Single-cell transcriptome analysis reveals subtype-specific clonal evolution and microenvironmental changes in liver metastasis of pancreatic adenocarcinoma and their clinical implications.

BACKGROUND: Intratumoral heterogeneity (ITH) and tumor microenvironment (TME) of pancreatic ductal adenocarcinoma (PDAC) play important roles in tumor evolution and patient outcomes. However, the precise characterization of diverse cell populations and their crosstalk associated with PDAC progression and metastasis is still challenging. METHODS: We performed single-cell RNA sequencing (scRNA-seq) of treatment-naïve primary PDAC samples with and without paired liver metastasis samples to understand the interplay between ITH and TME in the PDAC evolution and its clinical associations. RESULTS: scRNA-seq analysis revealed that even a small proportion (22%) of basal-like malignant ductal cells could lead to poor chemotherapy response and patient survival and that epithelial-mesenchymal transition programs were largely subtype-specific. The clonal homogeneity significantly increased with more prevalent and pronounced copy number gains of oncogenes, such as KRAS and ETV1, and losses of tumor suppressor genes, such as SMAD2 and MAP2K4, along PDAC progression and metastasis. Moreover, diverse immune cell populations, including naïve SELLhi regulatory T cells (Tregs) and activated TIGIThi Tregs, contributed to shaping immunosuppressive TMEs of PDAC through cellular interactions with malignant ductal cells in PDAC evolution. Importantly, the proportion of basal-like ductal cells negatively correlated with that of immunoreactive cell populations, such as cytotoxic T cells, but positively correlated with that of immunosuppressive cell populations, such as Tregs. CONCLUSION: We uncover that the proportion of basal-like subtype is a key determinant for chemotherapy response and patient outcome, and that PDAC clonally evolves with subtype-specific dosage changes of cancer-associated genes by forming immunosuppressive microenvironments in its progression and metastasis.

Single-cell RNA Sequencing Reveals Novel Cellular Factors for Response to Immunosuppressive Therapy in Aplastic Anemia.

Aplastic anemia (AA) is a lethal hematological disorder; however, its pathogenesis is not fully understood. Although immunosuppressive therapy (IST) is a major treatment option for AA, one-third of patients do not respond to IST and its resistance mechanism remains elusive. To understand AA pathogenesis and IST resistance, we performed single-cell RNA sequencing (scRNA-seq) of bone marrow (BM) from healthy controls and patients with AA at diagnosis. We found that CD34+ early-stage erythroid precursor cells and PROM1+ hematopoietic stem cells were significantly depleted in AA, which suggests that the depletion of CD34+ early-stage erythroid precursor cells and PROM1+ hematopoietic stem cells might be one of the major mechanisms for AA pathogenesis related with BM-cell hypoplasia. More importantly, we observed the significant enrichment of CD8+ T cells and T cell-activating intercellular interactions in IST responders, indicating the association between the expansion and activation of T cells and the positive response of IST in AA. Taken together, our findings represent a valuable resource offering novel insights into the cellular heterogeneity in the BM of AA and reveal potential biomarkers for IST, building the foundation for future precision therapies in AA.

Cellular plasticity and immune microenvironment of malignant pleural effusion are associated with EGFR-TKI resistance in non-small-cell lung carcinoma.

Malignant pleural effusion (MPE) is a complication of lung cancer that can be used as an alternative method for tissue sampling because it is generally simple and minimally invasive. Our study evaluated the diagnostic potential of non-small-cell lung carcinoma (NSCLC)-associated MPE in terms of understanding tumor heterogeneity and identifying response factors for EGFR tyrosine kinase inhibitor (TKI) therapy. We performed a single-cell RNA sequencing analysis of 31,743 cells isolated from the MPEs of 9 patients with NSCLC (5 resistant and 4 sensitive to EGFR TKI) with EGFR mutations. Interestingly, lung epithelial precursor-like cells with upregulated GNB2L1 and CAV1 expression were enriched in the EGFR TKI-resistant group. Moreover, GZMK upregulated transitional effector T cells, and plasmacytoid dendritic cells were significantly enriched in the EGFR TKI-resistant patients. Our results suggest that cellular plasticity and immunosuppressive microenvironment in MPEs are potentially associated with the TKI response of patients with EGFR-mutated NSCLC.