Circulating MicroRNA Biomarker Studies: Pitfalls and Potential Solutions

Abstract

BACKGROUND Circulating microRNAs have been proposed as disease biomarkers that may aid in risk assessment, diagnosis, prognosis, and monitoring of treatment response. The perceived opportunity has loomed particularly large in neoplastic disease, where alterations in cancer cells are thought to be reflected in the extracellular space as affected cells release upregulated miRNAs or fail to release apparently downregulated species. Despite the promise of miRNA biomarkers, evaluation of the diagnostic specificity and reproducibility of reported markers suggests that realizing this promise remains a work in progress. CONTENTS This review examines issues of diagnostic specificity and reproducibility that have afflicted circulating miRNA studies. Surveying the breast cancer literature as an example, few miRNAs are reported consistently. Furthermore, it is posited that the assumptions underlying models of direct contributions of diseased tissue to biofluid miRNA profiles may not hold. Suggestions for improving diagnostic specificity and reliability are provided. SUMMARY To maximize the likelihood of return on investment as miRNAs continue to be evaluated as specific and clinically useful markers, a focus is needed on miRNAs found in specific carriers, such as extracellular vesicles. Alternative sampling techniques should be developed, and nonblood biofluids should be considered. Careful optimization and standardization of preanalytical and analytical methods is needed to ensure that future results, positive or negative, are reliable.

MicroRNAs: A Puzzling Tool in Cancer Diagnostics and Therapy

“…Is it a biomarker of a particular condition or a simple sign of a general state of the disease? (29). As suggested by Witwer the answer to this question depends on what we contemplate as cell-free miRNAs (29).…”

“…(29). As suggested by Witwer the answer to this question depends on what we contemplate as cell-free miRNAs (29). We can assume that the pool of circulating miRNAs contains two types of miRNAs: miRNAs complexed with the Argonaute-2 proteins and vesicle-packed miRNAs (23).…”

“…The effort in finding specific miRNAs for each disease has failed (29). In fact, several miRNAs has been found associated with a broad range of conditions.…”

“…Conversely, miRNAs-Argonaute-2 proteins complexes represent only cell by-products that accumulate in the extracellular fluids (23). This suggests that one way to identify miRNAs suitable as biomarkers should be in focalizing the attention on vesicular miRNAs, instead of continuing the study of the entire circulating pool of molecules (29).…”

Circulating microRNAs and diabetes: potential applications in medical practice

“…These need to be standardised to allow the study of circulating miRNAs. Even though there are recent reports that seem to address this issue, to the best of our knowledge no global consensus exists at this time [72,75,76].…”