Drivers of Precision Medicine: Liquid Biopsy and Next-Generation Sequencing
James A Posey1 and Chen-Hsiung Yeh2*
1Sidney Kimmel Cancer Center, Thomas Jefferson University, USA
2Circulogene Theranostics, USA
Submission: December 11, 2016; Published: December 15, 2016
*Corresponding author: Chen-Hsiung Yeh, Circulogene Theranostics, 3125 Independence Drive, Suite 301, Birmingham, AL USA 35209; Tel: 1-205-234-0128
How to cite this article: James A. P, Chen-Hsiung Y. Drivers of Precision Medicine: Liquid Biopsy and Next-Generation Sequencing. Canc Therapy & Oncol Int J. 2016; 2(4): 555595. DOI: 10.19080/CTOIJ.2016.02.555595
Targeted therapy specifically aims at tumor genetic alterations s the hallmark of precision medicine. Companion diagnostic testing utilized to determine the presence or absence of certain oncogenic mutations prior to targeted treatment under the current medical guidelines will enable improved clinical outcome, and thus serves as a vital component for precision medicine. Standard clinical practice to assess genetic mutations in cancer patients has historically been through direct sampling of tumor tissue with biopsy or surgical resection. Unfortunately, tissue biopsy is one-time single-site sampling, painful, costly, and can miss the dynamics of tumor clonally evolution during disease progression. The emerging liquid biopsy by circulating cell-free DNA (cfDNA), on the other hand, can fulfill the gap, not only providing actionable information for decision-making prior to treatment, but also longitudinal surveillance during and after the treatment regimen to assess for patient response, drug resistance[1-3] and cancer recurrence .
Genomic content of cfDNA that is collected from a befouled specimen must be tested with an appropriate analytical method in order to identify existing mutant alleles. Traditional methods for mutation detection include assays such as quantitative PCR and Sanger sequencing [5,6]. PCR-based assays are sensitive but provide only limited information on a handful of genes and hotspot mutations, whereas Sanger sequencing does not possess the sensitivity adequate for detection in the context of liquid biopsy . Modern next-generation sequencing (NGS)-based technology involves sequencing DNA in a highly multiplex and parallel fashion with higher sensitivity .
Whole genome sequencing (WGS) or exome sequencing (WES) of tumor profiling using NGS is an unbiased approach that provides extensive and comprehensive genomic information about cancer. Although they can provide genetic resolution both at the single nucleotide level, as well as detect structural alterations such as large rearrangements, gross deletions and duplications [9-11],the cost of these sequencing are still too high and our current knowledge bases are too small for routine clinical application. Targeted gene panels are currently the best option for CLIA clinical utility in the field of liquid biopsy, as they allow multiple action able genes to be analyzed and can provide enough depth of coverage to detect minor allele frequencies in a clinically-relevant and cost-effective manner [12,13]. NGS gene panels can also eliminate the need for reflex testing and preserves precious specimens by limiting the number of tests required for full characterization. In order for NGS liquid biopsy diagnostics to be accurate and affordable, a balance must be reached between panel size and the level of multiplexing.
There are currently a handful of CLIA liquid biopsy companies who offer cfDNA NGS-based genetic testing. As more NGS gene panels begin to enter the market, it is important for users to understand the benefits and limitations of these methods and technologies being utilized. There are many nuances in designing NGS gene panels for liquid biopsy tumor profiling, and multiple options exist for each component of the workflow. Numerous factors can impact the accuracy, sensitivity and specificity of cfDNA sequencing including the sample preparation, target enrichment, library construction, sequencing platform and bioinformatics tool. Some degree of standardization of cfDNA
NGS assays used for the detection of somatic mutations is
essential before their deployment in clinical practice. Towards
this direction, in addition to the concordance studies between
tissue and liquid biopsies, comparison of cfDNA-based testing
between different methodologies on different platforms in
different laboratories are needed.
In the first side-by-side comparison study, the NGS results for
a12-patient cohort from 2 commercially available gene panels
(70 genes and 50 genes) are analyzed. These samples were
processed and tested in two independent CLIA liquid biopsy
laboratories, Guardant Health (GH) and Circulogene Theranostics
(CT), with distinct sample volume input, sample preparation,
library construction, NGS platform and different mutation calling
bioinformatics. GH and CT tests resulted in a very similar 40-
50% rate of clinically relevant actionable information, i.e., either
FDA drugs or active clinical trials. There are 2 cases reported
“no mutation” by both tests. TP53 is the most frequent mutated
gene identified in 41.7% (5/12) of cases. In 5 cases, there is at
least one gene mutation matched. In another 5 cases, mutations
did not match each other. It is noteworthy that there are 43
genes commonly covered by both panels, however, Guardant’s
test surveyed all axons of these 43 genes, while Circulogene’s
assay only scanned hotspot mutations within these genes.
Nevertheless, the overall concordance between both test results
was fairly high at the gene level, because the denominator in this
calculation was 516 (43 genes for 12 patients), and the majority
of genes were called wild-type alleles (no mutation detected).
A key issue in clinical oncology practice is the ability to
accurately interpret what is truly clinically meaningful and
actionable on a report from a commercially available NGS
test. The majority of cancer care is not delivered at tertiary
comprehensive cancer centers, but rather in busy community
oncology practices. Treating physicians will have time
constraints to read and digest NGS reports, so there is a need
for clear, concise reporting of clinically relevant targets. With the integration of additional levels of content enhancement
and scientific/clinical evidence, liquid biopsy NGS reports can
eventually deliver patient-centered context and substance that
will lead to better healthcare outcomes and toward the goal of
precision medicine (Figure 1).