and their extracellular matrix are important components of the microenvironment of hematopoietic tissues and are involved in the interaction of hematopoietic stem and stromal cells. Although previous studies have emphasized the role of HS proteoglycan synthesis by BM stromal cells, we have recently shown that the human hematopoietic progenitor cell line TF-1 also expressed an HS proteoglycan. Immunochemical, reverse transcriptase-polymerase chain reaction (RT-PCR), and Northern blot analysis of this HS proteoglycan showed that it was not related to the syndecan family of HS proteoglycans or to glypican. To answer the question of whether the expression of HS proteoglycans is associated with the differentiation state of hematopoietic progenitor cells, we have analyzed the proteoglycan synthesis of several murine and human hematopoietic progenitor cell lines. Proteoglycans were isolated from metabolically labeled cells and purified by several chromatographic steps. Isolation and characterization of proteoglycans from the cell lines HEL and ELM-D, which like TF-1 cells have an immature erythroid phenotype, showed that these cells synthesize the same HS proteoglycan, previously detected in TF-1 cells, as a major proteoglycan. In contrast, cell lines of the myeloid lineage, like the myeloblastic/promyelocytic cell lines B1 and B2, do not express HS proteoglycans. Taken together, our data strongly suggest that expression of this HS proteoglycan in hematopoietic progenitor cell lines is associated with the erythroid lineage. To prove this association we have analyzed the proteoglycan expression in the nonleukemic multipotent stem cell line FDCP-Mix-A4 after induction of erythroid or granulocytic differentiation. Our data show that HS proteoglycan expression is induced during early erythroid differentiation of multipotent hematopoietic stem cells. In contrast, during granulocytic differentiation, no expression of HS proteoglycans was observed.
AU  - Drzeniek, Z.*
AU  - Stöcker, G.*
AU  - Siebertz, B.*
AU  - Just, U.
AU  - Schroeder, T.
AU  - Ostertag, W.*
AU  - Haubeck, H.D.*
C1  - 23483
C2  - 31420
SP  - 2884-2897
TI  - Heparan sulfate proteoglycan expression is induced during early erythroid differentiation of multipotent hematopoietic stem cells.
JO  - Blood
VL  - 93
IS  - 9
PB  - American Society of Hematology
PY  - 1999
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Interleukin-12 (IL-12) and interferon-gamma (IFN-gamma) exert protective effects during experimental endotoxemia through upregulation of cellular immunity and phagocytic functions. They are part of a positive regulatory feedback loop that enhances the production of the other. Because critically ill patients show a marked suppression of T-cell and macrophage functions with a high susceptibility to infection, potential defects in the immunity/inflammation upregulating IL-12 IFN-gamma pathway were studied. As an ex vivo model of endotoxemia, lipopolysaccharide (LPS) stimulated whole blood from 25 critically ill patients and 12 healthy individuals was incubated with either recombinant human (rh) IL-12 or rhIFN-gamma, respectively. IFN-gamma dose-dependently (P < .05) increased the release of IL-12 p40 and p70 into LPS-stimulated whole blood from healthy humans without effect in whole blood from critically ill patients. RhIL-12 p70 enhanced (P < .05) the secretion of IFN-gamma in controls, while it was ineffective in LPS-stimulated whole blood from critically ill patients. The observed inhibition of the IL-12 IFN-gamma pathway is not specific to LPS, since Staphylococcus aureus Cowan strain I (SAC)-stimulated whole blood from critically ill patients showed similar suppression. The secretion of IL-12 and IFN-gamma was less reduced in critically ill patients when using isolated cultures of adherent cells or lymphocytes. Although preculture of whole blood from healthy humans with IL-10, but not with IL-4, mimicked suppression of the IL-12 IFN-gamma pathway similar to that observed during critical illness, the release of antiinflammatory reacting cytokines (IL-4, IL-10, transforming growth factor [TGF]-beta 1) was decreased into LPS-stimulated whole blood from critically ill patients. These results indicate at least two mechanisms responsible for dramatic disturbances of the IL-12 IFN-gamma pathway during critical illness: (1) deactivation of IL-12 and IFN-gamma producing leukocytes in vivo early after the primary insult, and (2) presence of serum suppressive factors different from IL-4, IL-10, or TGF-beta 1. Because IL-12 and IFN-gamma upregulate essential immune functions, the marked inhibition of IL-12 and IFN-gamma release may be pivotal for high susceptibility of critically ill patients to infection.
AU  - Ertel, W.*
AU  - Keel, M.*
AU  - Neidhardt, R.*
AU  - Steckholzer, U.*
AU  - Kremer, J.-P.
AU  - Ungethuem, U.*
AU  - Trentz, O.*
C1  - 24129
C2  - 31452
SP  - 1612-1620
TI  - Inhibition of the defense system stimulating interleukin-12 interferon-γ pathway during critical illness.
