Systemic circulating microRNA landscape in Lynch syndrome

Abstract

Circulating microRNAs (c-miRs) are small noncoding RNA molecules that migrate throughout the body and regulate gene expression. Global c-miR expression patterns (c-miRnomes) change with sporadic carcinogenesis and have predictive potential in early detection of cancers. However, there are no studies that have assessed whether c-miRnomes display similar potential in carriers of inherited pathogenic mismatch-repair gene variants (path_MMR), known as Lynch syndrome (LS), who are predisposed to highly increased cancer risk. Using high-throughput sequencing and bioinformatic approaches, we conducted an exploratory analysis to characterize systemic c-miRnomes of path_MMR carriers, sporadic rectal cancer patients and non-LS controls. We showed for the first time that cancer-free path_MMR carriers have a systemic c-miRnome of 40 differentially expressed c-miRs that can distinguish them from non-LS controls. The systemic c-miRnome of cancer-free path_MMR carriers also resembles the systemic c-miRnomes of cancer patients with or without path_MMR. Our pathway analysis linked the found differentially expressed c-miRs to carcinogenesis. A total of 508 putative target genes were identified for 32 out of 40 differentially expressed c-miRs, and 238 of them were enriched in cancer-related pathways. The most enriched c-miR-target genes include well-known oncogenes and tumor suppressor genes such as BCL2, AKT3, PIK3CA, KRAS, NRAS, CDKN1A and PIK3R1. Taken together, our findings suggest that LS and sporadic carcinogenesis share common biological pathways and alterations in these pathways can produce a c-miR signature which can track potential oncogenic stress in cancer-free path_MMR carriers. Therefore, c-miRs hold potential in monitoring the LS risk stratification patterns during clinical surveillance or cancer management.

Keywords: Lynch syndrome; bioinformatics; hereditary cancer; microRNA; next generation sequencing.

FIGURE 1

Healthy path_MMR carriers have a c‐miRnome that differ from non‐LS controls but resembles the c‐miRnomes of patients with sporadic or hereditary cancer. (A) DE c‐miRs in healthy path_MMR carriers vs non‐LS controls. (B) DE c‐miRs in sporadic rectal cancer patients vs path_MMR carriers with cancer. (C) DE c‐miRs in healthy path_MMR carriers vs sporadic rectal cancer patients. (D) DE c‐miRs in healthy path_MMR carriers vs path_MMR carriers with cancer. (E) DE c‐miRs in path_MMR carriers with cancer vs non‐LS controls. (F) DE c‐miRs in sporadic rectal cancers patients vs non‐LS controls. Blue dash lines indicate negative fold change of expression, red dash line indicate positive fold change of expression and gray dash line indicate FDR <0.05. Downregulated c‐miRs are highlighted in red, upregulated c‐miRs are highlighted in cyan and nonsignificantly expressed c‐miRs are highlighted in gray. Dots represents c‐miRs. c‐miR, circulating microRNA; FDR, false discovery rate

FIGURE 2

Dimension reduction analysis of multiple traits was unable to discern path_MMR carriers from sporadic rectal cancer patients. (A) Path_MMR carriers and sporadic rectal cancer patients. (B) Cancer status. Healthy, cancer‐free path_MMR carriers; path_MMR cancer, path_MMR carriers with cancer; path SR cancer, sporadic rectal cancer patients. (C) Cancer history. Current cancer, has cancer currently; Never, currently healthy, never had cancers; Previous cancer, currently healthy, had had cancer or multiple cancers; (D) path_MMR variant. (E) Dichotomous age. Over 60, persons >60‐years of age; Under 60, persons <60‐years of age. (F) Nondichotomous age. Over 60, persons >60‐years of age; Between 50 and 60, persons between 50 and 60‐years of age; Under 50, persons <50‐years of age. (G) BMI. Over 25, persons with BMI > 25; Under 25, persons with BMI < 25; NA, no reported BMI. (H) Sex. M, males; F, females. (I) Batch effect of three separate sequencing runs in running order. All t‐SNE plots are 2D constructions. Dots represent study subjects. BMI, body mass index

FIGURE 3

Pathway analysis revealed putative c‐miR‐target genes that are linked to biological processes and pathways associated with cancer. (A) Top 30 most enriched biological processes annotated to the identified target genes of 32 out of 40 DE c‐miRs found in healthy path_MMR carriers. FDR, false discovery rate; GO:BP, Gene Ontology:biological process; Hits, number of target genes annotated to the biological process. *Signal transduction by p53 class mediator resulting in cell cycle arrest. (B) Top 30 most enriched KEGG pathways annotated to the identified target genes of 32 out of 40 DE c‐miRs found in healthy path_MMR carriers. c‐miR, circulating microRNA; FDR, false discovery rate; KEGG, Kyoto Encyclopedia of Genes and Genomes pathway; Hits, number of target genes annotated to the pathway