Abstract PD2-01: Clinically dormant ER+ breast tumors exhibit AMPK activation

While adjuvant anti-estrogen therapy has shown immense clinical benefit for patients with estrogen receptor-positive (ER+) breast cancer, >30% of patients experience cancer recurrence within 15 years of initial diagnosis. Anti-estrogen therapies inhibit ER activity either directly (tamoxifen, fulvestrant) or by reducing systemic estrogen levels (aromatase inhibitors, AIs). Extending adjuvant anti-estrogen therapy from 5 to 10 years further prevents recurrence; however, with both 5- and 10-year treatment regimens, a large proportion of patients who relapse do so after cessation of therapy (“late recurrence”). Additionally, disseminated tumor cells in bone marrow have been found after 4 years of adjuvant anti-estrogen therapy in “disease-free” patients. These data collectively indicate that anti-estrogens elicit clinical benefit as adjuvant therapies, in part, by maintaining a population of residual micrometastatic cancer cells in a “clinically dormant” state (i.e., undetectable by routine clinical methods). Understanding how such dormant cancer cells survive will enable the development of more effective adjuvant therapies. We developed several luciferase-labeled xenograft models of ER+ breast cancer that recapitulate clinical dormancy in vivo. Low systemic levels of estrogen in mice can be further suppressed by ovariectomy, mimicking the effects of AI-induced estrogen deprivation seen in patients. In ovariectomized mice, palpable tumors form upon 17b-estradiol supplementation, but quickly regress upon estrogen withdrawal. While regressed tumors become non-palpable within 2 wk, a small proportion of cancer cells survive these estrogen-deprived conditions for >4 months in a clinically dormant, growth-suppressed state. This estrogen deprivation-induced clinically dormant cell population retains tumorigenic potential, as 17b-estradiol retreatment induces tumor recurrence. RNA expression profiling revealed AMPK alpha 2 as one of the most highly expressed genes in clinically dormant residual tumor cells compared to acutely estrogen-withdrawn tumors. Increased AMPK kinase activity was confirmed through immunohistochemical analysis of phospho-ACC, and AMPK substrate. AMPK activation using glucose deprivation or the anti-diabetes drug metformin promoted estrogen-independent survival and growth of ER+ breast cancer cells in vitro. Metformin treatment may also slow estrogen withdrawal-induced tumor regression and promote tumor cell survival in ER+ breast cancer xenografts. As a cellular energy sensor, AMPK has been shown to promote autophagy, a process linked with anti-estrogen resistance. Immunofluorescent staining of estrogen deprivation-induced clinically dormant residual tumor cells revealed decreased levels of the autophagy marker p62 compared to 17b-estradiol-driven tumors. Early data suggest that inhibition of autophagy with hydroxychloroquine abrogates cell survival conferred by metformin in estrogen-depleted conditions. Thus, AMPK may be promoting the survival of ER+ breast cancer cells in estrogen-deprived conditions by increasing autophagic flux. These data have implications for the interpretation of data from ongoing clinical studies testing metformin for the treatment of cancer. Citation Format: Hampsch RA, Dillon LM, Miller TW. Clinically dormant ER+ breast tumors exhibit AMPK activation [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr PD2-01.