Abstract 15: Midasin is required for mammosphere stability in Letrozole resistant breast cancer

Abstract While aromatase inhibitors (AI) (i.e., Letrozole) are first-line therapy for postmenopausal women with hormone dependent breast cancer, long-term treatment leads to resistance accompanied by the emergence of breast cancer stem cells (CSCs). Previously, our lab demonstrated that midasin, a novel chaperone protein involved in ribosomal biogenesis was increased 35-fold in the letrozole-resistant mammospheres and high levels of midasin was associated with increased relapse free survival in endocrine resistant breast tumors. Here, we hypothesize that elevated midasin expression poises AI-resistant cells for increased stemness, proliferation, and metastasis. To this end, we cultured AI-sensitive (AC-1) and AI-resistant (LTLT-Ca) cells two- and three dimensionally to study the consequences of midasin alteration on cellular behavior by utilizing a midasin inhibitor (i.e., ribozinoindole-2). Although results from proliferation assays indicate that 1-18 µM Rbin-2 treatment caused a 35-60% reduction in cell proliferation, the AC-1 cells were more sensitive. Similar results were observed when colony formation assays were performed. Overall, the LTLT-Ca cells expressed higher levels of midasin compared to the AC-1 cells, and 18µM Rbin-2 treatment reduced midasin expression in both cell lines. To understand the impact of midasin on the biology of endocrine resistance, epithelial to mesenchymal markers were examined and compared to AC-1 cells, LTLT-Ca cells expressed 3-fold higher levels of N-cadherin which were reduced by Rbin-2 treatment. Although the LTLT-Ca exhibited more mesenchymal properties and were more migratory, Rbin-2 treatment caused a similar dose-dependent decrease in wound healing in both cell lines. Interestingly, when mammosphere formation assays were conducted, Rbin-2 treatment induced mammosphere disintegration in both cell lines. When AC-1 and LTLT-Ca cells were treated with 18 µM Rbin-2, the AC-1 spheres were denser and larger (41.73 µm) while the LTLT-Ca spheres were hollow and smaller (5.80 µm), suggesting that midasin is required for stemness. LTLT-Ca mammospheres exhibited higher levels of N-cadherin and vimentin that were decreased with Rbin-2 treatment. Taken together, these changes suggest that increased midasin expression induces an EMT-like state which may in part be responsible for the mammosphere integrity, and enhanced migration which ultimately aids in maintenance of endocrine resistance. Midasin inhibition reverses this phenotype and may provide a novel strategy to target certain endocrine resistant breast tumors. Citation Format: Bipika Banjara, Mounika Pamukuntla, Manasa Kotina, Afia Ohemeng, Alicia Hudson, Selina Darling-Reed, Jillian L. Pope, Michael Davidson, Syreeta L. Tilghman. Midasin is required for mammosphere stability in Letrozole resistant breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 15.