Simultaneous inhibition of ATM, ATR, and DNA-PK causes synergistic lethality

Here we report that simultaneous inhibition of the three primary DNA damage recognition PI3 kinase-like kinases (PIKKs) —ATM, ATR, and DNA-PK— induces severe combinatorial synthetic lethality in mammalian cells. Utilizing Chinese hamster cell lines CHO and V79 and their respective PIKK mutants, we evaluated effects of inhibiting these three kinases on cell viability, DNA damage response, and chromosomal integrity. Our results demonstrate that while single or dual kinase inhibition increased cytotoxicity, inhibition of all three PIKKs results in significantly higher synergistic lethality, chromosomal aberrations, and DNA double-strand break (DSB) induction as calculated by their synergy scores. These findings suggest that the overlapping redundancy of ATM, ATR, and DNA-PK functions is critical for cell survival, and their combined inhibition greatly disrupts DNA damage signaling and repair processes, leading to cell death. This study provides insights into the potential of multi-targeted DDR kinase inhibition as an effective anticancer strategy, necessitating further research to elucidate underlying mechanisms and therapeutic applications. •Simultaneous inhibition of ATM, ATR, and DNA-PK induces synergistic lethality stronger than dual inhibitions of any of them.•Cellular lethality is associated with chromosome aberration induction rather than SCE.•This study suggests that overlapping redundancy of ATM, ATR, and DNA-PK functions in DNA damage responses.