TY - JOUR AB - The pro-survival MCL1 protein is overexpressed in many cancers, including B-cell non-Hodgkin lymphomas (B-NHL). S63845 is a highly specific inhibitor of MCL1. We analyzed mechanisms of sensitivity/resistance to S63845 in preclinical models of diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma (BL). Annexin V-based cytotoxic assays, western blot, protein co-immunoprecipitation, and cell clones with manipulated expression of BCL2 family proteins were used to analyze mechanisms of sensitivity to S63845. Experimental in vivo therapy with S63845 and/or venetoclax was performed using patient-derived xenografts (PDX) of treatment-refractory B-NHL. A subset of DLBCL and majority of BL cell lines were sensitive to S63845. The level of BCL2 protein expression was the major determinant of resistance to S63845: BCL2 serves as a buffer for pro-apoptotic proteins released from MCL1 upon exposure to S63845. While BCL2-negative lymphomas were effectively eliminated by single-agent S63845, its combination with venetoclax was synthetically lethal in BCL2-positive PDX models. Concerning MCL1, both, the level of MCL1 protein expression, and its occupational status represent key factors mediating sensitivity to S63845. In contrast to MCL1-BIM / BAK1 complexes that prime lymphoma cells for S63845-mediated apoptosis, MCL1-NOXA complexes are associated with S63845 resistance. In conclusion, MCL1 represents a critical survival molecule for most BLs and a subset of BCL2-negative DLBCLs. The level of BCL2 and MCL1 expression and occupational status of MCL1 belong to the key modulators of sensitivity/resistance to S63845. Co-treatment with venetoclax can overcome BCL2-mediated resistance to S63845, and enhance efficacy of MCL1 inhibitors in BCL2-positive aggressive B-NHL. AU - Klanova, M.* AU - Kazantsev, D.* AU - Pokorna, E.* AU - Zikmund, T. AU - Karolova, J.* AU - Behounek, M.* AU - Renesova, N.* AU - Sovilj, D.* AU - Kelemen, C.D.* AU - Helman, K.* AU - Jaksa, R.* AU - Havranek, O.* AU - Andera, L.* AU - Trneny, M.* AU - Klener, P.* C1 - 63422 C2 - 51528 CY - 615 Chestnut St, 17th Floor, Philadelphia, Pa 19106-4404 Usa SP - 89-99 TI - Anti-apoptotic MCL1 protein represents critical survival molecule for most Burkitt lymphomas and BCL2-negative diffuse large B-cell lymphomas. JO - Mol. Cancer Ther. VL - 21 IS - 1 PB - Amer Assoc Cancer Research PY - 2021 SN - 1535-7163 ER - TY - JOUR AB - The pattern recognition receptor RIG-I plays an important role in the recognition of nonself RNA and antiviral immunity. RIG-I's natural ligand, triphosphate RNA (ppp-RNA), is proposed to be a valuable addition to the growing arsenal of cancer immunotherapy treatment options. In this study, we present comprehensive data validating the concept and utility of treatment with synthetic RIG-I agonist ppp-RNA for the therapy of human cancer, with melanoma as potential entry indication amenable to intratumoral treatment. Using mRNA expression data of human tumors, we demonstrate that RIG-I expression is closely correlated to cellular and cytokine immune activation in a wide variety of tumor types. Furthermore, we confirm susceptibility of cancer cells to ppp-RNA treatment in different cellular models of human melanoma, revealing unexpected heterogeneity between cell lines in their susceptibility to RNA agonist features, including sequence, secondary structures, and presence of triphosphate. Cellular responses to RNA treatment (induction of type I IFN, FasK, MHC-I, and cytotoxicity) were demonstrated to be RIG-I dependent using KO cells. Following ppp-RNA treatment of a mouse melanoma model, we observed significant local and systemic antitumor effects and survival benefits. These were associated with type I IFN response, tumor cell apoptosis, and innate and adaptive immune cell activation. For the first time, we demonstrate systemic presence of tumor antigen-specific Cris following treatment with RIG-I agonists. Despite potential challenges in the generation and formulation of potent RIG-I agonists, ppp-RNA or analogues thereof have the potential to play an important role for cancer treatment in the next wave of immunotherapy. AU - Helms, M.W.