A Blue Light-Inducible CRISPR-Cas9 System for Inhibiting Progression of Melanoma Cells

Xia Wu,Jianzhong Zhang,Bo Yu,Haiyan Huang

doi:10.3389/fmolb.2020.606593

Abstract

Melanoma is an aggressive skin tumor that shows a high mortality rate and level of metastasis. BRAF gene mutation (BRAF V600E) is directly related to the occurrence of melanoma. In this study, a light-inducible gene expression system was designed to control the Cas9 transcription, which could then cleave the BRAF V600E. To prove the potential utility of this system in melanoma, the physiological function of melanoma cells was tested. It illustrated that the light-induced CRISPR-Cas9 system could inhibit the progression of G361 and A375 cells. Thus, this system may provide a novel therapeutic strategy of melanoma intervention.

Highlights

Melanoma is one of the most malignant skin tumors
After the action of Cas9 and sgRNA, the doublestrand DNA at the target site was cleaved to form a doublestrand break (DSB), and the mKate was activated through cell homologous recombination
The gRNA activity assay demonstrated that the CRISPR-Cas9 system cleaved the mutant BRAF V600E (Supplementary Figure S1), but had no effect on the wild type

Summary

Introduction

Melanoma is one of the most malignant skin tumors. Despite its low prevalence rate, its incidence increases every year (Hernandez-Davies et al, 2015; Chang et al, 2020; Strub et al, 2020). Over the past years, moleculetargeted drugs – such as BRAF and MEK inhibitors have been used in clinical practice (Luke et al, 2017; Dummer et al, 2018a,b; Chavda and Bhatt, 2020). Drug resistance often occurs after treatment for 2–18 months (Chang et al, 2020). To understand the underlying mechanism of this resistance, previous researchers found the reactivation of the BRAF/MEK/ERKs pathway accounted for 80% of the resistant tumors (Rizos et al, 2014). The PI3-K/AKT pathway that is closely interacted with the BRAF/MEK/ERKs could be another cause of drug resistance when reactivated (Davies, 2012; Rizos et al, 2014)

Methods

Results

Conclusion