An androgen receptor pre-mRNA stem loop structure determines splicing fate of AR-V7 in prostate cancer

ABSTRACT: In castration-resistant prostate cancer (CRPC), detection of androgen receptor (AR) splice isoform AR-V7 correlates strongly with resistance to AR pathway inhibitors (ARPI) and poor patient outcomes. AR-V7 incorporates transcript-terminating cryptic exon 3 (CE3), which excludes the ligand binding domain from the AR protein, thereby enabling AR-driven transcriptional regulation independent of androgens. While several CE3 splicing regulatory proteins have been described, the impact of pre-mRNA structures in this region is unknown. We performed SHAPE-MaP to develop the first experimentally informed secondary structure model of the CE3 3' splice site. This model predicted a previously undescribed RNA stem loop structure, which we hypothesized could inhibit AR-V7 biogenesis. Using an AR-V7 minigene assay, we observed decreased AR-V7 abundance in response to stabilization of this structure, leading us to term it the cryptic exon 3-splicing inhibitory stem loop (CE3-SISL). ∆SHAPE analysis supported a protein interaction at this site that is lost in androgen independence. Using quantitative proteomics, we nominated candidate interactors, such as neuro-oncological ventral antigen 1 (NOVA1), that may help explain the repressive function of CE3-SISL. Together, these data support the importance of this RNA structure in AR-V7 biogenesis and suggest a potential target for AR-V7-depleting agents. As a proof of concept, we developed several CE3-SISL-targeting antisense oligonucleotides (ASOs), which showed almost complete inhibition of AR-V7 biogenesis in both the minigene assay and in endogenous AR-V7-expressing cells with or without the ARPI enzalutamide. CE3-SISL is therefore functional in AR-V7 biogenesis and can serve as a structural target in therapeutic development for AR-V7-driven CRPC.

ORGANISM(S): Homo sapiens

PROVIDER: GSE307202 | GEO | 2026/08/29

REPOSITORIES: GEO