Thymidine rescues ATR kinase inhibitor-induced deoxyuridine contamination in genomic DNA, cell death, and interferon-a/f3 expression

ATR kinase is a central regulator of the DNA damage response (DDR) and cell cycle checkpoints. ATR kinase inhibitors (ATRi's) combine with radiation to generate CD8+ T cell-dependent responses in mouse models of cancer. We show that ATRi's induce cyclin-dependent kinase 1 (CDK1)-dependent origin firing across active replicons in CD8+ T cells activated ex vivo while simultaneously decreasing the activity of rate-limiting en-zymes for nucleotide biosynthesis. These pleiotropic effects of ATRi induce deoxyuridine (dU) contamination in genomic DNA, R loops, RNA-DNA polymerase collisions, and interferon-a/f3 (IFN-a/f3). Remarkably, thymi-dine rescues ATRi-induced dU contamination and partially rescues death and IFN-a/f3 expression in prolif-erating CD8+ T cells. Thymidine also partially rescues ATRi-induced cancer cell death. We propose that ATRi-induced dU contamination contributes to dose-limiting leukocytopenia and inflammation in the clinic and CD8+ T cell-dependent anti-tumor responses in mouse models. We conclude that ATR is essential to limit dU contamination in genomic DNA and IFN-a/f3 expression.

Automated summary

ATR kinase plays an important role in the DNA damage response and cell cycle checkpoints. ATR kinase inhibitors (ATRi's) have shown to enhance CD8+ T cell-dependent anti-tumor responses in mice. In this study, it is found that ATRi's induce CDK1-dependent origin firing and decrease nucleotide biosynthesis in activated CD8+ T cells. These effects result in dU contamination, R loops, RNA-DNA polymerase collisions, and interferon-alpha/beta (IFN-alpha/beta). Thymidine partially rescues ATRi-induced dU contamination and cell death in proliferating CD8+ T cells, as well as ATRi-induced cancer cell death. It is suggested that ATRi-induced dU contamination contributes to leukocytopenia and inflammation, and is important for CD8+ T cell-dependent anti-tumor responses.