CRISPR/Cas9 model of prostate cancer identifies Kmt2c deficiency as a metastatic driver by Odam/Cabs1 gene cluster expression

Abstract

Prostate cancer (PCa) is the second leading cause of cancer-related deaths in men, and treatment options for metastatic disease remain limited. In this study, we employed Adeno-associated virus (AAV) delivery and CRISPR-Cas9 technology to simultaneously mutate multiple genes in the mouse prostate, in order to investigate positive genetic interactions during PCa progression. By targeting five potential tumor suppressor genes (Pten, Trp53, Rb1, Stk11, and RnaseL), we induced advanced prostate tumors in mice without metastasis, and mice reached humane endpoint at eight weeks. When three epigenetic factors (Kmt2c, Kmt2d, and Zbtb16) were further depleted, tumor progression remained identical, but metastases were observed in the lung of all mice. Whole genome sequencing of metastatic tumors revealed few genetic alterations, confirming that mutations in the targeted genes are sufficient to transform prostatic cells into metastatic cancer. Mechanistic analysis uncovered multiple altered pathways, including impaired p-Src/p-Lyn-cMyc signaling, underscoring the importance of cMyc signaling in PCa metastasis. RNA sequencing revealed a significant dysregulation of overall transcriptome and increased expression of numerous genes in a genomic region at chr5qE1, which is highly conserved to human chr4q13.3 and regulated by KMT2C. Intriguingly, the depletion of Odam and Cabs1 in this gene cluster dismissed metastasis formation, indicating that they have critical functions for secondary tumor formation, which has not been reported before. Further genetic analysis demonstrated that Kmt2c deficiency is sufficient to facilitate lung metastasis, identifying Kmt2c as an essential gatekeeper for metastatic formation. Interestingly, the gene expression signature from mutations of the three epigenetic regulators were predictive of progression-free and overall survival and able to distinguish primary and metastatic human prostate cancer. Overall, this study highlights the positive genetic interactions between classical tumor suppressor genes and epigenetic modulators in the progression and onset of metastatic disease, providing predictions and insights into possible treatments for metastatic PCa.