Detecting cancer biomarkers in blood: challenges for new molecular diagnostic and point-of-care tests using cell-free nucleic acids

Abstract

As we move into the era of individualized cancer treatment, the need for more sophisticated cancer diagnostics has emerged. Cell-free (cf) nucleic acids (cf-DNA, cf-RNA) and other cellular nanoparticulates are now considered important and selective biomarkers. There is great hope that blood-borne cf-nucleic acids can be used for ‘liquid biopsies’, replacing more invasive tissue biopsies to analyze cancer mutations and monitor therapy. Conventional techniques for cf-nucleic acid biomarker isolation from blood are generally time-consuming, complicated and expensive. They require relatively large blood samples, which must be processed to serum or plasma before isolation of biomarkers can proceed. Such cumbersome sample preparation also limits the widespread use of powerful, downstream genomic analyses, including PCR and DNA sequencing. These limitations also preclude rapid, point-of-care diagnostic applications. Thus, new technologies that allow rapid isolation of biomarkers directly from blood will permit seamless sample-to-answer solutions that enable next-generation point-of-care molecular diagnostics.

Keywords:

• biomarkers
• cancer
• cell-free DNA
• cell-free RNA
• exosomes
• liquid biopsy
• molecular diagnostics
• nanoparticulates
• point-of-care
• sample preparation

Financial & competing interests disclosure

M Heller serves on the Scientific Advisory Board of Biological Dynamics, who produces a dielectrophoretic microarray device for the isolation of DNA. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

Key issues

• The analysis of blood-borne cell-free nucleic acids (cf-NA), demonstrated as a biomarker for multiple diseases, has tremendous potential to enable rapid, non-invasive molecular diagnostics for cancer.

• Recent advances in techniques to amplify and sequence nucleic acids (digital PCR and next-generation sequencing) now permit identification of the genomic signature of a tumor from minute concentrations of cf-NA contained within a blood sample.

• The ability to rapidly screen the blood for tumor-shed, aberrant nucleic acids will facilitate the move towards personalized medicine, starting with initial diagnosis, then providing critical feedback during treatment, and continuing through follow-up.

• Commonly used methods for isolating cf-NA from blood are relatively time-consuming and labor-intensive, which precludes their use in point-of-care (POC) diagnostic tests. This and a lack of isolation protocol standards ultimately impede development of rapid, POC tests based on cf-NA.

• New technologies will be required to resolve limitations of the multi-step processing of blood samples that is required by current protocols before the steps for nucleic acid amplification and analysis can begin.

• Dielectrophoresis technology has the potential to circumvent multi-step processing of blood samples, combining the sample preparation process with downstream analysis (PCR, sequencing, etc.), thus enabling seamless sample-to-answer systems that will lead to widespread use of liquid biopsies and viable POC molecular diagnostics.

Notes