JO  - Blood
VL  - 89
IS  - 5
PB  - WB Saunders
PY  - 1997
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Virally infected cells degrade intracellular viral proteins proteolytically and present the resulting peptides in association with major histocompatibility complex (MHC) class I molecules to CD8+ cytotoxic T lymphocytes (CTLs). These cells are normally prone to CTL-mediated elimination. However, several viruses have evolved strategies to avoid detection by the immune system that interfere with the pathway of antigen presentation. Epstein-Barr virus (EBV) expresses a predominantly late protein, the BCRF1 gene product vIL-10, that is similar in sequence to the human interleukin-10 (hIL-10). We show here that vIL-10 affects the expression of one of the two transporter proteins (TAPs) associated with antigen presentation. Similarly, hIL-10 showed the same activity. Expression of the LMP2 and TAP1 genes but not expression of TAP2 or LMP7 is efficiently downregulated, indicating a specific IL-10 effect on the two divergently transcribed TAP1 and LMP2 genes. Downregulation of TAP1 by IL-10 hampers the transport of peptide antigens into the endoplasmatic reticulum, as shown in the TAP-specific peptide transporter assay, their loading onto empty MHC I molecules, and the subsequent translocation to the cell surface. As a consequence, IL-10 causes a general reduction of surface MHC I molecules on B lymphocytes that might also affect the recognition of EBV-infected cells by cytotoxic T cells.
AU  - Zeidler, R.
AU  - Eissner, G.*
AU  - Meissner, P.*
AU  - Uebel, S.*
AU  - Tampé, R.*
AU  - Lazis, S.
AU  - Hammerschmidt, W.
C1  - 27624
C2  - 32772
SP  - 2390-2397
TI  - Downregulation of TAP1 in B lymphocytes by cellular and Epstein-Barr virus-encoded interleukin-10.
JO  - Blood
VL  - 90
IS  - 6
PY  - 1997
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Using animal models or healthy volunteers, injection of lipopolysaccharide (LPS) or bacteria causes activation of macrophages with excessive synthesis and secretion of proinflammatory cytokines. Although these models mimic the effects of LPS in the host, they may represent more of an experimental expression of endotoxemia than natural infection itself. Therefore, as an ex vivo model of sepsis, whole blood from 15 patients with severe sepsis and 20 control patients without infection was stimulated with LPS to study the kinetics of mRNA expression and release of proinflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1 beta, and IL-6. Stimulation of whole blood with 1 microgram/mL LPS resulted in a maximum increase of cytokine secretion in the control group, while a marked (P < .01) depression of TNF-alpha, IL-1 beta, and IL-6 release was observed in the septic group, which persisted up to 10 days after study enrollment. While IL-1 beta mRNA expression was similar in peripheral blood mononuclear cells (PBMCs) harvested from LPS-stimulated whole blood in septic and control patients, the half-life and consequently the expression of TNF-alpha and IL-6 mRNA were strongly reduced in the septic group. These data indicate a downregulatory mechanism of cytokine release in whole blood from patients with severe sepsis that occurs on different levels. Although excessive secretion of proinflammatory cytokines has been considered deleterious for the host, the reduced capacity of PBMCs in whole blood from septic patients to synthesize and secrete proinflammatory cytokines to an inflammatory stimulus may result in immunodeficiency, because these cytokines in low concentrations are involved in the upregulation of essential cellular and humoral immune functions.
AU  - Ertel, W.*
AU  - Kremer, J.-P.
AU  - Kenney, J.*
AU  - Steckholzer, U.*
AU  - Jarrar, D.*
AU  - Trentz, O.*
AU  - Schildberg, F.W.*
C1  - 24153
C2  - 31473
SP  - 1341-1347
TI  - Downregulation of proinflammatory cytokine release in whole blood from septic patients.
JO  - Blood
VL  - 85
IS  - 5
PB  - Amer. Soc. Hematology
PY  - 1995
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Burkitt lymphoma (BL) and immunoblastic lymphoma (IL) are the most frequent lymphoid tumors encountered in human immunodeficiency virus (HIV)-infected patients. Tumors with a morphology intermediate between BL and IL, and the existence of Burkitt's type translocations in some IL cases makes the classification of these tumors sometimes unclear. We have studied 14 cases of BL and IL in HIV-seropositive individuals with regard to clonality, Epstein-Barr virus (EBV) association, and the presence of c-myc rearrangement. Of seven tumors with morphology of BL, all were monoclonal, six showed a c-myc rearrangement and four were associated with EBV. Five tumors with morphology of IL were associated with EBV and devoid of c-myc rearrangement. Three were polyclonal representing EBV-driven lymphoproliferations similar to those observed in transplant recipients. Two tumors, one with a morphology of IL and the other intermediate between IL and BL were monoclonal, associated with EBV, and harbored a c-myc rearrangement. We propose that these last two tumors represent cases of BL that have adopted an immunoblastic morphotype in the context of acquired immunodeficiency syndrome (AIDS), reflecting the morphologic evolution of Burkitt lymphoma cells observed in culture.
AU  - Delecluse, H.J.
