Contribution of crosstalk of mesothelial and tumoral epithelial cells in pleural metastasis of lung cancer

Tumor metastasis commonly affects pleura in advanced lung cancer and is related to poor prognosis, but without systematic investigation on different cell types and their crosstalk at single cell resolution. Here, by integrating 180785 single cells from lung cancer and control samples, the most distinctive transcriptome profiles between tumor and control were revealed in mesothelial cells, which is the predominate cell type of pleura. Four subtypes were divided, including one predominately identified in malignant pleural effusion which was characterized by enriched cancer related pathways (e.g., cell migration) along evolutionary trajectory from normal mesothelial cells. Cancer-associated mesothelial cells exhibited varied interactions with different subtypes of malignant epithelial cells, and multiple ligands/receptors exhibited significant correlation with poor prognosis. Experimentally, mesothelial cells can increase the migration ability of lung cancer cells through co-culturing assay, and EGFR was the only affected gene in cancer cells that exhibited interaction with mesothelial cells and was associated with poor prognosis. Using EGFR antagonist cetuximab prevented the increased invasiveness of lung cancer cells induced by mesothelial cells. Moreover, EPGN-EGFR interaction was supported through spatial distribution analysis, revealing the significant proximity between EPGN+ mesothelial cells and EGFR+ epithelial cells. Our findings highlighted the important role of mesothelial cells and their interactions with cancer cells in pleural metastasis of lung cancer, which may facilitate cancer progression. Overall design: After co-culture, the RNA of A549 and MET-5A cells of control group and co-culture group was respectively extracted, which were further sequenced

胸膜转移是晚期肺癌的常见并发症，且与不良预后密切相关，但目前尚缺乏单细胞分辨率下针对不同细胞类型及其相互作用的系统性研究。本研究整合了肺癌样本与对照样本共180785个单细胞，鉴定发现胸膜的主要细胞类型——间皮细胞（mesothelial cells）中，肿瘤组与对照组的转录组特征差异最为显著。该研究将间皮细胞分为4个亚型，其中一个亚型主要见于恶性胸腔积液，其特征为在从正常间皮细胞演化的轨迹中富集了肿瘤相关通路（如细胞迁移）。肿瘤相关间皮细胞与不同亚型的恶性上皮细胞存在多样的相互作用，且多个配体/受体的表达与不良预后显著相关。实验层面，通过共培养实验证实间皮细胞可增强肺癌细胞的迁移能力；而在与间皮细胞存在相互作用且与不良预后相关的癌细胞基因中，表皮生长因子受体（EGFR）是唯一受影响的基因。使用EGFR拮抗剂西妥昔单抗（cetuximab）可阻断间皮细胞诱导的肺癌细胞侵袭能力增强。此外，空间分布分析验证了EPGN-EGFR的相互作用，证实EPGN阳性间皮细胞与EGFR阳性上皮细胞之间存在显著的空间邻近关系。本研究结果揭示了间皮细胞及其与癌细胞的相互作用在肺癌胸膜转移中的重要作用，该作用可能促进肿瘤进展。整体实验设计：分别提取共培养组与对照组的A549细胞及MET-5A细胞的RNA，随后进行测序。