* AU - Jahn-Hofmann, K.* AU - Gnerlich, F.* AU - Metz-Weidmann, C.* AU - Braun, M.* AU - Dietert, G.* AU - Scherer, P.* AU - Grandien, K.* AU - Theilhaber, J.* AU - Cao, H.* AU - Wagenaar, T.R.* AU - Schnurr, M.M.* AU - Endres, S. AU - Wiederschain, D.* AU - Scheidler, S.* AU - Rothenfußer, S. AU - Brunner, B.* AU - König, L.M.* C1 - 57603 C2 - 47860 CY - 615 Chestnut St, 17th Floor, Philadelphia, Pa 19106-4404 Usa SP - 2343-2356 TI - Utility of the RIG-I agonist triphosphate RNA for melanoma therapy. JO - Mol. Cancer Ther. VL - 18 IS - 12 PB - Amer Assoc Cancer Research PY - 2019 SN - 1535-7163 ER - TY - JOUR AB - Pancreatic ductal adenocarcinoma (PDAC) is likely the most aggressive and therapy-resistant of all cancers. Aim of this study was to investigate the emerging technology of matrix assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) as a powerful tool to study drug delivery and spatial tissue distribution in PDAC. We utilized an established genetically engineered mouse model of spontaneous PDAC to examine the distribution of the small molecule inhibitor erlotinib in healthy pancreas and PDAC. MALDI IMS was utilized on sections of single-dose or long-term-treated mice to measure drug tissue distribution. Histological and statistical analyses were performed to correlate morphology, drug distribution and survival. We found that erlotinib levels were significantly lower in PDAC compared to healthy tissue (p = 0.0078). Survival of long-term-treated mice did not correlate with overall levels of erlotinib or with overall histological tumor grade but both with the percentage of atypical glands in the cancer (p = 0.021, rs = 0.59) and the level of erlotinib in those atypical glands (p = 0.019, rs = 0.60). The results of this pilot study present MALDI IMS as a reliable technology to study drug delivery and spatial distribution compounds in a preclinical setting and supports drug imaging-based translational approaches. AU - Grüner, B.M.* AU - Winkelmann, I. AU - Feuchtinger, A. AU - Sun, N. AU - Balluff, B.* AU - Teichmann, N.* AU - Herner, A.* AU - Kalideris, E.* AU - Steiger, K.* AU - Braren, R.* AU - Aichler, M. AU - Esposito, I.* AU - Schmid, R.M.* AU - Walch, A.K. AU - Siveke, J.T.* C1 - 47787 C2 - 39477 CY - Philadelphia SP - 1145-1152 TI - Modeling therapy response and spatial tissue distribution of erlotinib in pancreatic cancer. JO - Mol. Cancer Ther. VL - 15 IS - 5 PB - Amer Assoc Cancer Research PY - 2016 SN - 1535-7163 ER - TY - JOUR AB - Monoclonal anti-HER2 antibody trastuzumab has significantly improved the survival of patients with HER2-overexpressing tumors. Nevertheless, systemic antibody therapy is expensive, limited in efficacy due to physical tumor barriers, and carries the risk of severe side effects such as cardiomyopathy. Oncolytic viruses mediate cancer-selective transgene expression, kill infected cancer cells while mounting antitumor immune responses, and have recently demonstrated promising efficacy in combination treatments. Here, we armed an oncolytic adenovirus with full-length trastuzumab to achieve effective in situ antibody production coupled with progressive oncolytic cancer cell killing. We constructed an infectivity-enhanced serotype 5 oncolytic adenovirus, Ad5/3-D24-tras, coding for human trastuzumab antibody heavy-and light-chain genes, connected by an internal ribosome entry site. Infected cancer cells were able to assemble full-length functional antibody, as confirmed by Western blot, ELISA, and antibody-dependent cell-mediated cytotoxicity assay. Importantly, oncolysis was required for release of the antibody into tumors, providing additional spatial selectivity. Ad5/3-D24-tras showed potent in vitro cytotoxicity and enhanced antitumor efficacy over oncolytic control virus Ad5/3-D24 or commercial trastuzumab