AU  - Raphaël, M.M.*
AU  - Magaud, J.P.*
AU  - Felman, P.*
C1  - 40244
C2  - 40021
SP  - 552-563
TI  - Variable morphology of human immunodeficiency virus-associated lymphomas with C-MYC rearrangements.
JO  - Blood
VL  - 82
IS  - 2
PY  - 1993
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Surprisingly little graft-versus-host disease (GVHD) has been observed in severe combined immunodeficient (SCID) mice injected intraperitoneally (IP) with human blood lymphocytes (hu-PBL-SCID), which raised the question as to whether GVHD in such a distant species is sporadic or suppressed because of immunologic reasons. After screening for blood T-cell chimerism, we hereby describe generalized lethal xenogeneic human GVHD in unconditioned SCID chimeras, which resembles GVHD in SCID mice injected with allogeneic lymphocytes. We adapted an immunocytochemical slide method for minute cell numbers, which allowed us to follow, by multimarker phenotyping of weekly mouse-tail bleeds, the chimeric status of 100 hu-PBL-SCID injected with 107 or 108 hu-PBL of Epstein-Barr virus- (EBV-) donors. More than half of the mice showed no or less than 2% T cells. However, 13% to 21% developed substantial blood T-lymphocyte chimerism (10% to 80% human CD+ cells) and high mortality. Immunohistology showed more human CD8+ than CD4+ T cells in the splenic white pulp. The cells developed HLA-DR activation markers and infiltrated the red pulp where human B cells also appeared. Expression of activation and proliferation markers increased within 5 to 6 weeks. Many human CD3+ cells were also found in the portal triads of the liver and in the lung, pancreas, and kidney. The thymus also became heavily infiltrated. The intestines and skin of hu-PBL-SCID were less infiltrated by donor cells than in SCID with allogeneic GVHD. The tongue contained almost no human T cells. Our data show that a relatively low overall incidence of human xenogeneic GVHD, even when high numbers of human PBL are injected, is the consequence of a dichotomy between mice with no or transient T-cell chimerism and a minority of mice with high-blood T-lymphocyte chimerism and GVHD mortality.
AU  - Hoffmann-Fezer, G.
AU  - Gall, C.
AU  - Zengerle, U.
AU  - Kranz, B.R.
AU  - Thierfelder, S.S.
C1  - 40411
C2  - 37990
SP  - 3440-3448
TI  - Immunohistology and immunocytology of human T-cell chimerism and graft- versus-host disease in SCID mice.
JO  - Blood
VL  - 81
IS  - 12
PY  - 1993
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - The first two mutations causing hereditary glucose-6-phosphate isomerase (GPI) deficiency associated with chronic nonspherocytic hemolytic anemia in nonhuman mammals are described in the mouse. As in humans, the hemolytic syndrome, which is characterized by a diminished erythrocyte number, lower hematocrit, lower hemoglobin, higher number of reticulocytes and plasma bilirubin concentration, as well as increased liver- and spleen-somatic indices, was exclusively manifested in homozygous mutants. In comparison with wild type, heterozygous individuals exhibited neither hematologic differences nor alterations of other physiologic parameters, including plasma concentration of glucose, pyruvate and lactate, body weight, organo-somatic indices of liver, lung, kidney, spleen, and heart, as well as viability. Glycolytic intermediates, adenine nucleotides, and metabolic rate were not significantly altered in erythrocytes from heterozygotes. On the contrary, if allowance is made for the young erythrocyte population, homozygous mutant erythrocytes showed an increased concentration of glucose-6-phosphate and normal or decreased concentrations of glycolytic metabolites following the enzymatic block. The concentration of adenosine triphosphate and the glycolytic rate also appeared to be reduced. Homozygous anemic mice showed hepatosplenomegaly and typical adaptations to hypoxia, such as an elevated heart-somatic index and, for one mutant line, an enhanced lung-somatic index. Further, these animals were characterized by a marked reduction of body weight and an increase of lethality both correlated with the degree of enzyme deficiency in tissues. The latter findings were attributed to a reduced glycolytic capability of the whole organism caused by the enzyme defect in tissues, rather than representing secondary consequences of GPI deficiency in erythrocytes. The similarity in physicochemical and kinetic properties of the mutant murine proteins reported earlier with those of allozymes found in human GPI deficiency, as well as the comparable metabolic and physiologic consequences of this enzyme defect in mice and humans support that these murine mutants are excellent animal models for the human disease.
AU  - Merkle, S.
AU  - Pretsch, W.
C1  - 40307
C2  - 13044
SP  - 206-213
TI  - Glucose-6-phosphate isomerase deficiency associated with nonspherocytic hemolytic anemia in the mouse: An animal model for the human disease.
