Single-cell RNA-sequencing as a potential approach for studying intratumor heterogeneity in pleural mesothelioma.

Unlabelled: Pleural mesothelioma (PM) is a rare and lethal cancer with limited treatment options. Intratumor heterogeneity (ITH) has been postulated as one of the reasons for the poor treatment response observed in most PM patients. In this regard, we aimed to characterize ITH in a multi-site tumor specimen using single-cell RNA-sequencing (scRNA-seq).

Methods: Tumor cells from three distant biopsies (costal, diaphragmatic, and mediastinal) of an epithelioid PM were analyzed with scRNA-seq.

Results: Three main cell states were identified in all regions: C1, stem-like; C2, epithelial-like; and C3, mesenchymal-like. C1 state was the most prominent globally, although it was less abundant in the mediastinal biopsy, compared to the other two studied regions. Trajectory analysis was suggestive of an epithelial-mesenchymal plasticity dynamic, including a stem-like intermediate state. Signatures of upregulated genes in each state (SigC1, SigC2, SigC3) were obtained and assessed in a large cohort of PM samples. Patients with tumors enriched in SigC3 were associated with worse survival and with reduced sensitivity to standard of care PM regimens. Additionally, SigC1 appeared to be potentially more sensitive to anti-angiogenic therapies.

Conclusions: This study highlights that scRNA-seq is useful to capture PM cellular and molecular heterogeneity and identifies gene-expression signatures with potential clinical relevance for future treatment tailoring.