Abstract C045: RP4010, a small molecule inhibitor of Store-Operated Calcium Entry (SOCE), potentiates activity of Gilteritinib and Cytarabine in preclinical models of AML

Abstract

Abstract Background: Store-operated calcium entry (SOCE) through calcium release activated calcium (CRAC) channels contribute to calcium influx in non-excitable cells. Elevation of cytosolic calcium through SOCE mediates an array of cellular responses including metabolism and gene expression, cell growth, and proliferation. Aberrant CRAC activity has been linked to various auto-immune disorders and certain cancers via the NFAT pathway. Calcium signaling via CRAC channels also mediates several physiological processes including cell proliferation and apoptosis. RP4010 is a potent inhibitor of CRAC channel activity (IC50=60 nM) with demonstrated efficacy across a range of cancer cell lines in vitro. The objective of this study was to evaluate the single agent activity and the combination of RP4010 and standard of care drugs in AML models in vitro and in vivo. Methods: Cell growth following incubation with RP4010 (5 μM), Gilteritinib (0.25-0.75 μM), or a combination of both agents was determined in the AML lines: THP-1, HL-60, KG-1, and U-937. Apoptosis was evaluated following incubation of cell lines (THP-1 and U-937) with compounds for 72 h, subsequent staining with Annexin-V-PE and 7-AAD, and analysis by flow cytometry. For cell cycle, cells were incubated with compounds for 72 h, fixed with 70% ethanol, incubated at 4°C for 3 h, and stained with Propidium Iodide prior to analysis by flow cytometry. AKT and Stat-5 phosphorylation was determined by Western blotting. In vivo efficacy was evaluated in a MV-4-11 mouse xenograft model. Results: Store-operated calcium entry with RP4010 potentiated Gliteritinib activity manifested by a significant (P<0.05) reduction in cell growth when compared to the activity of the individual agents. Similarly, the combination (3 µM RP4010 + 1 or 2 µM Gilteritinib) led to a 13% increase in the number of apoptotic cells besides increasing cells in the G0/G1 phase by 17%. Combining RP4010 (1 µM) and Gilteritinib (0.1 µM) reduced AKT and Stat-5 phosphorylation by 45 and 57%, respectively, when compared to Gilteritinib in the THP-1 cell line. In the MV-4-11 mouse xenograft model, RP4010 when administered orally at 30 mg/kg/BID for 21 days demonstrated significant (P<0.001) anti-tumor activity both as a single agent and in combination with Cytarabine with tumor growth inhibitions of 42 and 68 %, respectively. Both RP4010, as well as the combination of RP4010 + Cytarabine, were well tolerated throughout the study period with no appreciable change in body weight. Conclusions: RP4010, a potent SOCE inhibitor, attenuated AML tumor growth as a single agent besides demonstrating remarkable potentiation in activity of approved/standard of care drugs for treating AML. Findings provide a rationale for use of this molecule in clinical trials involving naïve or relapsed/refractory AML patients. RP4010 is currently being evaluated in a Phase-1 dose-escalation study in patients with relapsed/refractory lymphomas. Citation Format: Srikant Viswanadha, Satyanarayana Eleswarapu, Kumar V. Penmetsa, Swaroop Vakkalanka. RP4010, a small molecule inhibitor of Store-Operated Calcium Entry (SOCE), potentiates activity of Gilteritinib and Cytarabine in preclinical models of AML [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C045. doi:10.1158/1535-7163.TARG-19-C045