Collaboration of AR and ERα in conferring resistance to an aromatase inhibitor.

Abstract

579 Background: We have previously shown a role for AR overexpression in tamoxifen resistance in ERα-positive MCF-7 breast cancer cells; here we hypothesized that AR overexpression might similarly be involved in resistance to the aromatase inhibitor anastrazole (Anas). Methods: MCF-7 cells were transfected to express the aromatase gene (MCF-7 Arom), or the aromatase and AR (MCF-7 AR Arom cells). Western blot analysis was used to evaluate protein levels, MTT and soft agar assays to evaluate proliferation, luciferase reporter assays to evaluate transcriptional activities and confocal microscopy was used for localization. Results: Anas inhibited androstendione (AD)-stimulated growth in MCF-7 Arom cells but not in MCF-7 AR Arom cells. Similarly, Anas did not inhibit ERα transcriptional activity in MCF-7 AR Arom cells. Enhanced activation of pIGF-1R, pIRS-1, pAKT, and pMAPK were also observed in AR Arom cells, suggesting constitutive activation of nongenomic signaling in these cells. Consistent with activation of these potential treatment “escape” mechanisms, inhibitors of AKT and IGF-1R restored sensitivity to Anas. Sensitivity to Anas was also restored using the AR antagonist MDV3100, however use of Abiraterone acetate as a single agent most effectively blocked proliferation of AR-overexpressing cells. These results suggest that both AR and ERα must be blocked to restore sensitivity to hormonal therapies in AR overexpressing ERα-positive breast cancers. Unexpectedly, AR contributed to ERα transcriptional activity in MCF-7 AR Arom cells, as shown by inhibition with the AR antagonist bicalutamide. AR and ERα co-localized both in the cytoplasm and in the nucleus of AD+Anas-treated cells, suggesting potential activation of both non-genomic and nuclear-mediated effects when AR is overexpressed in ERα-positive cells. We confirmed these findings in breast cancer cells with acquired resistance to tamoxifen. Conclusions: These results show the necessity to block both AR and ER in patients whose tumors express elevated levels of AR. In addition, inhibitors to the AKT/IGF-1R signaling pathways or direct inhibition of androgen/estrogen synthesis provide alternative approaches to restore hormone sensitivity in resistant breast tumors.