RNA-Seq Blog Transcriptome Research & Industry News
Circulating biomarkers are urgently needed in HCC. miRNA are noncoding RNA that regulate mRNA expression and are detectable in tumor tissue and extracellular compartments. This pilot study explored the feasibility of using next-generation RNA sequencing to characterize miRNA abundance in serum from HCC patients at baseline and on targeted therapy.
Banked serum samples (400µL) were obtained from HCC patients enrolled on a phase I clinical trial of sorafenib plus the mTOR inhibitor, temsirolimus. Control sera were obtained from patients with non-malignant liver diseases (NMLD). Total RNA was purified using Trizol-LS and miRNeasy kit. Mature miRNA were size-selected by gel electrophoresis. Samples were sequenced by Illumina HiSeq 2500, 5M reads per sample. Raw reads were mapped using Novoalign, quantified, and normalized to total reads and exogenous spike-in miRNA recovery. Comparisons between HCC vs. NMLD and baseline vs. on treatment cohorts were performed using one-way ANOVA and false discovery rate (FDR) correction. Candidate signature miRNA were derived by fold change, p-value, and abundance cutoffs and cross-referenced to literature.
Cohorts: HCC baseline (n=23) and paired on treatment (n=20 cases, 30 samples), NMLD (n=12). HCC cohort: HBV+/dual 40%, HCV+ 32%. NMLD cohort: HCV+ 83%. 38 of 65 (58%) total samples qualified. Candidate miRNA were identified which showed up-regulation in HCC vs. NMLD (non-significant) and in elevated vs. normal alpha-fetoprotein (AFP) (FDR p<0.05). Multiple miRNA families showed a trend toward down-regulation on treatment with temsirolimus plus sorafenib including Let-7 and miR-17/92 family members. Serum signatures for HCC vs. NMLD and elevated AFP overlapped with miRNA expression in a published dataset of HCC tumors.
Serum miRNA in HCC patients can be characterized by next-generation sequencing. Differences in miRNA abundance were observed between HCC cases and NMLD controls, and within HCC cases according to clinical covariates including treatment status and AFP. Serum miRNA warrant further study as novel biomarkers in HCC.
