miR-548ah-3p suppresses ovarian cancer stemness and therapeutic resistance through network-level signaling reprogramming within a distinct alternative polyadenylation landscape

Abstract

Therapeutic resistance and cellular plasticity remain major challenges in high-grade serous ovarian cancer (HGSOC), yet the RNA-mediated mechanisms sustaining stem-like states remain incompletely understood. Here, we investigated microRNA-driven signaling reprogramming and alternative polyadenylation (APA) dynamics in an ovarian cancer stem cell (OCSC)-enriched spheroid model. Small RNA sequencing and targeted expression analyses identified reduced miR-548ah-3p abundance across OVCAR-3 spheroids, the evaluated drug-resistant ovarian cancer models, an additional HGSOC spheroid model, and therapy-resistant clinical tumor samples. Functional restoration of miR-548ah-3p suppressed stemness-associated phenotypes, impaired migration and invasion, and re-sensitized spheroids to platinum- and PARP inhibitor-based therapies. Transcriptome profiling revealed partial reversal of OCSC-associated transcriptional programs following prolonged miR-548ah-3p restoration. Integration of differential expression and target prediction analyses identified candidate rescue genes enriched in stemness, epithelial–mesenchymal transition (EMT), and survival-associated signaling pathways, supporting a network-level regulatory role rather than isolated single-target effects. APA analysis revealed a pronounced directional bias toward distal poly(A) site usage in spheroids, consistent with a lengthened 3′UTR landscape. miR-548ah-3p restoration reversed spheroid-associated gene expression patterns without broadly remodeling APA profiles, suggesting that signaling reprogramming occurs within a relatively stable isoform architecture. Together, these findings support miR-548ah-3p as a regulator of stemness-associated phenotypes in the investigated HGSOC models and of therapeutic resistance in the OVCAR-3 spheroid model.