Effects of Pre-analytic Variables on Circulating microRNAs
Subcontractor: Roswell Park Cancer Institute & University of Texas — MD Anderson
Principal Investigator: Hua Zhao, Ph.D.
Co-Principal Investigator: Christine Ambrosone, Ph.D.
Dr. Ambrosone, Dr. Zhao and his colleagues, realizing the clinical potential of circulating microRNAs in whole blood as biomarkers for cancer detection, performed biospecimen-focused research to identify possible preanalytic variables that may significantly affect the levels of circulating microRNAs. Their team accomplished these using two milestones, the first being to discover a panel of “housekeeping” circulating microRNAs, which are ubiquitous by utilizing a unique resource of the DataBank and BioRepository (DBBR) at Roswell Park Cancer Institute to conduct a two-step analysis to identify internal control circulating microRNAs in whole blood. They also sought to develop circulating microRNA QC tools by studying the effects of the pre-analytic variables on the “housekeeping” microRNAs and investigated how selected major preanalytic variables (namely, processing delay, storage condition, storage time, and freeze/thaw cycles) may affect the detection of circulating microRNAs. In the discovery phase of the first step, their team identified three microRNAs, including miR346, miR134, and miR934, whose levels exhibited the smallest variation between the case-control groups, as well as within each group inter-individually. In the further validation analysis, the consistency was validated for miR346 and miR134. At the second step, using miR346 and miR134 as internal controls, their team observed that as the increase in numbers of freeze/thaw cycles decreased the levels of both miR346 and miR134 (p< 0.0001); varying other processing and storage conditions did not affect miRNA levels. In a parallel analysis on plasma samples, levels of miR16 was found to be significantly decreased with increase in processing delay and increase in numbers of freeze/thaw cycles but was not affected by storage condition and duration. Their team identified several circulating microRNAs that could be used as internal controls to assess the quality of circulating microRNAs. These studies could have a great impact on assessing miRNA as cancer biomarkers in diagnostics and therapeutics. In addition the results from this study highlight the necessity of biospecimen-focused research on circulating microRNAs before clinical utilization.