Dataset Information

The BRCA2 p.N372 H i.a.1342A>C Could Regulate the Sensitivity of Ovarian Cancer Cells to Platinum-Based Drugs.

ABSTRACT:

Background and objective

We have previously reported that BRCA2 N372 H i.a.1342A>C heterozygous variation presented in platinum-resistant patients. This study aimed to further investigate the mechanism of BRCA2 N372 H mutation in the development of platinum resistance in ovarian cancer.

Methods

The BRCA2 N372 H i.a.1342A>C was synthesized and used to exchange 1 wildtype allele followed by sequencing to confirm the mutant allele sequence. Plasmids were constructed and transfected into the OVCAR-3 cells after lentiviral packaging. BRCA2 N372 H mRNA was detected by qPCR. BRCA2 protein was assessed by immunoblotting. Binding of the BRCA2 to Rad51 was detected by immunofluorescence staining. Sensitivity of the cells to cisplatin treatment was assessed with CCK-8 assay.

Results

It was found that expression of BRCA2 protein in ovarian cancer cells transfected with BRCA2 N372 H i.a.1342A>C gene (2.177 ± 0.003) was significantly increased compared to that of the cells transfected with lenti-EGFP only (1.227 ± 0.003, P < 0.001). Binding of the BRCA2 and Rad51 proteins was significantly increased in the cells with BRCA2 N372 H i.a.1342A>C mutation (3.542 ± 0.24) than that in the cells transfected with lenti-EGFP (1.29 ± 0.32) or empty cells (1.363 ± 0.32, P < 0.001). Cell viability significantly increased in the cells transfected with BRCA2 N372 H mutant gene. The IC50 value was significantly higher in the cells transfected with BRCA2 N372 H mutant gene (1.963 ± 0.04) than that of the cells transfected with lenti-EGFP (0.955 ± 0.03, P < 0.01) or empty cells (1.043 ± 0.007, P < 0.01).

Conclusion

Over expression of mRNA and protein of BRCA2 was detected in the cells with BRCA2 N372 H i.a.1342A>C mutation but not in the lentivirus negative control (lenti-EGFP) or the cells without transfection (empty cells), which may lead to resistance to platinum-based drugs in ovarian cancer cells through homologous recombination repair pathway.

SUBMITTER: Du ZH

PROVIDER: S-EPMC7768310 | biostudies-literature | 2020 Jan-Dec

REPOSITORIES: biostudies-literature

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The <i>BRCA2 p.N372 H i.a.1342A>C</i> Could Regulate the Sensitivity of Ovarian Cancer Cells to Platinum-Based Drugs.

Du Zhen-Hua ZH Xia Yu Y Yang Qing Q Gao Song S

Technology in cancer research & treatment 20200101

<h4>Background and objective</h4>We have previously reported that <i>BRCA2 N372 H i.a.1342A>C</i> heterozygous variation presented in platinum-resistant patients. This study aimed to further investigate the mechanism of <i>BRCA2 N372 H</i> mutation in the development of platinum resistance in ovarian cancer.<h4>Methods</h4>The <i>BRCA2 N372 H i.a.1342A>C</i> was synthesized and used to exchange 1 wildtype allele followed by sequencing to confirm the mutant allele sequence. Plasmids were construc ...[more]

PMID: 33357097

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