Abstract 1834: Src inhibition overcomes neratinib resistance in HER2-positive breast cancer

Abstract

Abstract Introduction: Despite the recent successes of HER2-targeted therapies, drug resistance still represents a significant clinical problem. Neratinib, an irreversible pan-HER tyrosine kinase inhibitor, has been recently FDA approved for the extended adjuvant treatment of early-stage HER2-positive breast cancer and little is known of its mechanism of resistance in breast cancer. Using a neratinib-resistant cell line model of HER2-positive breast cancer, HCC1954-N, the aim of this study was to characterise cross-resistance to other HER-targeting tyrosine kinase inhibitors (TKIs) and investigate the role of Src in acquired neratinib resistance. Methods: The effect of HER-family targeting TKIs neratinib, lapatinib, afatinib and, the Src inhibitor, dasatinib on proliferation of HCC1954-N and HCC1954-Par cells was assessed by acid phosphatase assays and three-dimensional Matrigel assays. Cell cycle progression was examined by flow cytometry using a propidium iodide assay. Caspase 3/7-Glo assay was used to assess apoptosis induction. To assess the prevention of the development of neratinib resistance, HCC1954 cells were treated twice weekly with neratinib, dasatinib, or the combination and stained with crystal violet when confluent. Results: The HCC1954-N cell line was resistant to neratinib and to afatinib and lapatinib compared to parental HCC1954, but neratinib was still the most potent of the three anti-proliferative agents. The combination of neratinib and dasatinib was synergistic in the neratinib-resistant HCC1954-N cell line in both adherent (CI value = 0.1 ± 0.03) and three-dimensional (CI value = 0.36 ± 0.02) conditions. Neratinib plus dasatinib did not result in cell cycle arrest; however, the combination caused a significant increase in the sub-G1 cell cycle population (p value = 0.018), indicating induction of apoptosis. This was further validated by increased activation of caspase 3 and 7 (p value = 0.015) in HCC1954-N cells treated with neratinib plus dasatinib. The addition of dasatinib to neratinib treatment prevented the development of neratinib resistance in parental HCC1954 cells. Conclusions: Further pre-clinical investigation of the combination of neratinib plus dasatinib is warranted as a potential therapeutic intervention for patients with neratinib-resistant HER2-positive breast cancer. Citation Format: Neil Conlon, Michelle Lowry, Susan Breslin, Lorraine O'Driscoll, Alexander J. Eustace, John Crown, Norma O'Donovan, Denis M. Collins. Src inhibition overcomes neratinib resistance in HER2-positive breast cancer [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 1834.