"One of the main reasons cancer cells are resistant to treatment is that they can inherently repair the DNA damage caused by radiation and chemotherapy," explains Christian Elias Badr, PhD, investigator in the Department of Neurology at MGH and co-senior author of the paper. The study's other co-senior author is Charles Pin-Kuang Lai, PhD, at the Academia Sinica in Taiwan.

Their study appeared this month as an online advance paper in Nucleic Acids Research.

DSB damage repair is key to maintaining genomic integrity and cell viability. It also plays a role in cancer treatment, which often includes chemoradiotherapy (radiation and chemotherapy), which disrupts DSB. A cell may recognize the damage and use its intrinsic DNA damage response (DDR) to reduce DSB-caused cell death. As a result, the cancer cell's own DNA repair mechanisms can promote drug resistance and recurrence in some malignancies. Researchers would like to know more about them.

The BLRR approach builds on earlier work members of the team did on enzymes called luciferases. These produce bioluminescence, making them useful for tracking molecules in cells. BLRR uses secreted Gaussia and Vargula luciferases to detect homology-directed repair (HDR) and non-homologous end joining (NHEJ) -- the two major pathways to DSB repair. Using BLRR. Researchers can track HDR and NHEJ-related activities over time in cells. It also detects DSB repairs in xenografted tumors in vivo.

"You can study DNA damage in cells with next generation sequencing (NGS), but that's more costly and time consuming," Badr says. "And our system's accuracy is comparable to NGS."

The researchers used their new tag to carry out multiple studies. In one, they found a significant difference in the efficiency of CRISPR/Cas9-mediated editing with guide RNAs 1-10bp apart. They also used BLRR analysis to detect altered dynamics for DSB repair induced by small molecule modulators. Lastly, they used the system to discover HDR-suppressing functions of anticancer cardiac glycosides in human glioblastomas and glioma cancer stem-like cells by inhibiting DNA repair protein RAD51 homolog 1.

In their paper, the authors describe the BLRR system as: "A highly sensitive platform to simultaneously and longitudinally track HDR and NHEJ dynamics that is sufficiently versatile for elucidating the physiology and therapeutic development of DSB repair." The authors plan on using this reporter system in high throughput drug screening to identify novel therapeutics that sensitize cancer cells to radiation and chemotherapy.

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Materials provided by Massachusetts General Hospital. Note: Content may be edited for style and length.

Journal Reference:

1. Jasper Che-Yung Chien, Elie Tabet, Kelsey Pinkham, Cintia Carla da Hora, Jason Cheng-Yu Chang, Steven Lin, Christian E Badr, Charles Pin-Kuang Lai. A multiplexed bioluminescent reporter for sensitive and non-invasive tracking of DNA double strand break repair dynamics in vitro and in vivo. Nucleic Acids Research, 2020; DOI: 10.1093/nar/gkaa669