Peak analysis of small cell-free RNA in oral cancer plasma finds recurrently protected narrow regions with clinical potential

Oral cancer is characterized by poor prognosis and low survival rate despite sophisticated surgical and radiotherapeutic modalities. Lymphatic metastasis of oral cancer is a complex process involving multiple post-transcriptional biological processes. Thus, feasible detection of cancer metastasis in plasma could be challenging and beneficial. We recruited patients with oral squamous cell carcinoma (OSCC), one of the common oral cancer types, and checked the lymphatic metastatic status and profiled small RNA in plasma samples. Overall design: Peripheral whole blood samples were collected from individuals before therapy or surgery using EDTA-coated vacutainer tubes. Plasma was separated within 2 hr after collection. All plasma samples were aliquoted and stored at -80°C. CfRNAs were extracted using miRNeasy serum/plasma miRNA isolation kit (QIAGEN, Shanghai, China), and DNA contamination was removed by Recombinant DNase I (RNase-free) (TAKARA, Beijing, China). Small libraries were prepared according to the manual of QIAseq miRNA library kit (QIAGEN, Shanghai, China). Library quantification was performed by Qubit dsDNA HS Kit. Library fragment size and quality were checked using Agilent 2100 Bioanalyzer. Libraries were sequenced on Illumina HiSeq X-ten with PE150.