Zahra Noroozi, Mehdi Shamsara, Elahe Valipour, Sahar Esfandyari, Alireza Ehghaghi, Amir Monfaredan, Zahra Azizi, Elahe Motevaseli, Mohammad Hossein Modarressi
Scientific reports 2022 Feb 09Human papillomavirus infections are associated with most cervical cancers, which are the fourth most common cancer in women. HPV-E6 protein binds to protein p53 and inhibits its function, leading to the switching of normal cells toward cancer cells. Here, we disrupted the HPV-E6 gene and investigated its effects on the proliferation and apoptosis of HeLa cells. The HPV18-E6 gene was targeted with two designed sgRNAs cloned into an AAV-CRISPR-based plasmid. The AAV-E6-CRISPR/Cas9 virions were prepared and titrated in HEK293t cells. The cleavage created in the HPV-E6 gene was detected using the T7E1 assay. Cell cycle profiling, MTT assay, and annexin V/PI staining were performed. Also, the p53 protein level was measured by Western blotting. Our data showed that disruption of the HPV-E6 gene led to increased cell apoptosis and decreased cell proliferation. A significant accumulation of infected cells in sub-G1 phase was observed in the cell profiling assay. Also, HPV-E6 gene disruption resulted in a significant increase in the level of P53 protein. Our findings indicated that AAV-mediated delivery of CRISPR/Cas9 can effectively target the HPV-E6 gene in HeLa cells, and its antiproliferative effects may provide therapeutic benefits of local administration of this gene-editing system for HPV-related cervical cancers. © 2022. The Author(s).
Zahra Noroozi, Mehdi Shamsara, Elahe Valipour, Sahar Esfandyari, Alireza Ehghaghi, Amir Monfaredan, Zahra Azizi, Elahe Motevaseli, Mohammad Hossein Modarressi. Antiproliferative effects of AAV-delivered CRISPR/Cas9-based degradation of the HPV18-E6 gene in HeLa cells. Scientific reports. 2022 Feb 09;12(1):2224
PMID: 35140292
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