Single cell RNA sequencing and TCR repertoire analysis of Adult T-cell Leukemia/Lymphoma (ATLL)

We examined ATLL using archived fresh frozen tissue after the biopsy by single-cell RNA sequencing (scRNA-seq) with T-cell receptor (TCR) clonal analysis. Highly clonal tumor cells showed multiple activating pathways, suggesting dynamic evolution of the malignancy. Dissecting diverse cell types comprising the TME led to the identification of a novel subset of cancer-associated fibroblast which showed enriched epidermal growth factor receptor (EGFR)-related transcripts. Human skin samples were obtained using remnants of biopsy tissue taken for diagnostic purposes.Specimens were placed in phosphate-buffered saline (PBS) on ice. Biopsy samples were cryopreserved in optimal cutting temperature (OCT) compound and stored at -80ºC. Single cell dissociates were loaded into the Chromium system (10x Genomics, USA) targeting 25,000 cells. The Chromium Single Cell 5’ Kit (10x Genomics, USA) was used to generate scRNA-seq and TCR libraries, according to the manufacturer’s instructions. scRNA-seq and TCR-seq data were aligned to the human reference genome (GRCh38) and processed using the CellRanger 4.0.0 pipeline (10x Genomics). In this study, we observed that the clonally expanded malignant tumor cells in ATLL are CD4 T-cells through scRNA-seq combined with TCR clonal analysis.