pf-477736 and Lymphoma

Checkpoint kinase 1 (CHK1) is critical for S-phase fidelity and preventing premature mitotic entry in the presence of DNA damage. Tumor cells have developed a strong dependence on CHK1 for survival, and hence, this kinase has developed into a promising drug target. Chk1 deficiency in mice results in blastocyst death due to G2/M checkpoint failure showing that it is an essential gene and may be difficult to target therapeutically. Here, we show that chemical inhibition of CHK1 kills murine and human hematopoietic stem and progenitor cells (HSPCs) by the induction of BCL2-regulated apoptosis. Cell death in HSPCs is independent of p53 but requires the BH3-only proteins BIM, PUMA, and NOXA. Moreover, Chk1 is essential for definitive hematopoiesis in the embryo. Noteworthy, cell death inhibition in HSPCs cannot restore blood cell formation as HSPCs lacking CHK1 accumulate DNA damage and stop dividing. Moreover, conditional deletion of Chk1 in hematopoietic cells of adult mice selects for blood cells retaining CHK1, suggesting an essential role in maintaining functional hematopoiesis. Our findings establish a previously unrecognized role for CHK1 in establishing and maintaining hematopoiesis.

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; Benzodiazepinones; Bone Marrow Cells; Cell Line, Tumor; Checkpoint Kinase 1; Embryo, Mammalian; Fetus; G2 Phase Cell Cycle Checkpoints; Hematopoiesis; Hematopoietic Stem Cells; Humans; Lymphoma; Melanoma, Experimental; Mice; Mice, Knockout; Primary Cell Culture; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-bcl-2; Pyrazoles; Quinolines; Quinuclidines; Tumor Suppressor Protein p53; Tumor Suppressor Proteins

CHK1 and CHK2 function as effectors of cell cycle checkpoint arrest following DNA damage. Small molecule inhibitors of CHK proteins are under clinical evaluation in combination with chemotherapeutic agents known to induce DNA damage. We examined whether CHK inhibitors could be effective as single agents in malignant cells with inherent DNA damage because of deregulated expression of the oncogene c-Myc. Eμ-myc lymphoma cells showed a dramatic increase in the extent of DNA damage and DNA damage response (DDR) signalling within 1 h of treatment with CHK1 inhibitors followed by caspase-dependent apoptosis and cell death. In p53 wild-type/ARF null Eμ-myc lymphoma cells, apoptotic cell death was preceded by accumulation of DNA damage and the amount of DNA damage correlated with the extent of cell death. This effect was not observed in normal B cells indicating that DNA damage accumulation following CHK inhibition was specific to Eμ-myc lymphoma cells that exhibit inherent DNA damage because of MYC-induced replication stress. Similar results were obtained with another structurally distinct CHK-inhibitor. Eμ-myc p53 null lymphoma cells were more sensitive to a dual CHK1/CHK2 inhibitor than to a CHK1-specific inhibitor. In all cases, the level of DNA damage following treatment was the most consistent indicator of drug sensitivity. Our results suggest that CHK inhibitors would be beneficial therapeutic agents in MYC-driven cancers. We propose that inhibitors of CHK can act in a synthetically lethal manner in cancers with replication stress as a result of these cancers being reliant on CHK proteins for an effective DDR and cell survival.

Topics: Animals; Apoptosis; Benzodiazepinones; Caspases; Cell Line, Tumor; Checkpoint Kinase 1; DNA Damage; Genes, myc; Genes, p53; Humans; Lymphoma; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neoplasm Transplantation; Protein Kinases; Pyrazoles