Fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) is one of the most common genetic lesions in acute myeloid leukemia patients (AML). Although FLT3 tyrosine kinase inhibitors initially exhibit clinical activity, resistance to treatment inevitably occurs within months. PIM kinases are thought to be major drivers of the resistance phenotype and their inhibition in relapsed samples restores cell sensitivity to FLT3 inhibitors. Thus, simultaneous PIM and FLT3 inhibition represents a promising strategy in AML therapy. For such reasons, we have developed SEL24-B489 - a potent, dual PIM and FLT3-ITD inhibitor. SEL24-B489 exhibited significantly broader on-target activity in AML cell lines and primary AML blasts than selective FLT3-ITD or PIM inhibitors. SEL24-B489 also demonstrated marked activity in cells bearing FLT3 tyrosine kinase domain (TKD) mutations that lead to FLT3 inhibitor resistance. Moreover, SEL24-B489 inhibited the growth of a broad panel of AML cell lines in xenograft models with a clear pharmacodynamic-pharmacokinetic relationship. Taken together, our data highlight the unique dual activity of the SEL24-B489 that abrogates the activity of signaling circuits involved in proliferation, inhibition of apoptosis and protein translation/metabolism. These results underscore the therapeutic potential of the dual PIM/FLT3-ITD inhibitor for the treatment of AML.

The phenomenon of cancer cell resistance to chemotherapeutics is the main cause of insensitivity to anticancer therapy. Thus, the current challenge remains searching for substances sensitising the activity of cytostatic drugs. In this respect, resveratrol is a very promising therapeutic agent. It has pleiotropic effect on cancer cells, which can play a key role in numerous resistance mechanisms, both classical and atypical. The purpose of the present study was to assess the effect of resveratrol on the inhibition of human pancreatic cancer cell proliferation and on the level of cytostatic resistance-associated proteins. The study was performed on human pancreatic cancer cell lines EPP85-181P (control), EPP85-181RDB (daunorubicin resistance) and EPP85-181PRNOV (mitoxantrone resistance). The effect of resveratrol on the viability and proliferation of the studied cell lines was evaluated by SRB assay, whereas cell cycle arrest and cytostatic accumulation by FACS. Western blot analysis was used to determine the level of P-glycoprotein, topoisomerase II α and β and immunofluorescence technique to visualise the proteins in the cells. Resveratrol inhibited proliferation of all studied cell lines. Phase-specific cell cycle arrest depended on the type of cancer cells. Resveratrol decreased the level and activity of P-gp in EPP85-181RDB cells. In EPP85-181PRNOV cells, expression of both TopoII isoforms increased in a statistically significant manner. The results of in vitro studies support the possibility of potential use of resveratrol in breaking cancer cell resistance to chemotherapeutic drugs.

Despite huge effort spent on understanding the pathogenesis of acute myeloid leukemia (AML), current standards of care are still based on the same chemotherapy agents as two decades ago - combinational treatment of cytarabine with an anthracycline. Fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) is one of the most common genetic lesions in AML. Although FLT3 inhibitors initially exhibit clinical activity, resistance to treatment inevitably occurs within months. PIM kinases are thought to be major drivers of the resistance phenotype and their inhibition in relapsed samples restores cell sensitivity to FLT3 inhibitors. Thus, simultaneous PIM and FLT3 inhibition represents a promising strategy in AML therapy. Selvita has developed a potent and selective first-in-class, dual PIM/FLT3 kinase inhibitor, the SEL24-B489 compound, and profiled its activity for in vitro and in vivo AML models showing significantly broader anti-tumor activity of SEL24-B489 than selective FLT3-ITD or PIM inhibitors. We compared SEL24-B489 head-to-head with a selective PIM inhibitor (AZD1208) and a selective FLT3-ITD inhibitor (AC220) in a panel of AML cell lines with FLT3-ITD or unmutated kinase (FLT3-WT) as well as peripheral AML cells and CD34+ bone marrow blasts. SEL24-B489 exhibited a significantly broader activity, irrespective of FLT3 status, than either of the selective inhibitors. Since PIM kinases have emerged as important mediators of FLT3-inhibitor resistance, we hypothesized that the dual specificity of SEL24-B489 might overcome the phenotype of resistance. We utilized previously developed MOLM-14 cells transduced with either FLT3-WT or FLT3 alleles containing TKD point mutations to show that neither of the these mutations decreased the cellular sensitivity to SEL24-B489. Higher cellular activity and biomarker response of SEL24-B489 than competitive inhibitors was shown by inhibition of specific biomarkers such as S6 and STAT5 phosphorylation at nanomolar concentrations in both FLT3-ITD positive and FLT3-WT cell lines in vitro. We have also demonstrated SEL24-B489 superior potency of SEL24-B489 in xenograft models in vivo. Consistent with the experiments in vitro showing marked synergy between SEL24-B489 and AraC, a combination of these agents resulted in almost completely blocked tumor growth in vivo. Most importantly, SEL24-B489 has been selected as a clinical candidate and is currently in phase I clinical trials. Citation Format: Wojciech Czardybon, Renata Windak, Aniela Gołas, Michał Gałęzowski, Aleksandra Sabiniarz, Izabela Dolata, Magdalena Salwińska, Paweł Guzik, Magdalena Zawadzka, Ewelina Gabor-Worwa, Bożena Winnik, Małgorzata Żurawska, Ewa Kolasińska, Ewelina Wincza, Marta Bugaj, Monika Danielewicz, Grzegorz Dubin, Ewa Jabłońska, Maciej Szydłowski, Tomasz Sewastianik, Bartosz Puła, Anna Szumera-Ciećkiewicz, Monika Prochorec-Sobieszek, Elżbieta Mądro, Ewa Lech-Marańda, Krzysztof Warzocha, Jerome Tamburini, Przemysław Juszczyński, Krzysztof Brzózka. Development of a potent, dual pan-PIM/FLT3 inhibitor for the treatment of heme malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4087. doi:10.1158/1538-7445.AM2017-4087