The DNA-dependent protein kinase (DNA-PK) plays an instrumental role in the overall survival and proliferation of cells. As a member of the phosphatidylinositol 3-kinase-related kinase (PIKK) family, DNA-PK is best known as a mediator of the cellular response to DNA damage. In this context, DNA-PK has emerged as an intriguing therapeutic target in the treatment of a variety of cancers, especially when used in conjunction with genotoxic chemotherapy or ionizing radiation. Beyond the DNA damage response, DNA-PK activity is necessary for multiple cellular functions, including the regulation of transcription, progression of the cell cycle, and in the maintenance of telomeres. Here, we review what is currently known about DNA-PK regarding its structure and established roles in DNA repair. We also discuss its lesser-known functions, the pharmacotherapies inhibiting its function in DNA repair, and its potential as a therapeutic target in a broader context.

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Amplification‐independent c‐MYC overexpression is suggested in multiple cancers. Targeting c‐MYC activity has therapeutic potential, but efforts thus far have been mostly unsuccessful. To find a druggable target to modulate c‐MYC activity in cancer, we identified two kinases, MAPKAPK2 (MK2) and the DNA‐dependent protein kinase catalytic subunit (DNA‐PKcs), which phosphorylate the Ser111 and the Ser93 residues of OCT4, respectively, to transcriptionally activate c‐MYC. Using these observations, we present here a novel cell‐based luminescence assay to identify compounds that inhibit the interaction between these kinases and OCT4. After screening approximately 80,000 compounds, we identified 56 compounds (“hits”) that inhibited the luminescence reaction between DNA‐PKcs and OCT4, and 65 hits inhibiting the MK2–OCT4 interaction. Using custom antibodies specific for pOCT4S93 and pOCT4S111, the “hits” were validated for their effect on OCT4 phosphorylation and activation. Using a two‐step method for validation, we identified two candidate compounds from the DNA‐PKcs assay and three from the MK2 assay. All five compounds demonstrate a significant ability to kill cancer cells in the nanomolar range. In conclusion, we developed a cell‐based luminescence assay to identify novel inhibitors targeting c‐MYC transcriptional activation, and have found five compounds that may function as lead compounds for further development.