in HER2-positive cancer models in vivo (both P < 0.05). Furthermore, Ad5/3-D24-tras resulted in significantly higher tumor-to-systemic antibody concentrations (P < 0.001) over conventional delivery. Immunological analyses revealed dendritic cell activation and natural killer cell accumulation in tumor-draining lymph nodes. Thus, Ad5/ 3-D24-tras is an attractive anticancer approach combining oncolytic immunotherapy with local trastuzumab production, resulting in improved in vivo efficacy and immune cell activation in HER2-positive cancer. Moreover, the finding that tumor cells can produce functional antibody as directed by oncolytic virus could lead to many valuable antitumor approaches. AU - Liikanen, I.* AU - Tahtinen, S.* AU - Guse, K.* AU - Gutmann, T. AU - Savola, P.* AU - Oksanen, M.* AU - Kanerva, A.* AU - Hemminki, A.* C1 - 49650 C2 - 40870 CY - Philadelphia SP - 2259-2269 TI - Oncolytic adenovirus expressing monoclonal antibody trastuzumab for treatment of HER2-positive cancer. JO - Mol. Cancer Ther. VL - 15 IS - 9 PB - Amer Assoc Cancer Research PY - 2016 SN - 1535-7163 ER - TY - JOUR AB - Trifunctional bispecific antibodies (trAb) are novel anticancer drugs that recruit and activate different types of immune effector cells at the targeted tumor. Thus, tumor cells are effectively eliminated and a long-lasting tumor-specific T-cell memory is induced. The trAb Ektomab is directed against human CD3 on T cells and the tumor-associated ganglioside GD2, which is an attractive target for immunotherapy of melanoma in humans. To optimize clinical applicability, we studied different application routes with respect to therapeutic efficacy and tolerability by using the surrogate trAb Surek (anti-GD2 x anti-murine CD3) and a murine melanoma engineered to express GD2. We show that subcutaneous injection of the trAb is superior to the intravenous delivery pathway, which is the standard application route for therapeutic antibodies. Despite lower plasma levels after subcutaneous administration, the same tumor-protective potential was observed in vivo compared to intravenous administration of Surek. However, subcutaneously delivered Surek showed better tolerability. This could be explained by a continuous release of the antibody leading to constant plasma levels and a delayed induction of proinflammatory cytokines. Importantly, the induction of counter-regulatory mechanisms was reduced after subcutaneous application. These findings are relevant for the clinical application of trifunctional bispecific antibodies and possibly also other immunoglobulin constructs. AU - Deppisch, N. AU - Ruf, P.* AU - Eissler, N. AU - Neff, F. AU - Buhmann, R.* AU - Lindhofer, H.* AU - Mocikat, R. C1 - 45187 C2 - 37255 CY - Philadelphia SP - 1877-1883 TI - Efficacy and tolerability of a GD2-directed trifunctional bispecific antibody in a preclinical model: Subcutaneous administration is superior to intravenous delivery. JO - Mol. Cancer Ther. VL - 14 IS - 8 PB - Amer Assoc Cancer Research PY - 2015 SN - 1535-7163 ER - TY - JOUR AB - In concurrent chemoradiotherapy, drugs are used to sensitize tumors to ionizing radiation. Although a spectrum of indications for simultaneous treatment with drugs and radiation has been defined, the molecular mechanisms underpinning tumor radiosensitization remain incompletely characterized for several such combinations. Here, we investigate the mechanisms of radiosensitization by the arabinoside nucleoside analogue 9-b-D-arabinofuranosyladenine (araA) placing particular emphasis on the repair of DNA double-strand breaks (DSB), and compare the results to those obtained with fludarabine (F-araA) and cytarabine (araC). Postirradiation treatment with araA strongly sensitizes cells to ionizing radiation, but leaves unchanged DSB repair by NHEJ in logarithmically growing cells, in sorted G1 or G2 phase populations, as well as in cells in the plateau phase of growth. Notably, araA strongly inhibits DSB repair by homologous recombination (HRR), as assessed by scoring ionizing radiation-induced RAD51 foci, and in functional assays using integrated reporter constructs. Cells compromised in HRR by RNAi-mediated transient knockdown of RAD51 show markedly reduced radiosensitization after treatment with araA. Remarkably, mutagenic DSB repair compensates for HRR inhibition in araA-treated cells. Compared with araA, F-araA and araC are only modestly radiosensitizing under the conditions examined. We propose that the radiosensitizing potential of nucleoside analogues is linked to their ability to inhibit HRR and concomitantly promote the errorprone processing of DSBs. Our observations pave the way to treatment strategies harnessing the selective inhibitory potential of nucleoside analogues and the development of novel compounds specifically utilizing HRR inhibition as a means of tumor cell radiosensitization. AU - Magin, S.* AU - Papaioannou, M.* AU - Saha, J.* AU - Staudt, C. AU - Iliakis, G.* C1 - 46891 C2 - 39011 SP - 1424-1433 TI - Inhibition of homologous recombination and promotion of mutagenic repair of DNA double-strand breaks underpins arabinoside-nucleoside analogue radiosensitization. JO - Mol. Cancer Ther. VL - 14 IS - 6 PY - 2015 SN - 1535-7163 ER - TY - JOUR AB - Activating mutations of the class III receptor tyrosine kinase FLT3 are the most frequent molecular aberration in acute myeloid leukemia (AML). Mutant FLT3 accelerates proliferation, suppresses apoptosis, and correlates with poor prognosis. Therefore, it is a promising therapeutic target. Here, we show that RNA interference against FLT3 with an internal tandem duplication (FLT3-ITD) potentiates the efficacy of the histone deacetylase inhibitor (HDACi) panobinostat (LBH589) against AML cells expressing FLT3-ITD. Similar to RNA interference, tyrosine kinase inhibitors (TKI; AC220/cpd.102/PKC412) in combination with LBH589 exhibit superior activity against AML cells. Median dose-effect analyses of drug-induced apoptosis rates of AML cells (MV4-11 and MOLM-13) revealed combination index (CI) values indicating strong synergism. AC220, the most potent and FLT3-specific TKI, shows highest synergism with LBH589 in the low nanomolar range. A 4-hour exposure to LBH589 + AC220 already generates more than 50% apoptosis after 24 hours. Different cell lines lacking FLT3-ITD as well as normal peripheral blood mononuclear cells are not significantly affected by LBH589 + TKI, showing the specificity of this treatment regimen. Immunoblot analyses show that LBH589 + TKI induce apoptosis via degradation of FLT3-ITD and its prosurvival target STAT5. Previously, we showed the LBH589-induced proteasomal degradation of FLT3-ITD. Here, we show that activated caspase-3 also contributes to the degradation of FLT3-ITD and that STAT5 is a direct target of this protease. Our data strongly emphasize HDACi/TKI drug combinations as promising modality for the treatment of FLT3-ITD-positive AMLs. AU - Pietschmann, K.* AU - Bolck, H.A.* AU - Buchwald, M.* AU - Spielberg, S.* AU - Polzer, H. AU - Spiekermann, K. AU - Bug, G.* AU - Heinzel, T.* AU - Böhmer, F.D.* AU - Krämer, O.H.* C1 - 10994 C2 - 30459 SP - 2373-2383 TI - Breakdown of the FLT3-ITD/STAT5 axis and synergistic apoptosis induction by the histone deacetylase inhibitor panobinostat and FLT3-specific inhibitors. JO - Mol. Cancer Ther. VL - 11 IS - 11 PB - Amer. Assoc. Cancer Research PY - 2012 SN - 1535-7163 ER - TY - JOUR AB - Constitutive activation of the phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR signaling cascade occurs in a variety of human malignancies, where it sustains tumor cell proliferation and survival. Pharmacologic blockade of this pathway exerts antineoplastic activity by triggering apoptosis and/or cell-cycle arrest. Pituitary adenomas show activation of the PI3K/AKT/mTOR pathway, but only a fraction of them respond in vitro to the antiproliferative action of rapamycin and RAD001 (mTOR inhibitors), possibly because of the described negative feedback