Tracing the stemness and malignant transition in a heritable colorectal cancer Lynch Syndrome by single-cell RNA-seq analysis.

Background: Lynch Syndrome (LS) is an autosomal dominant disease characterized by germline heterozygous mutations in DNA mismatch repair (MMR) genes. High-risk LS patients may proceed to colorectal cancer (CRC). However, the drivers or biomarkers of LS benign colon tissue approaching malignant CRC are not completely understood. This study aimed to understand the molecular and cellular changes during malignant transition in LS. Methods: Single-cell RNA sequencing (scRNA-seq) was used to analyze paired fresh biopsy samples from 3 LS patients (carcinoma vs. para-carcinoma, labeled as LS-CA vs. LS-paraCA). Single-nuclear RNA sequencing (snRNA-seq) was used to analyze a frozen biopsy sample of a LS patient. Datasets of Healthy controls and patients diagnosed with sporadic CRC (without LS-related germline or somatic mutations; labeled as nonLS-CRC) were downloaded from the open source. Integrative computational analysis was performed to conclude potential drivers of the malignant transition. Immuno-histo-fluorescence staining (IHF) were also performed for validating the proposed three key markers. Results: In the single-cell atlas, we observed an increase of primitive cancer stem-cells with high expression of biomarkers Conclusions: This study provides single-cell transcriptomic resource using affected tissues from patients with Lynch Syndrome and describes an integrative profile covering the alterations of cancer stem cell markers, mutation burden, and tumor immunity during the malignant transition from latency state to Lynch Syndrome and to colorectal cancer at the single-cell level.