Gemcitabine is a potent inhibitor of DNA replication and is a mainstay therapeutic for diverse cancers, particularly pancreatic ductal adenocarcinoma (PDAC). However, most tumors remain refractory to gemcitabine therapies. Here, to define the cancer cell response to gemcitabine, we performed genome-scale CRISPR–Cas9 chemical–genetic screens in PDAC cells and found selective loss of cell fitness upon disruption of the cytidine deaminases APOBEC3C and APOBEC3D. Following gemcitabine treatment, APOBEC3C and APOBEC3D promote DNA replication stress resistance and cell survival by deaminating cytidines in the nuclear genome to ensure DNA replication fork restart and repair in PDAC cells. We provide evidence that the chemical–genetic interaction between APOBEC3C or APOBEC3D and gemcitabine is absent in nontransformed cells but is recapitulated across different PDAC cell lines, in PDAC organoids and in PDAC xenografts. Thus, we uncover roles for APOBEC3C and APOBEC3D in DNA replication stress resistance and offer plausible targets for improving gemcitabine-based therapies for PDAC.

吉西他滨是一种有效的 DNA 复制抑制剂，是多种癌症的主要治疗药物，特别是胰腺导管腺癌 (PDAC)。然而，大多数肿瘤仍然对吉西他滨治疗无效。在这里，为了确定癌细胞对吉西他滨的反应，我们在 PDAC 细胞中进行了基因组规模的 CRISPR–Cas9 化学遗传筛选，发现胞苷脱氨酶APOBEC3C和APOBEC3D破坏后细胞适应性选择性丧失。吉西他滨治疗后，APOBEC3C 和 APOBEC3D 通过核基因组中的胞苷脱氨来促进 DNA 复制应激抵抗和细胞存活，以确保 PDAC 细胞中 DNA 复制叉的重新启动和修复。我们提供的证据表明，APOBEC3C或APOBEC3D与吉西他滨之间的化学-遗传相互作用在非转化细胞中不存在，但在不同的 PDAC 细胞系、PDAC 类器官和 PDAC 异种移植物中重现。因此，我们揭示了 APOBEC3C 和 APOBEC3D 在 DNA 复制应激抵抗中的作用，并为改善基于吉西他滨的 PDAC 疗法提供了合理的目标。