Abstract 6571: O-GlcNAc sites on progesterone receptor are critical for its ligand independent repression of interferon signaling in breast cancer

Abstract

Abstract The role of hormone receptors in breast cancer has been well established as a key contributor to breast cancer tumorigenesis. Estrogen receptor and progesterone receptors are present in nearly 70% of breast cancers. Estrogen receptor and estrogens in breast cancer has been extensively studied and targeted effectively through endocrine therapy, however the impact of progesterone and the progesterone receptor (PR) independent of estrogen is not well understood. We have previously demonstrated that PR promotes tumor growth and drives an immune suppressive environment, therefore understanding the biology of PR in breast cancer is critical. We demonstrated that PR attenuates type 1 interferon signaling via inhibition of STAT1 phosphorylation and increased degradation of STAT2. In this study we investigated the mechanism of PR regulation of IFN signaling. Post-translational modifications such as phosphorylation and SUMOylation can influence PR target gene promoter selectivity thereby altering its function. We have identified that PR is modified by O-GlcNAc, a single N-acetyl-glucosamine sugar that cycles on and off and serine or threonine amino acids in nuclear, cytoplasmic and mitochondrial proteins. Levels of total O-GlcNAc staining are higher in breast cancer tissue compared to adjacent normal tissue. Active O-GlcNAcylation, as evidenced by immunohistochemistry staining of O-GlcNAc-transferase (OGT), is elevated in patients with PR+ tumors compared to PR-. Using T47D breast cancer cells (an ER/PR-positive tumor line), mass spectrometry analysis revealed an O-GlcNAc site at S499, S733 and S735 on PR. Using a naturally occurring PR-negative variant of T47D cells, we introduced stable expression of a mutant PR with serine to alanine substitution at the O-GlcNAc sites (S499A, S733A, S75A), thereby blocking O-GlcNAc flux, in addition to wt PR as a control. RNA-Seq analysis of these cells following treatment with progesterone revealed ligand dependent PR gene regulation, however major differences were observed in gene expression of the two cell lines independent of ligand. These results indicate ligand independent PR suppression of IFN signaling requires O-GlcNAc at these three sites. Additionally, T47D-mutant PR cell lines implanted into immune deficient mice had significantly decreased growth compared to WT-PR control tumors. Together these findings indicate O-GlcNAcylation of PR contributes to PR driven breast cancer tumor growth and immune evasion via modulation of IFN signaling. Citation Format: Harmony Ivanna Saunders, Sean Holloran, Eilidh Chowanec, Julio Tinoco, Chad Slawson, Christy Hagan. O-GlcNAc sites on progesterone receptor are critical for its ligand independent repression of interferon signaling in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6571.