Nonviral gene editing via CRISPR/Cas9 delivery by membrane-disruptive and endosomolytic helical polypeptide

Abstract

SignificanceDelivery remains a significant challenge for robust implementation of CRISPR/Cas9. We report an efficient CRISPR/Cas9 delivery system comprising PEGylated nanoparticles based on the α-helical polypeptide PPABLG. Assisted by the high membrane-penetrating ability of the polypeptide, P-HNPs achieved efficient cellular internalization and endosomal escape. The CRISPR/Cas9 delivery system could reach 47.3% gene editing in cells, 35% gene deletion in vivo, and HeLa tumor growth suppression >71%, demonstrating an advantage over the existing conventional polycationic transfection reagents. Efficient also in knock-in and gene activation, the reported CRISPR/Cas9 delivery system serves to advance gene editing in vitro and in vivo.

Delivery of CRISPR/Cas9 for therapeutic genome editing

“…CRISPR/Cas9 plasmid was delivered by RRPHC to knock down the expression of the MutT homolog1 (MTH1), which resulted in more oxidized dNTP accumulation in the ovarian cancer peritoneal metastasis model to induce tumor apoptosis (Figure A) . In another study, Wang et al . developed a versatile gene‐editing platform that exhibited a high membrane‐penetrating ability for delivering the plasmid of Cas9 and sgRNA for cancer treatment.…”

“…The vector of PEGylated nanoparticles was constructed with the component of helical polypeptide poly( γ ‐4‐((2‐(piperidin‐1‐ yl ) ethyl) amino‐methyl) benzyl‐ l ‐glutamate). The nanoparticles showed a perfect deletion efficiency of polo‐like kinase 1 ( Plk1 ) in HeLa tumor tissue, which decreased the expression of Plk1 protein by 66.7%, and significantly inhibited tumor growth, thereby prolonging the survival time of tumor‐bearing mice (Figure B) . Inflammation is an immune response for the process of clearance of foreign invaders, and the generation of the obesity‐associated type 2 diabetes (T2D) has a strong relationship with chronic inflammation.…”

Genetic reprogramming for NK cell cancer immunotherapy with CRISPR/Cas9

“…26 However, in order to achieve gene knock-in by homology-directed repair, donor DNA also needs to be delivered into cells in addition to Cas9 protein and gRNA, therefore demanding higher loading capability of the vehicle. In addition to viral delivery, emerging nanomaterials-based delivery systems might be a possible direction due to the flexibility in modifications of the materials, and some preliminary studies already displayed their potential.…”

Engineering Precision Medicine

“…Current bioinformatics-based precision medicine is mostly based on our capability to read DNA. [187] Many exploratory studies are ongoing to integrate CRISPR into therapies to correct monogenic diseases. The prospect of treating hereditary diseases from their genetic roots sheds light on many untreatable diseases.…”