Inhibition of the checkpoint kinase Chk1 induces DNA damage and cell death in human Leukemia and Lymphoma cells.

BACKGROUND: Chk1 forms a core component of the DNA damage response and small molecule inhibitors are currently being investigated in the clinic as cytotoxic chemotherapy potentiators. Recent evidence suggests that Chk1 inhibitors may demonstrate significant single agent activity in tumors with specific DNA repair defects, a constitutively activated DNA damage response or oncogene induced replicative stress. METHODS: Growth inhibition induced by the small molecule Chk1 inhibitor V158411 was assessed in a panel of human leukemia and lymphoma cell lines and compared to cancer cell lines derived from solid tumors. The effects on cell cycle and DNA damage response markers were further evaluated. RESULTS: Leukemia and lymphoma cell lines were identified as particularly sensitive to the Chk1 inhibitor V158411 (mean GI50 0.17 µM) compared to colon (2.8 µM) or lung (6.9 µM) cancer cell lines. Chk1 inhibition by V158411 in the leukemia and lymphoma cell lines induced DNA fragmentation and cell death that was both caspase dependent and independent, and prevented cells undergoing mitosis. An analysis of in vitro pharmacodynamic markers identified a dose dependent decrease in Chk1 and cyclin B1 protein levels and Cdc2 Thr15 phosphorylation along with a concomitant increase in H2AX phosphorylation at Ser139 following V158411 treatment. CONCLUSIONS: These data support the further evaluation of Chk1 inhibitors in hematopoietic cancers as single agents as well as in combination with standard of care cytotoxic drugs.

Targeting conserved water molecules: design of 4-aryl-5-cyanopyrrolo[2,3-d]pyrimidine Hsp90 inhibitors using fragment-based screening and structure-based optimization.

Inhibitors of the Hsp90 molecular chaperone are showing promise as anti-cancer agents. Here we describe a series of 4-aryl-5-cyanopyrrolo[2,3-d]pyrimidine ATP competitive Hsp90 inhibitors that were identified following structure-driven optimization of purine hits revealed by NMR based screening of a proprietary fragment library. Ligand-Hsp90 X-ray structures combined with molecular modeling led to the rational displacement of a conserved water molecule leading to enhanced affinity for Hsp90 as measured by fluorescence polarization, isothermal titration calorimetry and surface plasmon resonance assays. This displacement was achieved with a nitrile group, presenting an example of efficient gain in binding affinity with minimal increase in molecular weight. Some compounds in this chemical series inhibit the proliferation of human cancer cell lines in vitro and cause depletion of oncogenic Hsp90 client proteins and concomitant elevation of the co-chaperone Hsp70. In addition, one compound was demonstrated to be orally bioavailable in the mouse. This work demonstrates the power of structure-based design for the rapid evolution of potent Hsp90 inhibitors and the importance of considering conserved water molecules in drug design.