JO  - Blood
VL  - 81
IS  - 1
PY  - 1993
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - APO-1 is a cell surface molecule that induces apoptosis when ligated with the monoclonal antibody anti-APO-1. Expression of APO-1 and response to anti-APO-1 was investigated in a number of Epstein-Barr virus (EBV)-positive and -negative Burkitt lymphoma (BL) cell lines, in EBV-immortalized lymphoblastoid cell lines, and in cells from fresh BL biopsies. APO-1 was not expressed in EBV-negative cell lines and in EBV-positive BL cell lines with a phenotype corresponding to BL tumor biopsy cells (CD10+, CD21-, CD23-, CD30-, CD39-, CDw70-, CD77+). Accordingly, fresh BL cells obtained from three BL biopsies were APO-1 negative. EBV-positive BL cell lines that had acquired a lymphoblastoid phenotype (CD10-, CD21+, CD23+, CD30+, CD39+, CDw70+, CD77-) upon prolonged in vitro cultivation, as well as normal B-lymphoblastoid cell lines, expressed a high density of APO-1. APO-1 may, therefore, be regarded as a B-cell activation marker. APO-1 expression is not the only prerequisite for anti-APO-1-induced apoptosis because 6 of 7 APO-1-expressing EBV-positive BL cell lines were not sensitive to anti-APO-1, whereas all lymphoblastoid cell lines were killed by anti-APO-1. The sensitivity of lymphoblastoid cell lines to anti-APO-1-mediated apoptosis may open a new therapeutic approach for the treatment of EBV-induced lymphoproliferative lesions in immunocompromised individuals, because these are composed of cells with a lymphoblastoid phenotype.
AU  - Falk, M.H.
AU  - Trauth, B.C.
AU  - Debatin, K.-M.
AU  - Klas, C.
AU  - Gregory, C.D.
AU  - Rickinson, A.B.
AU  - Calender, A.
AU  - Lenoir, G.M.
AU  - Ellwart, J.W.
AU  - Krammer, P.H.
AU  - Bornkamm, G.W.
C1  - 19224
C2  - 12295
SP  - 3300-3306
TI  - Expression of the APO-1 antigen in Burkitt Lymphoma Cell Lines Correlates with a Shift towards a Lymphoblastoid Phenotype.
JO  - Blood
VL  - 79
IS  - 12
PY  - 1992
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - A hamster antimouse CD3 monoclonal antibody (MoAb) opened the way to experimental studies on the suppression of allograft rejection and cytokine- related morbidity after treatment with antibodies modulating the CD3/T-cell receptor complex (CD3/TCR). Because earlier attempts to suppress graft- versus-host disease (GVHD) in patients by in vitro treatment of donor marrow with anti-CD3 MoAb had remained inconclusive, we used a rat IgG2b antimouse CD3 MoAb (17A2) with fewer side effects to analyze suppression of GVHD in the mouse model. Detailed phenotyping of blood, spleen, and lymphnode T cells after the injection of 400 μg 17A2 in C57BL/6 mice showed 60% CD3 downmodulation and 50% T-cell depletion for spleen cells. Injection of these spleen cells, together with bone marrow cells, in fully mismatched preirradiated CBA mice delayed GVHD by only 6 days. Ex vivo treatment of donor cells with 17A2 was not effective. In contrast, conditioning of marrow recipients with a single injection of 17A2 delayed 50% GVHD mortality by 100 days and prevented GVHD altogether after prolonged treatment, with survivors showing complete chimerism and specific transplantation tolerance. This difference in antibody effect contrasts with earlier experiences with nonmodulating but more strongly T-cell-depleting MoAbs of the same isotype, which prevent GVHD no matter whether applied in vitro or injected into donor or recipient mice. Our data indicate that CD3/TCR reexpression in marrow recipients with no circulating 17A2 is the reason why ex vivo donor cell treatment with anti-CD3 MoAb is comparatively ineffective. Our data, which allow separate evaluation of cell-depleting and cell-modulating antibody activity, help to explain previous clinical failure to suppress GVHD and provide evidence in favor of conditioning the marrow recipient with anti-CD3 MoAb as a therapeutic alternative.
AU  - Mysliwietz, J.
AU  - Thierfelder, S.S.
C1  - 40622
C2  - 38745
SP  - 2661-2667
TI  - Antilymphocytic antibodies and marrow transplantation. XII. Suppression of graft-versus-host disease by T-cell-modulating and depleting antimouse CD3 antibody is most effective when preinjected in the marrow recipient.
