Liquid biopsy next-generation sequencing (NGS) holds great promise in therapy selection for oncology, offering non-invasive and real-time insights into tumor genetics. While challenges remain, ongoing research and technological advancements are likely to significantly enhance the clinical utility and accessibility of liquid biopsy NGS, ultimately contributing to improved patient outcomes in cancer care.
The liquid biopsy landscape
Liquid biopsy testing can be used in the diagnosis of any cancer that secretes nucleic acid into the blood – both solid tumors and hematological cancers.
Saying that, it’s important to remember that solid tumors, in particular, are very heterogenous and shedding rates vary greatly between different tumor types. Additionally, early-stage cancers do not shed to the same extent as late-stage tumors, meaning liquid biopsy, for reasons of sensitivity, is not always as effective in early-stage disease detection.
In the health care setting, clinical testing labs offer tests for guideline-approved biomarkers. Lung cancer has the largest array of actionable mutations, according to current knowledge, and therefore makes up the largest proportion of tests run.
In the clinical trial sector, however, biopharma investigates a much larger array of exploratory biomarkers using liquid biopsy testing. This includes PK– PD studies and other investigations beyond efficacy and therapy selection.
Late-stage disease applications
In late-stage cancer management, the main applications for liquid biopsy are in diagnostics and therapeutic monitoring. For both these applications, liquid biopsy is complementary to traditional tissue testing. Current guidelines require that negative liquid biopsies are always reflexed to a tissue-based assay for confirmation. That is because false negatives are more likely when interrogating circulating tumor DNA than when testing the tumor tissue directly. However, a positive liquid biopsy result can be acted on, and several tests are validated and approved for this use.
Early-stage disease applications
Liquid biopsy offers enormous potential in assessing molecular minimal residual disease, in both solid and liquid cancers, via analysis of circulating free DNA. There are several technologies already available in this space that may find utility in monitoring in the early-stage. Whilst initially focused on metastatic disease, these technologies are now being developed for the early-stage disease setting.
The challenge with the technologies in use today in late-stage is that sensitivity really needs to be optimized for assays in this space. Currently, a negative liquid biopsy in an early-stage cancer patient does not mean there is no cancer; it means more investigation is required as the tumor DNA may not be in circulation or at very low levels. It’s also important to acknowledge that a false positive result can be devastating for the patient at the end of it.
The answer might be to develop liquid biopsies with a tumor-specificity in mind, for which we need more clinical evidence as well as technology research investments, and a reassessment of specimen collection and nucleic acid recovery methods in order to boost levels of ctDNA available for analysis.
Additionally, the reimbursement system also needs to catch up with the technology. If labs are not reimbursed to carry out early-stage disease testing, patients are going to miss out on that opportunity.
Advantages, limitations, and challenges
Tissue sections are a very scant resource, from which establishing the diagnosis is the first priority for pathologists, followed by molecular testing to establish genomic subtyping of a tumor. This is where the liquid biopsy is valuable – as often there is not enough tissue in a sample to support secondary testing. Overall, the benefit of the complementary tissue–liquid testing model is that of getting the patient on therapy sooner than with tissue biopsies alone, which, as oncology studies have proven, improves outcomes.
If this is the case, why don’t we just adopt a model of concurrent liquid and tissue biopsies for every patient? Well, the testing technologies and practice behaviors are not yet well aligned enough to support this, with tissue testing being largely based on immunohistochemistry, and liquid biopsies on molecular sequencing. Combining the techniques gives rise to a higher likelihood of confounding results than with single testing methods, which would introduce inefficiencies in the diagnostic process via retesting.
Liquid biopsy tests are relatively new to clinical diagnostics, and there is still a way to go to iron out the challenges. The way to address discordance between tissue and liquid test results for example, is through a good understanding of the cancer type being tested, as well as the technology types being used. With trust in the validation process of the assay, it becomes an investigative process to resolve the issue.
Establishing reliability and reproducibility of a liquid biopsy assay begins with establishing its goals. The next step is to set up verification and validation studies based on approved guidelines – of which plenty are now available from our learned organizations.
Then there are cost-effectiveness considerations. In the US, the reimbursement codes we use for our genomic tests are subject to regular revision, meaning the unit cost of tests is beyond our control. Pathologists, then, need to focus on the cost-effectives elements that are controllable, such as optimizing batch sizes or negotiating regional bulk discounts with technology suppliers.
Liquid biopsy techniques: rapid NGS versus digital PCR
Both technologies have their place in the liquid biopsy testing space.
Polymerase chain reaction (PCR)-based methods, being less expensive than NGS, are therefore far more accessible to many labs, but they can only provide limited variant information. Because of the lower cost and ease of use of PCR, it is most likely to be used for disease monitoring.
While NGS has relatively higher cost and complexity, it provides comprehensive biomarker information across hundreds of genes and is more likely to be used for baseline and landmark molecular tumor assessments and therapy guidance.
My experience with NGS technologies is in amplicon-based systems, which offer a comparable sensitivity to PCR testing. It also provides faster turnaround times than other NGS technologies due to its relatively low level of technical complexity in the test instrumentation – leading to a simplified workflow while retaining accurate and reliable results.
Hybrid capture technologies tend to have larger panels for more comprehensive screening, providing more molecular content than other technologies. In terms of informing treatment decisions, however, bigger isn’t always better. As more hybrid capture-based larger panels are developed, it will be interesting to see the comparisons against smaller panels in various applications.
Future directions of liquid biopsy NGS testing technology
I anticipate that NGS technologies in this area will be developed on a cancer-by-cancer basis. In a recently published study, our research team collected real-world liquid biopsy data across multiple cancers. We found that it is still only really the “big four” cancers for which we have significant data. We must invest more on research for monitoring of all cancer types.
I also predict we will see more standardization of the technologies, the reporting, and the therapeutic choices. For early-stage cancers, we need to see a revolution in pre-analytics, reimbursement models, technologies, and bioinformatics in order to address the large unmet need in this space.
Currently, testing and treatment of metastatic cancers is working well. However, if we can drive down healthcare costs by reimbursing liquid biopsy testing in the earlier stages of cancer diagnostics, patient outcomes and quality of life will undoubtedly improve.
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References
- BA McKelvey et al., “Advancing evidence generation for circulating tumor DNA: lessons learned from a multi-assay study of baseline circulating tumor DNA levels across cancer types and stages,” Diagnostics (Basel), 14, 9 (2024). PMID: 38732326.
- MF Mosele et al., “Recommendations for the use of next-generation sequencing (NGS) for patients with advanced cancer in 2024: a report from the ESMO Precision Medicine Working Group,” Ann Oncol, 35, 7 (2024). PMID: 38834388.
- C Rolfo et al., “Liquid biopsy for advanced NSCLC: a consensus statement from the International Association for the Study of Lung Cancer,” J Thorac Oncol, 16, 10 (2021). PMID: 34246791.
