cfDNA-based testing multi-cancer early detection and minimal residual disease monitoring represents a transformative approach to diagnosing cancer in its early stages and therapeutic monitoring. These tests analyze fragments of tumor-derived DNA circulating in the blood, offering a non-invasive way to detect and monitor cancer. However, developing a cfDNA-based in vitro diagnostic (IVD) requires navigating a complex regulatory framework, particularly to gain U.S. Food and Drug Administration (FDA) approval. This article explores the journey from the discovery of cfDNA to the FDA requirements for clinical validity, safety, and efficacy, while addressing the ethical considerations of such tests in a clinical trial setting. It ends with a discussion about the CONSORT 2010 clinical trial reporting guidelines with respect to cfDNA studies.
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History of cfDNA Based Multi-Cancer Early Detection and Minimal Residual Disease Monitoring
The history of cfDNA begins in 1948 with the pioneering discovery by Mandel and Metais, who first detected cfDNA in the blood plasma of healthy individuals (Mandel & Metais, 1948). Their discovery initially drew limited attention but would ultimately open new paths in medical diagnostics.
In the 1970s and 1980s, cfDNA research found early applications in prenatal diagnostics and organ transplant rejection monitoring (Lo et al., 1997). This was a key turning point, as scientists realized cfDNA could carry crucial biological information beyond that of endogenous DNA, including information about fetal DNA in maternal blood and donor DNA in organ recipients, which marked the beginning of its diagnostic utility.
In the 2000s, cfDNA research gained significant momentum in oncology due to advances in sequencing technology. Researchers discovered that cfDNA in cancer patients often contained tumor-specific genetic alterations, which could be analyzed for diagnostic purposes (Stroun et al., 1987). Tumor-derived cfDNA, or circulating tumor DNA (ctDNA), proved capable of revealing key genomic information, making it an ideal biomarker for cancer detection. In 2008, researchers (Diehl et al., 2008) established the concept of using circulating tumor DNA (ctDNA) as a “liquid biopsy” to monitor tumor mutations in the blood. Vogelstein and his team demonstrated that ctDNA could be used to detect specific mutations associated with cancer, making it a powerful tool for tracking tumor dynamics and potentially aiding in early diagnosis and monitoring of minimal residual disease (MRD).
Advancements in high-throughput sequencing technologies and the decreasing cost of genomic analysis have enabled cfDNA to become a realistic option for early cancer detection and MRD monitoring. These developments support the emergence of liquid biopsies, which have been analytically validated in studies examining accuracy and utility in cancer diagnostics (Bettegowda et al., 2014).
An excellent recent review article by Heitzer et al. (2023) provides an up-to-date summary of cfDNA’s applications in oncology, highlighting its potential in detecting early-stage cancers, monitoring treatment response, and guiding therapeutic decisions. This article underscores the importance of cfDNA technology in transforming cancer diagnostics and sets the scientific foundation for developing regulatory frameworks to evaluate these novel tests.
Regulatory Pathway for IVD Approval of cfDNA Based Tests
The FDA's new final rule on laboratory-developed tests (LDTs) marks a significant shift in regulatory oversight, with the agency now asserting direct control over LDTs, which were previously under the purview of the Centers for Medicare and Medicaid Services (CMS) via the Clinical Laboratory Improvement Amendments (CLIA). Traditionally, LDTs, which are diagnostic tests designed, manufactured, and used within a single clinical laboratory, were exempt from FDA's premarket review. This exemption allowed laboratories flexibility in developing customized tests for unique or emerging medical needs, including rare diseases and novel biomarkers. However, the new rule requires these tests to undergo FDA review for safety and efficacy if they are deemed high-risk, effectively classifying them similarly to in vitro diagnostic (IVD) devices. This shift means that laboratories must now ensure their tests meet FDA standards for clinical validity and analytical performance, which could include conducting preclinical studies, providing detailed documentation, and in some cases, submitting for premarket approval (PMA) or clearance under 510(k) pathways.
