Abstract
Chk1 kinase inhibitors are currently under clinical investigation as potentiators of cytotoxic chemotherapy and demonstrate potent activity in combination with anti-metabolite drugs that increase replication stress through the inhibition of nucleotide or deoxyribonucleotide biosynthesis. Inhibiting other metabolic pathways critical for the supply of building blocks necessary to support DNA replication may lead to increased DNA damage and synergy with an inhibitor of Chk1. A screen of small molecule metabolism modulators identified combinatorial activity between a Chk1 inhibitor and chloroquine or the LDHA/LDHB inhibitor GSK 2837808A. Compounds, such as 2-deoxyglucose or 6-aminonicotinamide, that reduced the fraction of cells undergoing active replication rendered tumour cells more resistant to Chk1 inhibitor-induced DNA damage. Withdrawal of glucose or glutamine induced G1 and G2/M arrest without increasing DNA damage and reduced Chk1 expression and activation through autophosphorylation. This suggests the expression and activation of Chk1 kinase is associated with cells undergoing active DNA replication. Glutamine starvation rendered tumour cells more resistant to Chk1 inhibitor-induced DNA damage and reversal of the glutamine starvation restored the sensitivity of tumour cells to Chk1 inhibitor-induced DNA damage. Chk1 inhibitors may be a potentially useful therapeutic treatment for patients whose tumours contain a high fraction of replicating cells.
Highlights
Maintaining the integrity of and faithfully copying genetic information are critical for cellular health
Inhibiting other metabolic pathways critical for the supply of building blocks necessary to support DNA replication may lead to increased replication stress and synergy with an inhibitor of Chk[1]
We screened a range of compounds capable of modulating cellular metabolism (Table 1) for their potential to increase γH2AX, a marker of DNA damage[19] or pChk[1] (S317), a marker of ATR activation, either alone or in combination with the Chk[1] inhibitor V158411 (Chk1i), in HT29 and U2OS cancer cells
Summary
Maintaining the integrity of and faithfully copying genetic information are critical for cellular health. Inhibition of Chk[1] following genotoxic stress induced by these agents results in checkpoint abrogation, inhibition of DNA repair and induction of cell death in cells with a defective p53 response. This approach is currently being evaluated in a range of Phase I and II clinical trials. A class of drugs termed the antimetabolites have been a component of cancer therapy for decades These drugs, which include pemetrexed, gemcitabine and hydroxyurea, generally work by inhibiting enzymes critical for nucleotide or deoxyribonucleotide biosynthesis decreasing the pool of dNTPs available for DNA synthesis thereby blocking cell proliferation and increasing replication stress. We evaluated the effect of numerous small molecule metabolism modulators to increase replication stress and activate the DNA damage response in combination with a novel Chk[1] inhibitor
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