Molecular minimal residual disease in resected non-small cell lung cancer: “flattening the curve”

The prognostic value of molecular residual disease (MRD) is well established across multiple solid tumor types. In non-small cell lung cancer (NSCLC), studies consistently show that patients who are MRD-negative—defined as having undetectable circulating tumor DNA (ctDNA) in peripheral blood—after surgery experience markedly higher disease-free survival (DFS) compared with MRD-positive patients. Reported hazard ratios for relapse range from 8.3 to 18.7, underscoring the strong and clinically meaningful prognostic impact of MRD status (1-6).

Clinicians and molecular biologists are increasingly exploring this non-invasive molecular tool to guide adjuvant therapy decisions. It holds particular promise for identifying patients who are likely cured after surgery and may therefore safely forgo adjuvant treatment.

Proof of concept has already been demonstrated in colorectal cancer (7). In the randomized DYNAMIC trial, patients with stage II colon cancer underwent R0 resection. In the experimental arm, MRD-positive patients received oxaliplatin-based or fluoropyrimidine chemotherapy, while MRD-negative patients received no further treatment. The ctDNA-guided approach was non-inferior to standard management for 2-year relapse-free survival (RFS), with only about half as many patients in the experimental group received adjuvant chemotherapy. After a median follow-up of 6 years, 5-year overall survival (OS) was similar in both groups (8), indicating that post-operative therapy can be safely de-escalated based on ctDNA-guided decisions.

To safely omit treatment in MRD-negative patients, however, it is essential to correctly identify them. Notably, DFS curves for MRD-negative patients are never completely flat (Figure 1), indicating that a subset still relapses. This group typically represents 10% to 20% of NSCLC patients (1,5,6,9-13).

Figure 1 Schematic representation of DFS according to MRD status. Repeated testing improves detection of residual disease in non-cured patients, while persistently negative results indicate a higher likelihood of cure and may flatten the DFS curve (arrow). Additional parameters, such as cerebrospinal fluid analysis and genetic/epigenetic profiling of the resected tumor, could further refine patient stratification and help identify those cured by surgery, in whom adjuvant therapy may be safely omitted. DFS, disease-free survival; MRD, molecular residual disease.

Several strategies aim to further improve the accuracy of MRD negativity and to “flatten” the DFS curve (Figure 1). The most common approach relies on ultra-sensitive, tumor-informed next-generation sequencing (NGS) assays that track specific genetic alterations identified in the primary tumor. Although highly sensitive, this technique is limited to patients with detectable mutations in tumor tissue. Emerging approaches using cell-free DNA (cfDNA) methylation (14) and fragmentation features (15) may enhance sensitivity. Multi-omic approaches (e.g., combining mutation tracking, methylation, and fragmentomics) may further improve both sensitivity and specificity.

Because the genomic alterations present in blood are rare events, repeated testing also improves the detection of patients at risk of relapse. This was shown by the Guangdong Lung Cancer Institute (China) in a prospective, observational study of 261 surgically treated NSCLC patients (stages I–III). A total of 948 blood samples were collected post-operatively on a predefined schedule (1 month post-surgery, then every 3 to 6 months). The initial results were reported after a median follow-up of 19.7 months (9), with a recent update at 43.4 months (10).

In the updated analysis, the negative predictive value (NPV)—the proportion of MRD-negative patients who remained relapse-free—was 76.5% when MRD testing was performed only once after surgery (landmark analysis; 56 of 238 MRD-negative patients relapsed). In contrast, with longitudinal testing every 3 to 6 months, the NPV rose to 93.2% (only 13 of the 192 MRD-negative patients relapsed). This clearly shows, even if MRD follow-up was not strictly conducted every 3 to 6 months for all patients, that repeated testing significantly improves the ability to identify truly cured patients.

Therefore, regular post-surgical MRD testing could improve therapeutic decision-making. However, the optimal testing frequency remains to be determined (every 3 months? every 6 months?). In a treatment strategy where MRD-negative patients are not given adjuvant therapy, treatment could be initiated either at the first MRD-positive result or after two consecutive positive tests.

Interestingly, Zhang et al. (9) reported that 13 patients who remained MRD-negative throughout the longitudinal follow-up still experienced relapse. In more than half of these cases (7/13; 53.8%), the brain was the only site of progression. This suggests a limitation of plasma ctDNA analysis in detecting central nervous system (CNS) involvement. Similar conclusions were reported by Aoki et al. (16), who described two patients that developed post-operative brain metastases after initial neoadjuvant EGFR tyrosine kinase inhibitor, and who had negative plasma EGFR tests after surgery. Cerebrospinal fluid (CSF) analysis may provide additional insights into CNS relapse risk (17), though it is more invasive and harder to repeat than blood sampling. Risk factors such as age, tumor burden, disease stage, histology, and tumor’s genetic or epigenetic characteristics can help identify patients at risk of CNS relapse (18). Serum markers, such as neurofilament light chain and glial fibrillary acidic protein, may also be used to detect patients with brain metastases (19,20).

Of note, in 7 of the 13 patients who experienced relapse, ctDNA was undetectable even prior to surgery, indicating low-shedding tumors (21). For such patients, the lack of detectable ctDNA in plasma—even when the tumor is present—limits MRD-based assessment of surgical efficacy. Some of the risk factors and potential serum markers mentioned above may also be used to predict relapse in these low-shedding tumors.

In conclusion, the vast majority of patients who remain MRD-negative during follow-up have a very favorable prognosis and are strong candidates for therapeutic de-escalation. Efforts should focus on optimizing the identification of these patients, and the validity of a de-escalation strategy must be tested in prospective interventional trials (22,23). For instance, the prospective multicenter CTONG 2201 trial (NCT05457049) is evaluating the hypothesis that no adjuvant therapy is needed for patients with longitudinal undetectable MRD. Stage IB–IIIA NSCLC patients who have undergone radical resection and have two consecutively undetectable MRD results (first at days 3–10, then at days 30±7 after surgery) will be enrolled and monitored with imaging and MRD testing every 3 months without adjuvant treatment (24). Analysis of 2-year DFS will allow conclusions to be drawn regarding such a strategy, including the optimal timing and frequency of blood testing. However, the absence of a control arm without MRD monitoring limits the overall evaluation of this approach. Beyond these clinical implications, several important challenges remain, including the costs and reimbursement of repeated tests, the feasibility of integrating their turnaround time into routine care, the specific difficulties associated with low shedders and brain progression, the impact on the quality of life of patients managed without adjuvant chemotherapy, and the ethnic and geographic generalizability of the findings. Considerable work is still needed before MRD testing can be implemented in routine clinical practice.

Acknowledgments

None.

Footnote

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