Pharmacologic targeting of midasin (MDN1) reveals a potential therapeutic vulnerability in ESR1-mutant breast cancer.

INTRODUCTION: Although most cases of estrogen receptor-positive (ER+) breast cancer initially respond to endocrine therapy, many patients ultimately develop resistance. A major contributor to endocrine resistance in metastatic disease is the acquisition of constitutively active somatic mutations in the estrogen receptor ligand-binding domain (LBD). We previously identified midasin (MDN1), a ribosome biogenesis protein, as significantly overexpressed in letrozole-resistant MCF-7 cells. Because these cells are ERlow/- and represent only a subset of endocrine-resistant tumors, we hypothesized that ESR1 mutations cooperate with MDN1 dysregulation to confer a survival advantage.

METHODS: To address this, the cBioPortal database was queried to assess correlations between breast cancer subtypes and MDN1 expression levels. MCF-7 cell lines harboring ER point mutations were evaluated by RNA sequencing and immunoblot analysis to measure ER and MDN1 expression. To identify pharmacologic inhibitors of midasin, computational docking analyses were performed using a panel of ribozinoindole (Rbin) analogs, followed by biological evaluation studies.

RESULTS: Initial analyses demonstrated that MDN1 expression is elevated in human breast cancer tumors, including luminal, HER2+, and triple-negative breast cancer. RNA sequencing of parental MCF-7 cells and ESR1-mutant derivatives, MCF-7 Y537S and MCF-7 D538G, revealed comparable MDN1 transcript levels across all cell lines. In contrast, immunoblot analysis showed mutation-dependent differences in MDN1 protein expression, with the highest levels observed in MCF-7 D538G cells, followed by MCF-7 Y537S cells and then parental MCF-7 cells. Computational docking analyses of Rbin analogs led to the selection of Rbin-1 and Rbin-2 for biological evaluation. Viability assays revealed minimal activity for Rbin-1, whereas Rbin-2 reduced proliferation by 30%-55% across all three cell lines, with the most pronounced effects observed in MCF-7 D538G cells at 24 and 48 h. Consistent with these findings, Rbin-2 treatment decreased MDN1 protein expression by approximately 50% in all cell lines, while ER levels remained largely unchanged.

DISCUSSION: Collectively, these results establish the feasibility of pharmacologically targeting midasin in mammalian cell lines and support a functional link between MDN1 expression and ESR1 mutation-driven endocrine resistance. This work provides a foundation for future mechanistic studies of midasin as a potential therapeutic vulnerability in ER-mutant breast cancer.