Targeting ATR Kinase as a Strategy for Canine Lymphoma and Leukaemia Treatment.

Abstract

Ataxia telangiectasia and Rad3-related (ATR) kinase is one of the main regulators of cell response to DNA damage and replication stress. Effectiveness of ATR targeting in human cancers has been confirmed in preclinical studies and ATR inhibitors are currently developed clinically in human oncology. In the presented study, we tested the anticancer efficacy of ATR inhibitor berzosertib in an in vitro model of canine haematopoietic cancers. Using MTT assay and flow cytometry, we assessed the cytotoxicity of berzosertib in four established canine lymphoma and leukaemia cell lines and compared it with its activity against noncancerous canine cells. Further, we estimated the level of apoptosis in berzosertib-treated cells via flow cytometry and assessed H2AX phosphorylation as a marker of DNA damage using western blot technique. In flow-cytometric analysis, we also evaluated potential synergism between berzosertib and chlorambucil and assessed the influence of berzosertib on cell cycle disturbances induced by the drug. The results demonstrated that berzosertib, even without additional DNA damaging agent, can be effective against canine lymphoma and leukaemia cells at concentrations that were harmless for noncancerous cells, although sensitivity of individual cancer cell lines varied greatly. Cell death occurred through caspase-dependent apoptosis via induction of DNA damage. Berzosertib also acted synergistically with chlorambucil, probably by preventing DNA damage repair as a consequence of S-phase arrest abrogation. In conclusion, ATR inhibition may provide a new therapeutic option for the treatment of canine lymphomas and leukaemias, but further studies are required to determine potential biomarkers of their susceptibility.