Abstract 4088: Th1 cytokines and oncodriver inhibition: A combination treatment strategy in TNBC

Abstract

Abstract Triple Negative Breast Cancer (TNBC) is characterized by lack of hormone receptors and targeted therapies, resistance to existing treatment and poor survival outcome, highlighting the need for novel therapy development. Oncogene addiction in breast cancer (BC) cells to drive malignancy up presents oncodrivers as promising therapeutic candidates in BC. Our group has shown loss of anti-oncodriver CD4+-T-helper-type-1 (Th1) response in TNBC patients. Here, we studied effects of combination treatment with Th1 cytokines (IFN-γ+TNF-α) and oncodriver blockade in human and mouse TNBC cells (MDA-MB-231, MDA-MB-468, HCC1143, BT549, 4T1). We measured expression of 3 oncodriver proteins: EGFR, HER3 and cMET and observed multiple oncodrivers in majority of the cell lines, suggesting combined oncodriver inhibition may have stronger antitumor effects. Based on oncodriver status, we tested combination treatment with Th1 cytokines and a) monoclonal α-EGFR antibody cetuximab, b) cMET RTK inhibitor crizotinib and c) siRNA inhibition of three oncodrivers. No significant difference in proliferation after IFN-γ or IFN-γ+TNF-α treatment suggested IFN-γ is the primary effector cytokine. IFN-γ alone reduced proliferation >50% in majority of TNBC cells. In EGFRhigh/HER3pos/cMETlow MDA-MB-468, IFN-γ and cetuximab treatment significantly increased apoptosis compared to untreated cells. Combined EGFR and HER3 siRNA inhibition, followed by IFN-γ treatment, increased apoptosis in those cells. In EGFRpos/HER3low/cMEThigh MDA-MB-231 and EGFRlow/HER3high/cMEThigh 4T1, IFN-γ and crizotinib combination significantly decreased proliferation (p<0.0001) compared to single agent. In EGFRpos/HER3pos/cMETpos HCC1143, combined siRNA inhibition of 3 oncodrivers followed by IFN-γ treatment increased apoptotic signal compared to control. Protein expression showed increased STAT1Tyr701 phosphorylation as primary mechanism of IFN-γ mediated proliferation inhibition. Western blot after cetuximab/crizotinib and Th1 cytokines treatment suggests altered phosphorylation of signaling effectors, PI3K/AKT, p44/42MAPK and STATs 1, 3 and 5 may contribute to anti-proliferative, apoptotic effects of combination treatment. siRNA inhibition of one oncodriver resulted in a compensatory increase in expression of the other oncodriver (increased HER3 level in siEGFR transfected MDA-MB-468 cells and vice versa), suggesting a complex signaling crosstalk between oncodrivers in TNBC. We are currently investigating effects of combination treatment on cellular distribution of oncodrivers, % of ALDHpos stem-like cells, cytokines and chemokines secretion profile and protein expression in TNBC cells. Our study suggests combination treatment with Th1 cytokines and oncodriver inhibition in TNBC may contribute to improved therapeutic benefit and clinical outcome. Citation Format: Amrita Basu, Yongsheng Jia, Krithika Kodumudi, Doris Wiener, Brian Czerniecki. Th1 cytokines and oncodriver inhibition: A combination treatment strategy in TNBC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4088.