Moving forwards, any cfDNA-based cancer diagnostic test to reach patients in the U.S., must first receive FDA approval as an in vitro diagnostic (IVD) device. The FDA evaluates IVDs based on a risk classification system that determines the required level of review, with cfDNA-based tests typically categorized as Class II or Class III due to their potential impact on patient outcomes. Three key attributes of a cfDNA must be rigorously demonstrated:
Analytical Validity: To obtain FDA approval, the analytical validity of a cfDNA test must be thoroughly validated. This involves confirming that the test reliably detects cfDNA with high sensitivity and specificity, particularly at the low concentrations typical of early-stage cancer and MRD. Critical metrics like the limit of detection, specificity, and accuracy must be rigorously demonstrated.
Clinical Validity: Clinical validity is a key requirement for FDA approval, necessitating robust evidence that the test can accurately distinguish between individuals with and without cancer (or with and without residual disease). Clinical validity often involves large-scale clinical trails demonstrating that the cfDNA test can detect cancer at early stages or identify MRD accurately, with minimal false positives or negatives.
Clinical Utility: The final regulatory requirement is to show that the test has clinical utility, meaning it benefits patient outcomes or assists in medical decision-making. This can involve demonstrating that the test results lead to improved cancer detection rates, more timely intervention, or better post-treatment monitoring and is typically a component of the clinical trials supporting an IVD application.
Clinical Trial Design Considerations for MCED and MRD Tests
Designing clinical trials for multi-cancer early detection (MCED) and MRD tests presents unique challenges due to the complexity of the test and the diversity of cancer types. A robust design framework for clinical trials involving cfDNA-based tests will include essential elements of the CONSORT 2010 clinical trial reporting guidelines including ethical considerations, randomization, endpoint selection, diagnostic metrics like sensitivity, specificity, and positive predictive value, all contextualized for laboratory tests.
Ethical Considerations:
Clear, informed consent is critical in trials involving cfDNA-based tests. Patients must understand the implications of a positive or negative test result, especially given the potential anxiety surrounding cancer risk. Managing the psychological impact of false positives (which could lead to unnecessary anxiety and follow-up procedures) and false negatives (which may give patients a false sense of security). Because cfDNA-based tests may carry significant costs, ensuring equitable access and diverse representation in clinical trials is essential. This helps prevent healthcare disparities and ensures that the test performs effectively across various demographic groups.
Randomization
Randomization in clinical trials is fundamental to reducing bias and ensuring that any observed effects are likely due to the intervention rather than other confounding variables. For cfDNA-based tests, randomization might entail assigning participants to an intervention group (where the cfDNA test is used alongside standard methods) and a control group (where only standard diagnostic procedures are used). This approach helps isolate the cfDNA test's performance in detecting cancer or minimal residual disease (MRD) and enables a clearer understanding of its potential clinical benefit.
Endpoint Selection
Endpoints in clinical trials for cfDNA-based tests should align with the test's purpose—whether for screening, early detection, or monitoring disease recurrence. In the context of cancer, common primary endpoints include disease-free survival, overall survival, and progression-free survival. For cfDNA tests, however, surrogate endpoints such as the test's ability to detect recurrence or new cancer cases earlier than traditional methods may be appropriate.
Types of Endpoints:
Primary Endpoint: For cfDNA-based screening, a primary endpoint might be the proportion of early-stage cancers detected accurately compared to a standard method. In MRD monitoring, the primary endpoint might be the time to detection of recurrence.
Secondary Endpoints: Secondary endpoints could include test accuracy, patient adherence, and impact on treatment decisions. Including these helps provide a comprehensive evaluation of the test’s performance and utility.
Sensitivity, Specificity, and Positive Predictive Value (PPV)
Diagnostic accuracy metrics are critical in evaluating cfDNA-based tests.
Sensitivity: In this context, sensitivity is the test’s ability to correctly identify individuals with the disease (true positive rate). High sensitivity is crucial for early-stage cancer detection because false negatives can delay treatment. For an MRD test, high sensitivity ensures that residual disease is detected as early as possible, allowing timely intervention.
