Dampening Enthusiasm for Circulating MicroRNA in Breast Cancer

Abstract

Genome-wide platforms for high-throughput profiling of circulating miRNA (oligoarray or miR-Seq) offer enormous promise for agnostic discovery of circulating miRNA biomarkers as a pathway for development in breast cancer detection. By harmonizing data from 15 previous reports, we found widespread inconsistencies across prior studies. Whether this arises from differences in study design, such as sample source or profiling platform, is unclear. As a reproducibility experiment, we generated a genome-wide plasma miRNA dataset using the Illumina oligoarray and compared this to a publically available dataset generated using an identical sample size, substrate and profiling platform. Samples from 20 breast cancer patients, 20 mammography-screened controls, as well as 20 breast cancer patients after surgical resection and 10 female lung or colorectal cancer patients were included. After filtering for miRNAs derived from blood cells, and for low abundance miRNAs (non-detectable in over 10% of samples), a set of 522 plasma miRNAs remained, of which 46 were found to be differentially expressed between breast cancer patients and healthy controls (p<0.05), of which only 3 normalized to baseline levels in post-resection cases and were unique to breast cancer vs. lung or colorectal cancer (miR-708*, miR-92b* and miR-568, none previously reported). We were unable to demonstrate reproducibility by various measures between the two datasets. This finding, along with widespread inconsistencies across prior studies, highlight the need for better understanding of factors influencing circulating miRNA levels as prerequisites to progress in this area of translational research.

Direct Comparison of Metastasis-Related miRNAs Expression Levels in Circulating Tumor Cells, Corresponding Plasma, and Primary Tumors of Breast Cancer Patients

“…Although several groups have been focused on studying the expression levels of circulating miRNAs in plasma, it is remarkable that limited overlap has been observed between the findings of even very similar studies of the same disease. Regarding breast cancer, as Thompson et al mentioned, there is a dampening enthusiasm for circulating miRNAs because after the analysis of 15 reports on circulating miRNAs in breast cancer patients revealed a very low reproducibility between the datasets published previously (45 ). The lack of reproducibility may have several reasons: (a) sample type (plasma, serum, whole blood), (b) differences in blood processing protocols, (c) differences in study populations, (d) differences in time points of sample collections (46 ).…”

Circulating MicroRNA Biomarker Studies: Pitfalls and Potential Solutions

“…There was no overlap of significantly differentially expressed miRNAs between the Leidner et al data set and others. The authors noted that consistently discordant results outnumbered concordant results, justifying what they called "dampening enthusiasm" in their article title (27 ). Unfortunately, the more recent findings have not helped to build consensus.…”

“…In some studies, these potential markers may have been overlooked. Interestingly, Leidner et al discarded common blood miRNAs before analysis, effectively focusing on just these rare miRNAs (27 ). The possibility of reanalysis and discovery is another argument for public availability of raw and normalized data.…”

Acute Hepatitis C Virus Infection Induces Consistent Changes in Circulating MicroRNAs That Are Associated with Nonlytic Hepatocyte Release

“…Little is understood about the early dynamics and release of circulating miRNAs under specific disease conditions. Importantly, circulating miRNA signatures have been challenging to confirm in separate cohorts, raising questions as to whether consistent changes in circulating miRNAs occur (9,10).…”