JO  - Blood
VL  - 80
IS  - 10
PY  - 1992
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Cytokine expression was analyzed in CD14++ regular monocytes and in the novel subset of CD14+/CD16+ small monocytes. Biologic activity for tumor necrosis factor (TNF), interleukin-1 (IL-1), and IL-6 in the supernatant of elutriator-enriched, cell sorter-purified small monocytes was about 10-fold lower compared with regular monocytes when stimulated with lipopolysaccharide (LPS) for 12 hours. In CD14++ regular monocytes levels were 1,157 U x 10(-3)/mL, 158 U/mL, and 1,337 U/mL for TNF, IL-1, and IL-6, respectively. By contrast, CD14+/CD16+ small monocytes exhibited 137 U x 10(-3)/mL, 14 U/mL, and 60 U/mL for TNF, IL-1, and IL-6, respectively. Additional treatment with interferon-gamma enhanced production of TNF in both subsets, but CD14+/CD16+ small monocytes still exhibited lower levels. Stimulation of the monocyte subsets by platelet-activating factor gave the same pattern of results. Hybridization with 32P-labeled oligonucleotides specific for the respective cytokine messenger RNAs (mRNAs) showed a 10-fold lower prevalence of transcripts for TNF, IL-1, and IL-6, as well. By contrast, the constitutive expression of Glyceraldehyde-3-phosphate-dehydrogenase mRNA was 1.7-fold higher in the CD14+/CD16+ small monocytes. These data indicate that the novel subset of small monocytes is selectively suppressed in the expression of the cytokines TNF, IL-1, and IL-6, suggesting that these cells may comprise a deactivated type of cell. The expression of class II transcripts in the small monocytes is, however, similar to the regular monocytes, and the cell surface expression of class II protein about threefold increased. Thus, the novel subset of small monocytes appears to be a functionally distinct type of cell.
AU  - Ziegler-Heitbrock, L.
AU  - Ströbel, M.
AU  - Kieper, D.
AU  - Fingerle, G.
AU  - Schlunck, T.
AU  - Petersmann, I.
AU  - Ellwart, J.
AU  - Blumenstein, M.
AU  - Haas, J.G.
C1  - 19446
C2  - 12540
SP  - 503-511
TI  - Differential expression of cytokines in human blood monocyte subpopulations.
JO  - Blood
VL  - 79
IS  - 2
PY  - 1992
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Mergenthaler, H.-G.
AU  - Hültner, L.
AU  - Dörmer, P.
C1  - 18819
C2  - 11938
TI  - Generation of Interleukin 6 in Human Micro Longterm Bone Marrow Cultures.
JO  - Blood
VL  - Suppl. 1
PY  - 1991
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Remarkable differences in the suppression of graft-versus-host disease (GVHD) have been found for anti-Thy-1 antibodies to relate to (1) antigen density and antibody coating on the target cells, (2) antibody isotype, and (3) uptake of complement subcomponent C1q. Regarding (2) and (3) we now demonstrate that depletion of the third complement component C3 by cobra venom factor (CVF) differentiates two T-cell elimination pathways in mice: four rat IgG2c anti-Thy-1 monoclonal antibodies (MoAbs) with low uptake of mouse C1q lost most of their T-cell-depleting and consequently GVHD-preventing effect in C3-depleted H2 IA incompatible semiallogeneic (C57BL/6xCBA)F1 mice. In contrast, eight rat IgG2b, mouse IgG2a, and 2b anti-Thy-1 MoAbs with high affinity for C1q still remained strongly T-cell-depleting and prevented GVHD even in fully mismatched CBA mice depleted of C3. In conjunction with our observation that anti-Thy-1 MoAbs also suppress GVHD in C5-deficient AKR mice, we conclude that complete complement activation until T-cell lysis is not required for our antibodies to be effective in vivo. Activation, but only until deposition of C3b on target-cells for opsonisation via C3b receptors, is necessary with the less immunosuppressive anti-Thy-1 IgG2c isotype with low affinity for C1q. Mouse C1q uptake and C3/C4 deposition on target cells were measured with labeled antibodies and localized in T-cell areas. Interestingly, not even activation until C3b is necessary with the most immunosuppressive C1q-high-affine isotypes. As far as the latter is concerned, we discuss whether elimination of antibody-coated cells via Fc receptors is enhanced by binding to C1q-receptors and/or by intercalating C1q expressed on macrophages.
AU  - Thierfelder, S.
AU  - Mysliwietz, J.
AU  - Hoffmann-Fezer, G.
AU  - Kummer, U.
C1  - 19021
C2  - 12062
SP  - 2285-2291
TI  - Antilymphocytic Antibodies and Marrow Transplantation. XIV. Antibody-Induced Suppression of Graft-Versus-Host Disease in C3-Decomplemented Mice Differentiates Two T-Cell-Depletion Pathways.
JO  - Blood
VL  - 77
IS  - 10
PY  - 1991
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Acute graft-versus-host disease, interstitial pneumonitis, endothelial leakage syndrome, and veno-occlusive disease are major complications of bone marrow transplantation. Though several new regimens for prophylaxis and treatment of these syndromes have been introduced, the overall incidence has been only slightly reduced over the last few years. We retrospectively analyzed tumor necrosis factor alpha (TNF alpha) serum levels between day -8 and day 100 after bone marrow transplantation in 56 patients transplanted in our unit for a variety of hematological diseases. In 34 patients with uneventful courses, mean TNF alpha levels rose to a maximum of 76 +/- 29 pg/mL. In contrast, 22 patients with major transplant related complications showed mean increases of TNF alpha of 492 +/- 235 pg/mL (P less than .0001). Increases of TNF alpha occurred before interstitial pneumonitis and severe acute graft-versus-host disease with a latency of 25 to 54 days. Early complications such as endothelial leakage syndrome and veno-occlusive disease were closely associated with increases of TNF alpha serum levels. Our study suggests two pathways of TNF alpha release: activation of host macrophages and stimulation of donor cells in the course of acute graft-versus-host disease. Cytokine monitoring should be helpful for prediction and earlier treatment of major transplant related complications.
