The Establishment of Prostate-specific, SKP2 Humanized Mice by CRISPR Knock-in Method Reveals Neoplastic Initiation and Microenvironmental Reprogramming

Abstract

A recent study has shown that SKP2 inactivation can prevent cancer initiation by extension of total cell cycle duration without perturbing normal division, which suggests a new strategy for cancer prevention. However, direct in vivo evidence for human SKP2 on cancer initiation and prostatic microenvironment is still lacking and a prostate-specific SKP2 humanized mouse model is critical for developing prostate cancer immunoprevention approaches through targeting human SKP2. We therefore have established a prostate-specific human SKP2 (hSKP2) knock-in mouse model by a CRISPR knock-in approach. Overexpression of hSKP2, which is driven by an endogenous mouse probasin promoter, induces prostatic lesions including hyperplasia, mouse prostate intraepithelial neoplasia (mPIN), and low-grade carcinoma and increases prostate weights. Transcriptional profiling by RNA-sequencing analysis revealed significant gene expression alterations in epithelial to mesenchymal transition (EMT), extracellular matrix, and interferon signaling in the prostate of hSKP2 knock-in mice compared to wild-type mice. Single cell deconvolution showed an increase of fibroblasts population and a decrease of CD8+ T cell and B cell populations in the prostate of hSKP2-knock-in mice. Consistently with these results from the SKP2 humanized mouse, overexpression of hSKP2 in human prostate cancer PC3 cells markedly increased cell migration and invasion and induced the gene expression of EMT and interferon pathways, including FMOD, THY1, PFKP, USP18, IL15, etc. In addition, paired prostate organoids were derived from SKP2 humanized and wild-type mice for drug screening and validated by known SKP2 inhibitors, Flavokawain A and C1. Both of which selectively decrease the viability and alter the morphologies of organoids of hSKP2 knock-in rather than wild-type mice. Our studies provide a well-characterized prostate-specific hSKP2 knock-in mouse model and offer new mechanistic insights for understanding the oncogenic role of SKP2 in shaping the prostatic microenvironment during early carcinogenesis.