Abstract P5-05-06: Progesterone receptor (PR) isoform-specific expansion of breast cancer stem-like cells occurs through a phospho-PR/FOXO1 driven signaling axis

Abstract

Luminal breast cancers account for ˜75% of cases and are positive for progesterone receptor (PR) and estrogen receptor (ER) expression. PR is a classical ER-target gene and is used as a biomarker of ER activity; however, a growing body of evidence supports the role of PR as an important modifier of ER actions and a key driver of luminal breast cancer progression. Progesterone signaling is mediated by two PR isoforms: full-length PR-B and truncated PR-A, which lacks the N-terminal 164 amino acids. Little is known about PR isoform-specific actions in PR-expressing breast tumors, given that total PR expression is measured clinically. Herein, we sought to identify phenotypic differences in luminal breast cancer cells (T47D) overexpressing PR-A (T47D-YA) or PR-B (T47D-YB). We demonstrate that PR-B expression is required for anchorage-independent colony formation, while PR-A expressing cells fail to proliferate in soft agar. PR-B driven proliferation has been mapped to PR phosphorylation events, which include MAPK or CDK consensus sites such as Ser294. We demonstrate that in contrast to previous reports, PR-A is well phosphorylated at Ser294 in response to progestins (e.g. R5020) using our custom phospho-PR (Ser294) polyclonal antibody. Interestingly, Ser294 phosphorylation of PR-A occurs more rapidly and robustly following hormone treatment compared to PR-B expressing cells. Our findings indicate that PR-A is a dominant driver of stem-like expansion in breast cancer cells. PR-A tumorspheres exhibited enriched ALDH+ and CD44+/CD24- populations compared to PR-B and promoted heightened gene expression of stem-like genes (e.g. FOXO1). We demonstrate that the PR target gene and co-activator FOXO1 promotes both PR-A and PR-B phosphorylation at Ser294 and augments tumorsphere formation. Direct inhibition of FOXO1 levels abrogates phospho-PR (Ser294) levels and tumorsphere formation in PR expressing cells. Finally, we show that Ser294 is required for PR-A induced expression of stem-like genes (e.g. FOXO1) and stem-like behavior as measured by ALDH+/CD44+ tumorspheres. Taken together, our data reveal unique functions of PR isoforms as modulators of distinct and opposing pathways (i.e. proliferation versus stem-like expansion) in luminal breast cancer models. A clear understanding of PR isoform-specific actions, phosphorylation events, and the role of co-factors such as FOXO1 may lead to novel biomarkers of advanced tumor behavior and reveal new approaches to pharmacologically target PR isoforms in luminal breast cancers. Citation Format: Truong TH, Dwyer AR, Diep CH, Lange CA. Progesterone receptor (PR) isoform-specific expansion of breast cancer stem-like cells occurs through a phospho-PR/FOXO1 driven signaling axis [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P5-05-06.