21/08/1402
The chicken chorioallantoic membrane model for isolation of CRISPR/cas9-based HSV-1 mutant expressing tumor suppressor p53
Oncolytic viruses (OVs) have emerged as a novel cancer treatment
modality, which selectively target and kill cancer cells while sparing
normal ones. Among them, engineered Herpes simplex virus type 1 (HSV-1)
has been proposed as a potential treatment for cancer and was moved to
phase III clinical trials. Previous studies showed that design of OV
therapy combined with p53 gene therapy increases the anti-cancer
activities of OVs. Here, the UL39 gene of the ICP34.5 deleted HSV-1 was
manipulated with the insertion of the EGFP-p53 expression cassette
utilizing CRISPR/ Cas9 editing approach to enhance oncoselectivity and
oncotoxicity capabilities. The ΔUL39/Δγ34.5/HSV1-p53 mutant was isolated
using the chorioallantoic membrane (CAM) of fertilized chicken eggs as a
complementing membrane to support the growth of the viruses with gene
deficiencies. Comparing phenotypic features of
ΔUL39/Δγ34.5/HSV1-p53-infected cells with the parent Δγ34.5/HSV-1 in
vitro revealed that HSV-1-P53 had cytolytic ability in various cell
lines from different origin with different p53 expression rates.
Altogether, data presented here illustrate the feasibility of exploiting
CAM model as a promising strategy for isolating recombinant viruses
such as CRISPR/Cas9 mediated HSV-1-P53 mutant with less virus
replication in cell lines due to increased cell mortality induced by
exogenous p53.