Abstract 213: NEK1-mediated phosphorylation of YAP1 is key to prostate cancer progression

Abstract The key to preventing metastatic castration-resistant prostate cancer (mCRPC) progression is understanding how androgen-sensitive (AS) PCa cells progress to independence and modify accordingly their transcriptional repertoire. We recently identified a novel axis of the Hippo pathway characterized by the sequential kinase cascade induced by androgen deprivation, AR−mTORTLK1BNEK1pYAP1-Y407, leading to CRPC adaptation. Phosphorylation of YAP1-Y407 increases upon androgen deprivation therapy (ADT), correlated with increased NEK1 activity, and this is suppressed in a YAP-Y407F mutant. Overexpression of wt-GFP-YAP but not the Y407F SDM resulted in dramatic morphologic changes. In fact, LNCaP expressing wt-YAP were AI whereas the Y407F mutant were AS, and LNCaP expressing wt-GFP-YAP underwent EMT transformation. This, largely reflecting transcriptional differences in both AR-dependent genes (FKBP5, PSA) and some involved in EMT (ZEB1, TWIST, E-CAD) ; all of which could be effectively reversed with J54-mediated inhibition of TLK1NEK!YAP signaling. We found that J54, a pharmacological inhibitor of the TLK1NEK1YAP1 nexus led to degradation of YAP1, suggesting that the Y407 phosphorylation is critical for transcriptional activation and stabilization of YAP. Specifically, Nek1-mediated phosphorylation of YAP-Y407 increases its productive interaction with transcriptional activators like TEAD or AR, resulting in its nuclear retention and stabilization. This was later demonstrated by coIP of TEAD4 or the AR with GFP-YAP antiserum, as well as via LUC expression transfections using ARE and Hippo reporter plasmids in LNCaP expressing wt-GFP-YAP or Y407F proteins, as well as by ChIP for key AR-responsive gene promoters. Further, in NEK1 haploinsufficient TRAMP mice we determined reduced YAP1 expression, a key transcriptional activator of CRPC progression, and, if castrated at 12 weeks, the mice failed to show to overt prostate carcinomas, even while displaying reduced E-Cadherin (E-Cad) expression in hyperplastic ductules. Finally, IHC examination of prostate cancer biopsies revealed that pYAP1-Y407 nuclear signal is low in samples of low-grade cancer but elevated in high GS specimens. Citation Format: Damilola M. Olatunde, Ishita Ghosh, Arrigo De Benedetti. NEK1-mediated phosphorylation of YAP1 is key to prostate cancer progression abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 213.