Discovery and Development of Novel DNA-PK Inhibitors by Targeting the unique Ku-DNA Interaction

Gavande, N. S.; VanderVere-Carozza, P. S.; Pawelczak, K. S.; Vernon, T. L.; Hanakahi, L. A.; Summerlin, M.; Dynlacht, J. R.; Farmer, A. H.; Sears, C. R.; Al Nasrallah, N.; Garrett, J.; Turchi, J. J.

doi:10.1101/2020.08.24.261875

bioRxiv · 10.1101/2020.08.24.261875

Discovery and Development of Novel DNA-PK Inhibitors by Targeting the unique Ku-DNA Interaction

Gavande, N. S.·VanderVere-Carozza, P. S.·Pawelczak, K. S.·Vernon, T. L.·Hanakahi, L. A.·Summerlin, M.·Dynlacht, J. R.·Farmer, A. H.·Sears, C. R.·Al Nasrallah, N.·Garrett, J.·Turchi, J. J.

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Abstract

DNA-dependent protein kinase (DNA-PK) plays a critical role in the non-homologous end joining (NHEJ) repair pathway and the DNA damage response (DDR). DNA-PK has therefore been pursued for the development of anti-cancer therapeutics in combination with ionizing radiation (IR). We report the discovery of a new class of DNA-PK inhibitors that act via a novel mechanism of action, inhibition of the Ku-DNA interaction. We have developed a series of highly potent and specific Ku-DNA binding inhibitors (Ku-DBis) that block the Ku-DNA interaction and inhibit DNA-PK kinase activity. Ku-DBis directly interact with the Ku and inhibit in vitro NHEJ, cellular NHEJ, and potentiate the activity of IR and radiomimetics. Analysis of Ku-null cells demonstrates that Ku-DBis cellular activity is a direct result of Ku inhibition, as Ku-null cells are insensitive to Ku-DBis. The utility of Ku-DBis was also demonstrated in a CRISPR gene-editing model where we demonstrate that the efficiency of gene insertion events was increased in cells pre-treated with Ku-DBis, consistent with inhibition of NHEJ and activation of homologous recombination to facilitate gene insertion. These data demonstrate the discovery and application of new series of compounds that modulate DNA repair pathways via a unique mechanism of action.

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Gavande, N. S., VanderVere-Carozza, P. S., Pawelczak, K. S., Vernon, T. L., Hanakahi, L. A., Summerlin, M., Dynlacht, J. R., Farmer, A. H., Sears, C. R., Al Nasrallah, N., Garrett, J., Turchi, J. J.. 2020-08-24. Discovery and Development of Novel DNA-PK Inhibitors by Targeting the unique Ku-DNA Interaction. https://doi.org/10.1101/2020.08.24.261875

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