Open AccessEffective inhibition of cancer cells by recombinant adenovirus expressing EGFR-targeting artificial microRNA and reversed-caspase-3
Author(s) -
Mengmeng Yan,
Jia Chen,
Hua Jiang,
Yuqiong Xie,
Chunchun Li,
Lihong Chen,
Baoyu Yang,
Jiashu Cao
Publication year - 2020
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0237098
Subject(s) - mtt assay , cetuximab , flow cytometry , apoptosis , microbiology and biotechnology , in vivo , cancer research , biology , western blot , protein kinase b , chemistry , immunology , antibody , monoclonal antibody , biochemistry , gene
The EGFR-targeting cancer therapies are commonly facing drug resistance, mostly due to mutations. Gene therapy with artificial microRNA targeting EGFR conserved sequence may avoid such problem. In this study, we constructed a recombinant adenovirus expressing EGFR-targeting artificial microRNA and active revCASP3 (Ad-EC), under the control of tumor-specific SLPI promoter, and evaluated its inhibitory effect on HEP-2 cancer cells both in vitro and in vivo . MTT assay showed that cell growth inhibition rate at 72h was 44.0% in Ad-EC group at MOI 50, while the rate was 7.7% in the control virus Ad-GFP group and 3.6% in Cetuximab (500 μg/ml) group respectively. Flow cytometry analysis revealed the late apoptotic cells rate was 36.1% in Ad-EC group, significantly higher than 6.5% of Ad-GFP group ( p < 0.001). When Ad-EC (MOI 50) was combined with CDDP (0.25 μg/ml), late apoptotic cells rate increased to 61.2%, significantly higher than each monotherapy group ( P < 0.001). The real-time xCELLigence system recorded an effective cell growth inhibition in Ad-EC and CDDP groups, and more enhanced effect in Ad-EC plus CDDP group. Western blot revealed that Ad-EC could inhibit the activation of AKT pathway and ERK1/2 pathway, while Cetuximab had the AKT pathway over-activated. In vivo experiments with HEP-2 xenograft in nude mice confirmed the tumor inhibition in Ad-EC, CDDP and Ad-EC plus CDDP groups compared with PBS group ( P < 0.01). Collectively, these data support the effective inhibition of cancer cells by this novel gene therapy strategy.