Ewa Theressa, Panchwagh Neel, Tai Chin-Hsien, Avula Leela Rani, Joseph Sarah, Rudloff Michael W, Malik Nargis, Zhang Xianyu, Alewine Christine
Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.
FASEB J. 2024 Dec 13;38(24):e70247. doi: 10.1096/fj.202400446R.
Peritoneum is the second most common site of metastasis in patients with pancreatic ductal adenocarcinoma (PDAC). Peritoneal colonization is impaired in PDAC cells with knockout (KO) of the cancer surface antigen mesothelin (MSLN) or by introducing Y318A mutation in MSLN to prevent binding to mucin-16 (MUC-16). MSLN has a membrane-bound form but is also shed to release soluble MSLN (sMSLN). Their individual roles in peritoneal metastasis are unknown. Here, a C-terminal truncated MSLN mutant (∆591) incapable of cell membrane insertion but proficient in secretion was engineered. Expression of ∆591 MSLN failed to rescue peritoneal metastasis in MSLN KO cells and inhibited peritoneal colonization when overexpressed in WT PDAC cells. Exposing PDAC cells to conditioned medium (CM) containing excess sMSLN impaired cancer cell clustering in vitro and in peritoneal fluid in vivo, while CM containing only Y318A sMSLN did not. These data demonstrate that interaction of membrane-bound MSLN with MUC-16 promotes cell clustering that is critical for efficient peritoneal metastasis. However, peritoneal colonization by MSLN KO cells was rescued by expression of ∆591 mutant MSLN bearing Y318A mutation, suggesting that sMSLN also has a MUC-16-independent role in peritoneal spread. Alterations in inflammatory signaling pathways occurred following KO cell exposure to CM containing sMSLN, and CM from cancer cells with intact peritoneal metastasis provoked increased KO cell secretion of IL-1α. While excess sMSLN inhibits cell clustering and peritoneal colonization, sMSLN may also promote PDAC peritoneal metastasis independent of MUC-16.
腹膜是胰腺导管腺癌（PDAC）患者中第二常见的转移部位。在敲除癌症表面抗原间皮素（MSLN）的PDAC细胞中，或通过在MSLN中引入Y318A突变以阻止其与粘蛋白-16（MUC-16）结合，腹膜定植会受到损害。MSLN有膜结合形式，但也会脱落以释放可溶性MSLN（sMSLN）。它们在腹膜转移中的各自作用尚不清楚。在此，构建了一种C端截短的MSLN突变体（∆591），其不能插入细胞膜但分泌功能正常。∆591 MSLN的表达未能挽救MSLN敲除细胞中的腹膜转移，并且在野生型PDAC细胞中过表达时会抑制腹膜定植。将PDAC细胞暴露于含有过量sMSLN的条件培养基（CM）中，会损害体外癌细胞聚集以及体内腹膜液中的癌细胞聚集，而仅含有Y318A sMSLN的CM则不会。这些数据表明，膜结合的MSLN与MUC-16的相互作用促进了对有效腹膜转移至关重要的细胞聚集。然而，携带Y318A突变的∆591突变体MSLN的表达挽救了MSLN敲除细胞的腹膜定植，这表明sMSLN在腹膜扩散中也具有不依赖于MUC-16的作用。在敲除细胞暴露于含有sMSLN的CM后，炎症信号通路发生改变，并且来自具有完整腹膜转移的癌细胞的CM会促使敲除细胞分泌更多的IL-1α。虽然过量的sMSLN会抑制细胞聚集和腹膜定植，但sMSLN也可能独立于MUC-16促进PDAC的腹膜转移。
