Title: Dual Therapy with Insulin-Like Growth Factor-I Receptor/Insulin Receptor (IGF1R/IR) and Androgen Receptor (AR) Antagonists Inhibits Triple-Negative Breast Cancer Cell Migration In Vitro

Abstract

Background: Few targeted therapies are currently available for triple-negative breast cancers (TNBC) due to their lack of estrogen receptor-α and progesterone receptor expression and HER-2 amplification. In the clinic, TNBC is associated with a range of adverse biologic features including a highly metastatic phenotype and early patient relapse, leading to poor clinical outcomes. Identifying effective treatment strategies to inhibit or prevent TNBC metastasis could help to improve patient treatment options and clinical outcomes.

Methods: Using immunohistochemistry, archival breast specimens from women with TNBC were screened for androgen receptor (AR), Slug and insulin-like growth factor-2 (IGF2) expression. The effects of IGF2 and combinations of insulin-like growth factor-1 receptor/insulin receptor (IGF1R/IR) and AR antagonists on TNBC cell migration was assessed in vitro using scratch wound assays, and cell death initiated by dual combinations was assessed using caspase assays. In parallel, gel electrophoresis and Western immunoblot assays were used to test effects of IGF1R/IR and AR antagonists on Slug expression. Finally, using the model of Response Additivity, drug synergy was assessed using different therapeutic treatment strategies.

Results: Based on modified Allred scoring results, IGF2, Slug and AR were identified in archival TNBC specimens. Further, IGF2 stimulated migration of TNBC cells MDA-MB-231 and HCC 1937 in vitro. Combination therapies of a) IGF1R/IR inhibitor BMS-754807 plus IGF1R inhibitor NVP-AEW807 or b) AR inhibitor enzalutamide together with either BMS-754807 or NVP-AEW807 led to inhibition of TNBC cell migration (P<0.001) and reduced expression of Slug. Additionally, combination treatments promote TNBC cell apoptotic activity in vitro (P<0.005). Significant synergy was detected (CI < 1) with enzalutamide plus NVP-AEW541 or BMS-754807 in BT549 and HCC 1937 cultures for cell migration in vitro; while combination of BMS-754807 plus NVP-AEW541 elicited antagonist effects (CI > 1) in BT549, MDA-MB-231 and HCC 1937 cultures.

Conclusion: These findings indicate that combinations of IGF1R/IR and AR antagonists can prevent TNBC migration, induce apoptosis and impact known mediators of metastasis, thus suggesting a potential for future application of such dual treatments to manage TNBC progression in the clinic.