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Abstract
Androgen receptor (AR) signaling continues to drive disease progression in a large fraction of castration-resistant prostate cancer (CRPC) after androgen deprivation therapy. CRISPR/Cas9-mediated gene editing presents a promising technology to silence the AR gene at the genomic DNA level. However, delivery of CRISPR machinery remains challenging for potential therapeutic application. Here, we developed a platform using extracellular vesicles (EVs) as a vehicle to deliver Cas9/single guide RNA (sgRNA) ribonucleoprotein (RNP) complex to silence the AR gene in prostate cancer (PCa) cells. A genetic modification conferred the N-myristoylation to the Cas9 protein, which enhanced the encapsulation of Cas9/sgRNA RNP into EVs and silenced both ectopic and endogenous AR gene. Interestingly, gene editing efficiency varied across PCa cell lines, associated with different chromatin accessibility at the target site. Functional analyses demonstrated that Cas9/sgRNA RNP (targeting the N-terminal domain of the AR gene) did not change gene-edited AR mRNA levels, but significantly inhibited expression levels of AR downstream genes, and thereby attenuating PCa cell proliferation. Importantly, EV-mediated delivery of the Cas9/sgRNA RNP introduced indels into the AR gene and inhibited proliferation of enzalutamide-resistant PCa cells. This study highlights a therapeutic strategy for treatment of CRPC using a programmable EV-mediated delivery platform.