Liquid biopsy and its role in an advanced clinical trial for lung cancer

Abstract

Liquid biopsy methodologies, for the purpose of plasma genotyping of cell-free DNA (cfDNA) of solid tumors, are a new class of novel molecular assays. Such assays are rapidly entering the clinical sphere of research-based monitoring in translational oncology, especially for thoracic malignancies. Potential applications for these blood-based cfDNA assays include: (i) initial diagnosis, (ii) response to therapy and follow-up, (iii) tumor evolution, and (iv) minimal residual disease evaluation. Precision medicine will benefit from cutting-edge molecular diagnostics, especially regarding treatment decisions in the adjuvant setting, where avoiding over-treatment and unnecessary toxicity are paramount. The use of innovative genetic analysis techniques on individual patient tumor samples is being pursued in several advanced clinical trials. Rather than using a categorical treatment plan, the next critical step of therapeutic decision making is providing the "right" cancer therapy for an individual patient, including correct dose and timeframe based on the molecular analysis of the tumor in question. Per the 21st Century Cures Act, innovative clinical trials are integral for biomarker and drug development. This will include advanced clinical trials utilizing: (i) innovative assays, (ii) molecular profiling with cutting-edge bioinformatics, and (iii) clinically relevant animal or tissue models. In this paper, a mini-review addresses state-of-the-art liquid biopsy approaches. Additionally, an on-going advanced clinical trial for lung cancer with novelty through synergizing liquid biopsies, co-clinical trials, and advanced bioinformatics is also presented. Impact statement Liquid biopsy technology is providing a new source for cancer biomarkers, and adds new dimensions in advanced clinical trials. Utilizing a non-invasive routine blood draw, the liquid biopsy provides abilities to address perplexing issues of tumor tissue heterogeneity by identifying mutations in both primary and metastatic lesions. Regarding the assessment of response to cancer therapy, the liquid biopsy is not ready to replace medical imaging, but adds critical new information; for instance, through a temporal assessment of quantitative circulating tumor DNA (ctDNA) assay results, and importantly, the ability to monitor for signs of resistance, via emerging clones. Adjuvant therapy may soon be considered based on a quantitative cfDNA assay. As sensitivity and specificity of the technology continue to progress, cancer screening and prevention will improve and save countless lives by finding the cancer early, so that a routine surgery may be all that is required for a definitive cure.

Keywords: Bioinformatics; biomarkers; cancer; genomics; liquid biopsy; precision medicine.

Figure 1.

Cell-free DNA (cfDNA) origins. cfDNA is released by normal cells and cells involved with pathologic processes (e.g. inflammation, neoplasia). Circulating tumor DNA (ctDNA) is a subset of cfDNA released by tumor cells, and may contain a variety of genomic alterations (e.g. point mutations, chromosomal rearrangements, copy number variations). The vast majority of ctDNA is felt not to be derived from circulating tumor cells (CTCs). Although scientifically interesting, circulating RNA and exosomes are not quite ready for the clinic. (A color version of this figure is available in the online journal.)

Figure 2.

Consensus analysis of reads by molecular tagging. (a) Reads that align to the same region of the genome and contain the same molecular tag are grouped, and analyzed for true positive variants and false positives. Error correction is applied by eliminating false positives. Only true positive variants will appear in all of the raw aligned reads with the common molecular tag. (b) High-level consensus analysis pipeline. (A color version of this figure is available in the online journal.)

Figure 3.

Clinical trial design and information flow. (a) The UAMS Clinical Trials Office coordinates all aspects of the approved Institutional Review Board (IRB) protocol, as well as patient enrollment, de-identification of all patient information and samples, and performs protocol monitoring. (b) The cohorts and sample types included in the clinical trial are: (i) lung cancer patients, all stages, and histologies, (ii) inflammatory disease patients (Rheumatoid Arthritis and Hepatitis C), (iii) heavy smokers (≥ 30 pack years) without cancer per low dose CT (LDCT) testing and, (iv) a normal volunteer exercise group. Regarding liquid biopsy (LBx), both plasma and urine are collected from all cohorts for the study of cfDNA in both health and different disease states. (c) UAMS genomics shared resource. (d) The Google Cloud is utilized to enhance collaborative endeavors across the member organizations of the Arkansas Bioinformatics Consortium (AR-BIC), which is an alliance consisting of the research universities in Arkansas and the National Center for Toxicological Research (NCTR), a center of the Food and Drug Administration (FDA). (A color version of this figure is available in the online journal.)

Figure 4.

Towards patient-based prognostic assays. (a) Tumor material is acquired from patients with early stage disease during surgical resection for cure or a research biopsy is obtained from patients with more advanced disease. Tumor tissue is used for both patient-derived xenograft (PDX) and tumor explant studies. (b) Tumor tissue is transplanted into NOD scid gamma (NSG) mice to initiate the PDX. If molecular profiling reveals a hotspot mutation (e.g. EGFR L858R) in the patient’s tumor, a genetically engineered mouse (GEM) will be purchased from a commercial vendor or developed in-house at UAMS. (c) A custom database captures all clinical and molecular profiling experimental results from patients and their corresponding mouse models. (d) Temporal analyses are performed on patients and mouse models via liquid biopsies, e.g. for investigation of therapeutic response or resistance. (A color version of this figure is available in the online journal.)