Oncogenic potential of truncated-Gli3 via the Gsk3β/Gli3/AR-V7 axis in castration-resistant prostate cancer

The functional activation of the androgen receptor (AR) and its interplay with the aberrant Hh/Gli cascade are pivotal in the progression of castration-resistant prostate cancer (CRPC) and resistance to AR-targeted therapies. Our study unveiled a novel role of the truncated form of Gli (t-Gli3) in advancing CRPC. Investigation into Gli3 regulation revealed a Smo-independent mechanism for its activation. Despite lacking a transactivation domain, t-Gli3 relies on androgen receptor variant 7 (AR-V7) for its action. Mechanistically, Gsk3β activation led to the t-Gli3 generation, and inhibition of Gsk3β supported the accumulation of full-length Gli3 expression through a non-canonical mechanism. Knockdown of Gsk3β (Gsk3β KD) reduces CRPC cell proliferation, induces apoptosis via mitochondrial fragmentation, and triggers metabolomic reprogramming. The in vivo studies with Gsk3β KD cells in the mouse prostate resulted in tumor growth retardation compared to scramble cells. RNA-seq HALLMARK Gene Set Enrichment Analysis (GSEA) analysis of Gsk3β KD revealed a positive enrichment of apoptosis, tumor suppressor gene, and negative enrichment of oncogenic pathway. Furthermore, combinational use of a Gsk3β inhibitor with anti-Smo or Gli1 significantly inhibited the CRPC cell growth, which is resistant to individual Smo or Gli1 inhibitor targeting. Intriguingly, solely targeting Gli3 showed effectiveness in inhibiting CRPC cell growth. Overall, our study underscores the clinical significance of Gli3, emphasizing t-Gli3, and provides novel insights into the interplay of the Gsk3β/t-Gli3/AR-V7 axis in CRPC.