AU  - Holler, E.
AU  - Kolb, H.-J.
AU  - Möller, A.
AU  - Kempeni, J.
AU  - Liesenfeld, S.
AU  - Pechumer, H.
AU  - Lehmacher, W.
AU  - Ruckdeschel, G.
AU  - Gleixner, B.
AU  - Riedner, C.
AU  - Ledderose, G.
AU  - Brehm, G.
AU  - Mittermüller, J.
AU  - Wilmanns, W.
C1  - 19729
C2  - 12858
SP  - 1011-1016
TI  - Increased serum levels of tumor necrosis factor alpha precede major complications of bone marrow transplantation.
JO  - Blood
VL  - 75
IS  - 4
PY  - 1990
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Severe microangiopathy has been reported as a rare complication of cyclosporine A (CsA) prophylaxis in allogeneic bone marrow transplantation (BMT). We found morphological and biochemical changes indicative of generalized endothelial damage in 49 of 66 allogeneic marrow graft recipients receiving cyclosporine, but none in 11 patients treated with methotrexate for prophylaxis of graft-v-host disease (GVHD). Changes occurred after engraftment of bone marrow and consisted of intravascular hemolysis with red cell fragmentation and de novo thrombocytopenia. They were preceded by a decrease in activated partial thromboplastin time and fibrinogen indicating activation of coagulation. Endothelial damage as the central lesion of microangiopathy was confirmed by a simultaneous increase of factor VIII related antigen. Severe microangiopathy was observed in ten patients and was fatal in seven. Risk factor analysis revealed a highly significant association of microangiopathy with severity of acute GVHD (aGVHD) (P less than .001) and use of CsA prophylaxis (P less than .001). Our data suggest endothelial damage as a result of cellular activation and subsequent release of cytokines in the course of a aGVHD, which is not inhibited by CsA prophylaxis.
AU  - Holler, E.
AU  - Kolb, H.-J.
AU  - Hiller, E.
AU  - Mraz, W.
AU  - Lehmacher, W.
AU  - Gleixner, B.
AU  - Seeber, Ch.
AU  - Jehn, U.
AU  - Gerhartz, H.H.
AU  - Brehm, G.
AU  - Wilmanns, W.
C1  - 17589
C2  - 10801
SP  - 2018-2024
TI  - Microangiopathy in Patients on Cyclosporine Prophylaxis Who Developed Acute Graft-Versus- Host Disease After HLA-Identical Bone Marrow Transplantation.
JO  - Blood
VL  - 73
IS  - 7
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Use of immunocytology for accurate identification of malignant cells in cerebrospinal fluid (CSF) has so far been hampered by high cell requirements of the immunologic methods hitherto used. In an attempt to minimize cell loss in cytopreparation, electrostatic binding of cells to poly-L-lysine (PLL)-coated multispot slides, followed by immunocytochemistry, was investigated. Using optimized conditions of cell attachment and fixation and performing all washing procedures on the slide made multimarker analysis possible even in paucicellular specimens, while preserving excellent cell morphology and yielding high sensitivity in the detection of antigens. In a study of 26 CSF specimens with inconclusive cytomorphology, comprising 335 single marker determinations, we were able to discriminate reliably between resting or activated benign cells and a wide range of types of malignant lymphoid cell. A definitive diagnosis was reached in all cases by one tap only. Malignant meningitis was ruled out in ten specimens and proved in 16, including five in which the type of malignancy could only be determined by immunophenotyping. We conclude that immunocytochemistry on PLL-coated slides constitutes the method of choice for immunologic cell differentiation in CSF, which allows equivocal morphologic findings to be clarified.
AU  - Kranz, B.R.
AU  - Thiel, E.
AU  - Thierfelder, S.
C1  - 17812
C2  - 10726
SP  - 1942-1950
TI  - Immunocytochemical Identification of Meningeal Leukemia and Lymphoma: Poly-L-Lysine- Coated Slides Permit Multimarker Analysis even with Minute Cerebrospinal Fluid Cell Specimens.
JO  - Blood
VL  - 73
IS  - 7
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Use of immunocytology for accurate identification of malignant cells in cerebrospinal fluid (CSF) has so far been hampered by high cell requirements of the immunologic methods hitherto used. In an attempt to minimize cell loss in cytopreparation, electrostatic binding of cells to poly-L-lysine (PLL)-coated multispot slides, followed by immunocytochemistry, was investigated. Using optimized conditions of cell attachment and fixation and performing all washing procedures on the slide made multimarker analysis possible even in paucicellular specimens, while preserving excellent cell morphology and yielding high sensitivity in the detection of antigens. In a study of 26 CSF specimens with inconclusive cytomorphology, comprising 335 single marker determinations, we were able to discriminate reliably between resting or activated benign cells and a wide range of types of malignant lymphoid cell. A definitive diagnosis was reached in all cases by one tap only. Malignant meningitis was ruled out in ten specimens and proved in 16, including five in which the type of malignancy could only be determined by immunophenotyping. We conclude that immunocytochemistry on PLL-coated slides constitutes the method of choice for immunologic cell differentiation in CSF, which allows equivocal morphologic findings to be clarified.