Specificity: Specificity represents the test’s ability to correctly identify individuals without the disease (true negative rate). In cancer screening, high specificity minimizes false positives, reducing unnecessary follow-up testing and associated patient anxiety. Specificity is equally important in MRD monitoring to prevent misclassification and overtreatment.
Positive Predictive Value (PPV): PPV indicates the likelihood that a positive test result accurately reflects disease presence. In cfDNA testing, a high PPV is particularly valuable in screening programs where a low prevalence of disease might otherwise lead to more false positives.
Population Selection
Choosing an appropriate trial population is crucial for the success of these studies. For MCED tests, this may involve enrolling a diverse cohort, including individuals with an elevated cancer risk (e.g., older adults or those with family histories of cancer). For MRD trials, patients who have completed primary treatment for cancer but remain at risk for recurrence are often included.
Statistical Considerations
Rigorous statistical methods are essential to assess the test’s performance accurately, especially given the low prevalence of early-stage cancer in the general population. Large sample sizes are often necessary to achieve statistical significance and ensure that the test’s positive predictive value remains high.
Applying CONSORT 2010 Guidelines to Reporting cfDNA based MCED and MRD clinical trials
The CONSORT 2010 guidelines were developed as part of an ongoing effort to improve the transparency, reliability, and utility of clinical trial reports, thereby supporting evidence-based clinical decision-making. Included in the guidelines is a checklist that helps to ensure comprehensive and consistent reporting across clinical trials. Key aspects that apply to cfDNA-based trials (and others) include:
Trial Design Description: Describing the randomization process, intervention, and control groups clearly.
Participant Flow Diagram: Showing the flow of participants through the trial phases (enrollment, intervention allocation, follow-up, and analysis) to improve transparency.
Outcome Reporting: Clearly specifying and distinguishing between primary and secondary outcomes, reporting on diagnostic metrics, and including an analysis of sensitivity and specificity in the results.
Adherence and Loss-to-Follow-Up: For cfDNA-based tests, patient adherence and potential loss to follow-up can impact diagnostic outcomes. CONSORT recommends detailing this aspect, as it provides insight into the real-world applicability of the test.
Integrating these elements will help create a comprehensive design framework for cfDNA-based clinical trials, enhancing their quality, transparency, and impact..
Conclusion
The development of cfDNA-based IVDs for cancer detection and MRD monitoring represents a significant advance in precision oncology. By enabling earlier detection and close monitoring of residual disease, these tests have the potential to improve patient outcomes significantly. However, obtaining FDA approval requires careful attention to analytical and clinical validity, clinical utility, and ethical considerations throughout the trial design process. Rigorous trials, regulatory oversight, and transparent publications are essential to ensure that these tests offer safe, accurate, and accessible solutions for patients. As technology advances, it is vital to prioritize innovation and ethical integrity, ensuring that cfDNA diagnostics fulfill their promise of improving cancer diagnostics.
Suggested Additional Reading:
Mandel, P., & Metais, P. (1948) Les acides nucléiques du plasma sanguin chez l'homme. Comptes Rendus des Seances de la Societe de Biologie et de ses Filiales, 142, 241–243.
Lo, Y. M. D., et al. (1997) Presence of fetal DNA in maternal plasma and serum. The Lancet, 350(9076), 485–487.
Stroun, M., et al. (1987) Isolation and characterization of DNA from the plasma of cancer patients. Eur J Cancer Clin Oncol. Jun;23(6):707-12.
Diehl, F., et al. (2008). Circulating mutant DNA to assess tumor dynamics. Nature Medicine, 14, 985–990.
Bettegowda, C., et al. (2014). Detection of circulating tumor DNA in early- and late-stage human malignancies. Science Translational Medicine, 6(224), 224ra24.
Heitzer, E., et al. (2019). Current and future perspectives of liquid biopsies in genomics-driven oncology. Nat Rev Genet 2019 Vol. 20 Issue 2 Pages 71-88
Moher, D., et al. (2010) CONSORT 2010 explanation and elaboration: updated guidelines for reporting parallel group randomised trials. BMJ Vol. 340 Pages c869