Nuclear cGAS mediated replication stress and mitotic catastrophe can overcome gemcitabine resistance.

Gemcitabine, a ribonucleotide reductase (RNR) inhibitor, is active in pancreatic ductal carcinoma (PDAC) patients, but unfortunately has a limited impact on long term outcomes. Gemcitabine induces nucleotide deficiency, DNA damage including single stranded DNA (ssDNA) and replication stress (RS). DNA damage can activate cyclic GMP-AMP synthase (cGAS), leading to genome instability, micronucleus generation, and immune activation. In model systems, gemcitabine resistance can be overcome by combination treatment with the ataxia telangiectasia and Rad3-related inhibitor (ATRi; AZD6738) that blocks S and G2 checkpoints, although underlying mechanisms remain to be fully elucidated. We show that cells with low basal RS are resistant to gemcitabine, which could be overcome by combination treatment with AZD6738 through elevation of RS, phospho-RPA32 exhaustion, and mitotic catastrophe in PDAC cell models. Gemcitabine induces nuclear cGAS accumulation independent of STING-mediated immune activation. The binding of nuclear cGAS to γH2AX at double strand DNA breaks (DSBs) plays a pivotal role in RS activation and mitotic catastrophe in gemcitabine and AZD6738 treated cells.