CRISPR/Cas9-mediated knock-in of BRCA1/2 mutations restores response to olaparib in pancreatic cancer cell lines

• Published in: Scientific reports Vol. 13; no. 1; p. 18741
• Main Authors: Witz, Andréa, Dardare, Julie, Francois, Aurélie, Husson, Marie, Rouyer, Marie, Demange, Jessica, Merlin, Jean-Louis, Gilson, Pauline, Harlé, Alexandre
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 31.10.2023; Nature Publishing Group; Nature Portfolio
• Subjects: 631/1647/1513/1967/3196; 631/67/1059; 692/4028; Adenocarcinoma; Adenocarcinoma - genetics; Apoptosis; BRCA1 protein; BRCA1 Protein - genetics; BRCA2 protein; BRCA2 Protein - genetics; Breast cancer; CRISPR; CRISPR-Cas Systems; Germ-Line Mutation; Humanities and Social Sciences; Humans; Life Sciences; MCF-7 Cells; Metastases; multidisciplinary; Mutation; Pancreatic cancer; Pancreatic Neoplasms; Pancreatic Neoplasms - drug therapy; Pancreatic Neoplasms - genetics; Pancreatic Neoplasms - pathology; Phthalazines - pharmacology; Science; Science (multidisciplinary); Transfection; Tumor cell lines
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-023-45964-w

Pancreatic cancer is one of the most aggressive diseases with a very poor outcome. Olaparib, a PARP inhibitor, as maintenance therapy showed benefits in patients with metastatic pancreatic adenocarcinoma bearing germline BRCA1/2 mutations. However, germline BRCA mutation has been described in only 4–7% of patients with pancreatic adenocarcinoma. A CRISPR/Cas9-mediated system was used to knock-in the c.763G > T p.(Glu255*) and c.2133C > A p.(Cys711*) mutations in cell lines to obtain truncated BRCA1 and BRCA2 proteins, respectively. A CRISPR/Cas9 ribonucleoprotein complex was assembled for each mutation and transfected into two pancreatic cell lines (T3M4 and Capan-2) and into a breast cancer cell lines (MCF7) as control. BRCA protein levels were significantly decreased in all BRCA-depleted cells ( P  < 0.05), proving the transfection efficiency of our CRISPR/Cas9 systems. As expected, the calculated olaparib IC50 were significantly reduced for all cell lines harbored BRCA1 or BRCA2 mutations compared to wild-type BRCA1 / 2 cells ( P  < 0.01). Furthermore, we observed a higher induction of apoptosis after 72 h olaparib treatment in BRCA-depleted cells than in wild-type cells. This strategy might offer new insights into the management of patients with pancreatic cancer and open up new perspectives based on the in vivo use of CRISPR/Cas9 strategy.