Multi-Faceted Attributes of Salivary Cell-free DNA as Liquid Biopsy Biomarkers for Gastric Cancer Detection

Recent advances in circulating cell free DNA (cfDNA) analysis from biofluids has opened new avenues for liquid biopsy (LB). However, current cfDNA LB assays are limited by the availability of existing information on established genotypes associated with tumor tissues. Certain cancers present with a limited list of established mutated cfDNA biomarkers and thus, non-mutated cfDNA characteristics along with alternative biofluids is needed to broaden the available cfDNA targets for cancer detection. In this report, we have employed a low-coverage single stranded (ss) library NGS pipeline "Broad-Range cell-free DNA-Seq" (BRcfDNA-Seq) using saliva to comprehensively investigate the characteristics of salivary cfDNA (ScfDNA). Identification of cfDNA features has been made possible by applying novel cfDNA processing techniques which permits the incorporation of ultra-short, ss, and jagged DNA fragments. As a proof of concept using 10 gastric cancer (GC) and 10 non-cancer samples, we have examined if ScfDNA characteristics including fragmentomics, end motif profiles, microbial contribution, and human chromosomal mapping could differentiate between these two groups. Individual and integrative analysis of these ScfDNA features demonstrated significant differences between the two cohorts, suggesting that disease state may affect ScfDNA population by altering nuclear cleavage or the profile of contributory organism cfDNA to total ScfDNA. These novel features of ScfDNA characteristics could be clinically useful for improving LB-based detection and the eventual monitoring of local or systemic diseases.