Abstract 1889: Synergistic effects of glutaminase and proteasome inhibition in multiple myeloma cell lines

Abstract Background: Multiple Myeloma (MM) accounts for approximately 2.1% of all cancer deaths with a 5-year median survival rate of 49.6%. Besides chemotherapy, treatment options include monoclonal antibodies, immunomodulatory agents, and proteasome inhibitors. Carfilzomib is a selective proteasome inhibitor administered intravenously in patients with relapsed or refractory MM. With cancer cell metabolism emerging as a critical regulator of tumor progression, combining carfilzomib with an inhibitor of the metabolic machinery represents a novel therapeutic strategy to prevent MM progression. RP10107 is a potent, and selective glutaminase (GLS-1) inhibitor that demonstrated high potency against mouse (IC50=21.2 nM), rat (IC50=18.2 nM) and human (IC50=26.4 nM) enzymes with selectivity over GLS-2 (>380-fold). The objective of this study was to evaluate the effect of a combination of carfilzomib and RP10107 in MM cell lines. Methods: Glutamate concentrations in MM cell lines (MM-1S and RPMI-8226) following treatment with RP10107 was estimated by LC-MS/MS. Synergism between RP10107 and carfilzomib was determined using different concentrations of the compounds in a 5 x 5 grid. Synergism, additivity, or antagonism was calculated based on the BLISS score. Apoptosis was determined by Annexin V/7AAD staining using a MUSE® Annexin V and dead cell assay kit (Millipore) while the effect on cell cycle was estimated using Guava Cell Cycle Reagent (Millipore). Expression of cleaved PARP and cleaved caspase-8 were determined by Western Blotting. Results: Treatment of MM cell lines with RP10107 resulted in an increased ratio of glutamine to glutamate with a doubling noticed at concentrations as low as 30 nM. A high degree of synergism between RP10107 and carfilzomib was noticed in both the MM cell lines. Addition of 3 nM Carfilzomib to 30 or 500 nM RP10107 resulted in near complete inhibition of RPMI-8226 and MM-1S cell growth, respectively. Additionally, incubation with the combination (1 μM RP10107 + 15 nM Carfilzomib) for 72 h caused a G0/G1 arrest with a corresponding increase in the percent of apoptotic cells (2.5-fold) compared to the individual agents. Incubating RPMI-8226 cells with a combination of RP10107 and Carfilzomib for 48 h resulted in a 2.2- and 13.6-fold increase in expression of cleaved caspase-8 and cleaved PARP, respectively. Conclusions: Addition of RP10107, a glutaminase inhibitor, potentiated Carfilzomib activity in MM cells. Findings provide a rationale for use of the combination in future clinical trials involving MM patients thereby providing a safer and more effective alternative to currently available therapy. Citation Format: Srikant Viswanadha, Satyanarayana Eleswarapu, Kumar V. Penmetsa, Swaroop Vakkalanka. Synergistic effects of glutaminase and proteasome inhibition in multiple myeloma cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1889.