我们和其他人最近表明，在体外，DNA损伤响应（DDR）所涉及的蛋白质对KRAS突变的胰腺导管腺癌（PDAC）细胞生长至关重要。然而，使我们能够识别这些关键蛋白质的CRISPR-Cas9文库的DDR相关基因受到限制。为了进一步研究DDR在这种情况下的作用，我们进行了全面的、DDR-focused的CRISPR-Cas9 loss-of-function筛选。本次筛选确定了总裁含蛋白（VCP）是KRAS突变PDAC细胞系中的必需基因。我们观察到，基因和药物对VCP的抑制限制了细胞生长并诱导了细胞凋亡。针对VCP依赖性生长的基础，我们首先评估了VCP对DDR的贡献，发现VCP的缺失导致DNA双链断裂的积累。接下来，我们研究了它在蛋白质稳态中的作用，发现VCP的缺失导致多泛素化蛋白质的积累。我们还发现，VCP的缺失增加了自噬作用。因此，我们推断，同时抑制VCP和自噬可能是一种有效的组合。因此，我们发现，VCP抑制剂与自噬抑制剂氯喹具有协同作用。我们得出结论，同时靶向自噬可以增强KRAS突变PDAC中VCP抑制剂的疗效。

We and others have recently shown that proteins involved in the DNA damage response (DDR) are critical for KRAS-mutant pancreatic ductal adenocarcinoma (PDAC) cell growth in vitro. However, the CRISPR-Cas9 library that enabled us to identify these key proteins had limited representation of DDR-related genes. To further investigate the DDR in this context, we performed a comprehensive, DDR-focused CRISPR-Cas9 loss-of-function screen. This screen identified valosin-containing protein (VCP) as an essential gene in KRAS-mutant PDAC cell lines. We observed that genetic and pharmacologic inhibition of VCP limited cell growth and induced apoptotic death. Addressing the basis for VCP-dependent growth, we first evaluated the contribution of VCP to the DDR and found that loss of VCP resulted in accumulation of DNA double-strand breaks. We next addressed its role in proteostasis and found that loss of VCP caused accumulation of polyubiquitinated proteins. We also found that loss of VCP increased autophagy. Therefore, we reasoned that inhibiting both VCP and autophagy could be an effective combination. Accordingly, we found that VCP inhibition synergized with the autophagy inhibitor chloroquine. We conclude that concurrent targeting of autophagy can enhance the efficacy of VCP inhibitors in KRAS-mutant PDAC.