HPV Oncogene Manipulation Using Nonvirally Delivered CRISPR/Cas9 or Natronobacterium gregoryi Argonaute

Advanced Science
Yeh-Hsing LaoKam W Leong

Abstract

CRISPR/Cas9 technology enables targeted gene editing; yet, the efficiency and specificity remain unsatisfactory, particularly for the nonvirally delivered, plasmid-based CRISPR/Cas9 system. To tackle this, a self-assembled micelle is developed and evaluated for human papillomavirus (HPV) E7 oncogene disruption. The optimized micelle enables effective delivery of Cas9 plasmid with a transient transgene expression profile, benefiting the specificity of Cas9 recognition. Furthermore, the feasibility of using the micelle is explored for another nucleic acid-guided nuclease system, Natronobacterium gregoryi Argonaute (NgAgo). Both systems are tested in vitro and in vivo to evaluate their therapeutic potential. Cas9-mediated E7 knockout leads to significant inhibition of HPV-induced cancerous activity both in vitro and in vivo, while NgAgo does not show significant E7 inhibition on the xenograft mouse model. Collectively, this micelle represents an efficient delivery system for nonviral gene editing, adding to the armamentarium of gene editing tools to advance safe and effective precision medicine-based therapeutics.

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Methods Mentioned

BETA
transfection
FACS
confocal microscopy
PCR
gene knockdown
transmission electron microscopy
antisense oligonucleotide
xenograft
transfect
Scanning

Software Mentioned

Protein BLAST
Cas9
Cas
FlowJo
BLAST
OFFinder

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