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Nguyen, Truc

University of British Columbia

Telomerase is a specialized RNA-dependent DNA polymerase that maintains the length of telomeres at chromosomal ends by synthesizing telomeric repeats de novo from an internal RNA template. Without telomerase activity, telomeres shorten with each cell division, eventually reaching a length that would prompt either senescence or apoptosis. Telomerase reverse transcriptase (TERT), the catalytic component of telomerase, is upregulated in the majority of human cancers, often at a level that is more than sufficient to keep telomere length above the critical threshold. Previously, our laboratory reported that recombinant TERT expression protects transformed fibroblasts that utilize an alternative telomerase-free telomere lengthening mechanism against drug-induced DNA damage, indicating that TERT has a role in DNA damage response which is independent from its canonical telomeric function. In the present study, we treated recombinant TERT-expressing ovarian cancer cells with genotoxic agents of distinct mechanisms of action to identify possible DNA damage response pathways that TERT participates in. Specifically, our cell model is the recombinant TERT-expressing PEO1 ovarian cancer cells (PEO1+TERT) which have innately impaired capacity for the error-free DNA repair pathway homologous recombination (HR). Genotoxic drugs tested include CX5461 (a G-quadruplex stabilizer currently in clinical trials for HR-deficient cancers) and the following FDA-approved chemotherapeutic agents: bleocin (direct double-strand break inducer); etoposide (topoisomerase II inhibitor); niraparib and olaparib (poly-ADP ribose polymerase (PARP) inhibitors); cisplatin and oxaliplatin (platinum compounds); and mitomycin C (non-platinum crosslinking agent). Additionally, we performed immunofluorescence to detect γH2AX foci – a DNA damage biomarker – in drug-treated PEO1+TERT in comparison to vector-control cells. We found that recombinant TERT confers significant protection against bleocin, CX5461, etoposide, and mitomycin C, and insignificant protection against platinum compounds, on HR-deficient PEO1, and that this protective effect of TERT is not dependent on PARP activity. Additionally, we observed that PEO1+TERT escapes cell cycle arrest and proliferates better than vector-control cells subjected to the same treatment despite sustaining higher level of γH2AX foci formation. Thus, we conclude that TERT promotes DNA damage tolerance and/or error-prone repair in HR-deficient PEO1 cells, and as such is a potential chemotherapy-sensitizing target for telomerase-positive HR-deficient cancers.

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2024-10-31

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/82907

Pharmaceutical Sciences

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/