AU  - Kranz, B.R.
AU  - Thiel, E.V.
AU  - Thierfelder, S.S.
C1  - 34006
C2  - 36515
SP  - 1942-1950
TI  - Immunocytochemical identification of meningeal leukemia and lymphoma: Poly-L-lysine-coated slides permit multimarker analylsis even with minute cerebrospinal fluid cell specimens.
JO  - Blood
VL  - 73
IS  - 7
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Acute and chronic lymphatic leukemias were investigated on the single- cell level for the activity of genes coding for the IgM heavy chain and the alpha and beta chains of the T-cell antigen receptor (TCR). We used a new method for preparing highly fluorochrome-labeled gene probes for in situ hybridization, which allowed rapid and quantitative detection of mRNA at the individual cell level. Leukemic cell populations classified as belonging to the B lineage according to their surface antigenic patterns revealed increasing expression of mRNA for the IgM heavy chain (mu mRNA) in a maturation-dependent fashion, which was not correlated to rearrangement of the immunoglobulin mu chain gene--only 66% of the leukemias with rearranged mu gene also transcribed it. TCR mRNA was detected in B-antigen positive leukemic cells. High levels of both TCR and mu mRNA expression in all cells of some of these leukemias allowed the conclusion that these cells simultaneously transcribed the genes for T and B cell antigen receptors. TCR mRNA was also found in what are considered relatively mature B leukemias, lineage cross-over on the mRNA level being observed at a frequency of 23% (five of 22 cases), comparable with that of “inappropriate” receptor gene rearrangement. The quantitation of mRNA with fluorochrome-labeled gene probes in situ may allow determining the degree of gene activation in individual antigenically defined cells and may thus contribute a new tool for characterization of normal and malignant cells.
AU  - Mar, P.
AU  - Pachmann, K.
AU  - Reinecke, K.
AU  - Emmerich, B.
AU  - Thiel, E.
C1  - 17824
C2  - 10792
SP  - 638-644
TI  - Quantitative Tracing of mRNAs for T- and B-Lymphocyte Receptor Genes in Individual Cells by In Situ Hybridization With Fluorochrome-Labeled Gene Probes. I. Expression in Malignancies Carrying B-Lineage Associated Antigens.
JO  - Blood
VL  - 74
IS  - 2
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Reisbach, G.
AU  - Sindermann, J.
AU  - Kremer, J.P.
AU  - Hültner, L.
AU  - Wolf, H.
AU  - Dörmer, P.
C1  - 17826
C2  - 10790
SP  - 959-964
TI  - Macrophage Colony-Stimulating Factor (CSF-1) Is Expressed by Spontaneously Outgrown EBV- B Cell Lines and Activated Normal B Lymphocytes.
JO  - Blood
VL  - 74
IS  - 3
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Human B lymphocytes activated by mitogens or infected by Epstein Barr virus (EBV) have previously been shown to release colony-stimulating activity (CSA) supporting the growth of normal human bone marrow progenitors. We established five different human EBV-B cell lines spontaneously outgrown from nonmalignant peripheral blood cells and long-term bone marrow cultures. CSA derived from all of these lines induces the growth of murine macrophage colonies, whereas virtually no human bone marrow cell progenitors were stimulated. As observed in the tumor cell line MIA PaCa-2, a 4.3 kilobase (kb) transcript was detected in all cases using a human colony-stimulating factor (CSF)-1 probe. Expression of this transcript can be further stimulated within three hours upon addition of phorbol myristate acetate (PMA). The highly purified native protein exerting macrophage colony-stimulating activity (M-CSA) exhibits a molecular size of approximately 75 to 97 Kd in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The identity of EBV-B cell derived M-CSA with human urinary CSF-1 was confirmed by a complete neutralization of macrophage CSA by an antihuman urinary CSF-1 antiserum. Normal human B lymphocytes purified from tonsils or from mononuclear blood cells also express CSF-1 upon stimulation with Staphylococcus aureus Cowan I. No CSF-1 expression, however, could be detected in normal resting B lymphocytes or in the Burkitt lymphoma cell line RAJI.
AU  - Reisbach, G.
AU  - Sindermann, J.R.
AU  - Kremer, J.P.
AU  - Hültner, L.
AU  - Wolf, H.J.
AU  - Dörmér, P.G.
C1  - 41925
C2  - 36494
SP  - 959-964
TI  - Macrophage colony-stimulating factor (CSF-1) is expressed by spontaneously outgrown EBV-B cell lines and activated normal B lymphocytes.
