Synthetic Lethal Interaction Between BRCA1 and FEN1 in Ovarian Cancer Cells
- Author(s): Huang, Yueyang
- Advisor(s): Wang, Jean Y.J.
- et al.
The concept of synthetic lethality (SL) can be applied to selectively kill cancer cells by targeting SL-partners of a cancer-specific mutation. The SL-interaction between homologous recombination (HR) and the flap endonuclease was first established in yeast, and recently confirmed in human cells by showing that HR-deficient (BRCA1-deficient) cell lines are more sensitive to flap endonuclease 1 (FEN1)-inhibitors than BRCA1-proficient cell lines. To further establish that BRCA1-loss is sufficient to cause FEN1-inhibitor sensitivity in ovarian cancer, I used the CRISPR/Cas9 gene-editing methods to correct the BRCA1 mutation in the FEN1-inhibitor sensitive UWB1.289 cells, and to knockout BRCA1 in the FEN1-inhibitor resistant OV90 cells, followed by measuring the response of BRCA1-edited clones to FEN1-inhibitor SMD2485. With UWB1.289, I was unable to propagate plasmid-transfected cells, suggesting a possible sensitivity to exogenous DNA. With OV90, I found that the majority of edited BRCA1-alleles produced proteins with small in-frame deletions (∆BRCA1) and that the few null-alleles underwent further mutations to produce ∆BRCA1 upon propagation in culture. I also found that ∆BRCA1-expressing clones were as resistant as the parental OV90 cells to SMD2485. Due to the sensitivity of UWB1.289 cells to transfection and the instability of BRCA1-null alleles in OV90 cells, this study failed to establish the sufficiency of BRCA1-loss in causing FEN1-inhibitor sensitivity. Alternative strategies to express BRCA1-wt in UWB1.289 cells and to overcome the instability of the BRCA1-null alleles in OV90 cells are discussed as future approaches to test the direct causal relationship between BRCA1-loss and FEN1-dependency in ovarian cancer.