loop on AKT which reactivates the signaling cascade. Rats affected by the multiple endocrine neoplasia-like syndrome (MENX) develop pituitary adenomas showing increased activated AKT. In this study, we comparatively investigated the antitumor potential of the novel dual PI3K/mTOR inhibitor NVP-BEZ235 and the single mTOR inhibitor RAD001 on rat pituitary adenoma cells in primary culture. NVP-BEZ235 inhibits the PI3K pathway both upstream and downstream of AKT, thereby preventing the negative feedback loop. NVP-BEZ235 was more effective than RAD001 in reducing cell viability of pituitary adenomas. Consistently, NVP-BEZ235 treatment decreased Akt and S6 phosphorylation and triggered apoptosis. Because MENX is caused by a germline loss-of-function mutation in the cell-cycle inhibitor p27Kip1, we investigated the relationship between this defect and response to NVP-BEZ235 treatment. The levels of p27Kip1 positively correlate with the response to NVP-BEZ235 treatment. Combined treatment with NVP-BEZ235 and the proteasome inhibitor bortezomib, which increases p27Kip1 amount, shows synergistic antiproliferative effects on pituitary adenoma cells. Our data suggest that NVP-BEZ235 may represent an effective therapeutic modality for pituitary adenomas and that p27Kip1 levels represent a potential predictor of response to dual PI3K/mTOR inhibition. AU - Lee, M.S. AU - Theodoropoulou, M.* AU - Graw, J. AU - Roncaroli, F.* AU - Zatelli, M.C.* AU - Pellegata, N.S. C1 - 6653 C2 - 29043 CY - Philadelphia, PA , USA SP - 1450-1459 TI - Levels of p27 sensitize to dual PI3K/mTOR inhibition. JO - Mol. Cancer Ther. VL - 10 IS - 8 PB - American Assoc. for Cancer Research PY - 2011 SN - 1535-7163 ER - TY - JOUR AB - Perineural invasion, the growth of tumor cells along nerves, is a key feature of pancreatic cancer. The cardinal symptom of pancreatic cancer, abdominal pain often radiating to the back, as well as the high frequency of local tumor recurrence following resection are both attributed to the unique ability of pancreatic tumor cells to invade the neuronal system. The molecular mechanisms underlying the neuroaffinity of pancreatic tumors are not completely understood. In this study, we developed a novel method to monitor ex vivo perineural invasion into surgically resected rat vagal nerves by different human pancreatic tumor cell lines. Genome-wide transcriptional analyses were employed to identify the consensus set of genes differentially regulated in all highly nerve-invasive (nerve invasion passage 3) versus less invasive (nerve invasion passage 0) pancreatic tumor cells. The critical involvement of kinesin family member 14 (KIF14) and Rho-GDP dissociation inhibitor beta (ARHGDI beta) in perineural invasion was confirmed on RNA and protein levels in human pancreatic tumor specimens. We found significant up-regulation of KIF14 and ARHGDI beta mRNA levels in patients with pancreatic cancer, and both proteins were differentially expressed in tumor cells invading the perineural niche of pancreatic cancer patients as detected by immunohistochemistry. Moreover, functional knockdown of KIF14 and ARHGDI beta using small interfering RNA resulted in altered basal and/or perineural invasion of pancreatic tumor cells. Our work provides novel insights into the molecular determinants of perineural invasion in pancreatic cancer. The established nerve invasion model and the consensus signature of perineural invasion could be instrumental in the identification of novel therapeutic targets of pancreatic cancer as exemplified by KIF14 and ARHGDI beta. AU - Abiatari, I.* AU - DeOliveira, T.* AU - Kerkadze, V.* AU - Schwager, C.* AU - Esposito, I. AU - Giese, N.A.* AU - Huber, P.* AU - Bergman, F.* AU - Abdollahi, A.* AU - Friess, H.* AU - Kleeff, J.* C1 - 517 C2 - 26765 SP - 1494-1504 TI - Consensus transcriptome signature of perineural invasion in pancreatic carcinoma. JO - Mol. Cancer Ther. VL - 8 IS - 6 PB - Amer Assoc Cancer Research PY - 2009 SN - 1535-7163 ER -