JO  - Blood
VL  - 74
IS  - 3
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Pretreatment blast cells from 739 adults with acute lymphoblastic leukemia (ALL) were immunophenotyped as part of a prospective treatment protocol study. Among 192 patients (26%) with T lineage ALL, 47 (6%; 24% of T lineage ALL) had lymphoblasts without sheep erythrocyte rosette formation, but with pan-T antigen CD7 on the membrane and intracellular CD3 proteins mostly in perinuclear accumulation. The T-cell surface antigens CD5 and/or CD2 and focal acid phosphatase were additional markers of this subgroup traditionally called pre-T ALL, whereas thymocyte antigen CD1 as well as CD4 and CD8 antigens were not expressed. Hematopoietic progenitor cell markers, namely terminal deoxynucleotidyl transferase (TdT), and in part common ALL antigen (CD10), HLA-DR antigens, and/or My-10 (CD34), a unique antigen of marrow cells absent in thymus cells, further characterized this immature T-ALL form of putative prothymocytic phenotype (CD7+/intracellular CD3+/TdT+/My-10+/-/HLA-DR+/-/CD10+/-). The prethymic T cell character was supported by germ-line T-cell receptor beta genes found in 21 of 36 patients analyzed. In five cases only T gamma-chain genes were rearranged. Fifteen patients, however, had rearrangements of both T beta and T gamma genes. Immunoglobulin heavy chain genes were rearranged only in two cases. Pre-T ALL differed significantly from E-rosette+ T-ALL in some presenting clinical features, namely mediastinal mass, lymphoadenopathy, and platelet count, and independently of clinical factors in prognosis (P = .02, median remission duration: 15.7 v 33.5 months, and P = .02, median survival time: 24.6 v 50.7 months). We conclude that ALL classification based solely on T- or B-cell lineage affiliation is not sufficient but needs further subdivision according to relevant maturation stages as exemplified here within the T-cell axis. The putative prethymic T cell progenitor phenotype described might help elucidate the sequence of genetic events that commit normal hematopoietic cells to the T-cell lineage.
AU  - Thiel, E.
AU  - Kranz, B.R.
AU  - Raghavachar, A.
AU  - Bartram, C.R.
AU  - Löffler, H.
AU  - Messerer, D.
AU  - Ganser, A.
AU  - Ludwig, W.-D.
AU  - Büchner, T.
AU  - Hoelzer, D.
C1  - 18063
C2  - 10904
SP  - 1247-1258
TI  - Prethymic Phenotype and Genotype of Pre-T (CD7+/ER-)-Cell Leukemia and its Clinical Significance Within Adult Acute Lymphoblastic Leukemia.
JO  - Blood
VL  - 73
IS  - 5
PY  - 1989
SN  - 0006-4971
ER  - 

TY  - JOUR
AU  - Riccardi, A.
AU  - Danova, M.
AU  - Wilson, G.
AU  - Ucci, G.
AU  - Dörmer, P.
C1  - 17327
C2  - 10138
SP  - 267a 
TI  - Cell Proliferation in Hematological and Solid Tumors: in Vivoistudy with Bromodeoxyuridine and Flow Cytometry.
JO  - Blood
VL  - 5
PY  - 1987
SN  - 0006-4971
ER  - 

TY  - JOUR
AB  - Peripheral blood leukemia cells from patients with acute monoblastic leukemia (AMoL) were tested for killer cell activity against target cells that detected natural killer cell-mediated or monocyte-mediated spontaneous cytotoxicity. The fibrosarcoma cell line Wehi 164, pretreated with actinomycin D to induce susceptibility to lysis, specifically detects the activity of unstimulated human monocytes. In four of six cases of AMoL, high killer cell activity could be measured against this target. In three of these four cases, the killer cell activity could be assigned exclusively to the leukemic clone, based on the high leukocyte counts and the resultant dilution of normal cells, as evidenced by marker and by functional analysis. While leukemic cells with killer cell activity against Wehi 164 contained 34% to 45% cells that were positive for binding of the 63D3 monoclonal antibody, the two leukemic samples without killer cell activity contained only 1% and 12% 63D3-positive cells. Cell sorting of 63D3-positive and -negative cells from two leukemias with killer cell activity demonstrated that the killer cell activity was restricted to the 63D3-positive fraction of AMoL cells. These data demonstrate that monoblastic leukemia cells can be potent killer cells and that killing activity is linked to the 63D3-defined cell surface molecule.
AU  - Ziegler-Heitbrock, L.
AU  - Munker, R.
AU  - Thiel, E.
AU  - Krebs, I.
AU  - Riethmüller, G.
C1  - 30787
C2  - 33867
SP  - 8-14
TI  - Killer cell activity of human monoblastic leukemia cells as detected with a monocyte-specific target cell.
JO  - Blood
VL  - 65
IS  - 1
PY  - 1985
SN  - 0006-4971
ER  -