In Vitro Prostate Cancer Treatment via CRISPR‐Cas9 Gene Editing Facilitated by Polyethyleneimine‐Derived Graphene Quantum Dots

Abstract

AbstractCRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)‐Cas9 (CRISPR associated protein 9) is a programmable gene editing tool with a promising potential for cancer gene therapy. This therapeutic function is enabled in the present study via the non‐covalent delivery of CRISPR ribonucleic protein (RNP) by cationic glucosamine/PEI‐derived graphene quantum dots (PEI‐GQDs) that aid in overcoming physiological barriers and tracking genes of interest. PEI‐GQD/RNP complex targeting the tumor protein 53 (TP53) gene mutation overexpressed in ∽50% of cancers successfully produces its double‐stranded breaks in solution and in prostate cancer (PC‐3) cells. Restoring this cancer “suicide” gene can promote cellular repair pathways and lead to cancer cell apoptosis. Its repair to the healthy form performed by simultaneous PEI‐GQD delivery of CRISPR RNP and a gene repair template leads to a successful therapeutic outcome: 40% apoptotic cancer cell death, while having no effect on non‐cancerous (HeK293) cells. The translocation of PEI‐GQD/RNP complex into PC‐3 cell cytoplasm is tracked via GQD intrinsic fluorescence, while enhanced green fluorescent protein (EGFP)‐tagged RNP is detected in the cell nucleus, showing the successful detachment of the gene editing tool upon internalization. Using GQDs as non‐viral delivery and imaging agents for CRISPR‐Cas9 RNP sets the stage for image‐guided cancer‐specific gene therapy.