EXONERATE-TRaCE: A Liquid Biopsy for Tracking Response to Anti–Epidermal Growth Factor Receptor–Based Therapy in Metastatic Colorectal Cancer

PURPOSE The EXOsome and cell-free microRNAs (miRNAs) of anti–epidermal growth factor receptor (EGFR) Resistance (EXONERATE) study previously developed and validated a liquid biopsy to predict the efficacy of first-line anti-EGFR therapy in metastatic colorectal cancer (mCRC) from treatment-naïve blood samples. This study evaluates the performance of the EXONERATE assay during therapy as a tool for monitoring the emergence of anti-EGFR resistance before it becomes clinically or radiologically apparent. METHODS Retrospective, single-center, longitudinal, REMARK-compliant biomarker study assessing the ability of a liquid biopsy to predict response to anti-EGFR monoclonal antibodies and to track resistance development longitudinally. Fifty-two patients with RAS wild-type mCRC provided 142 plasma samples during administration of first-line anti-EGFR–based therapy. RESULTS Baseline EXONERATE scores predicted response with 83.3% specificity and 77.3% sensitivity (area under the receiver operating characteristic curve AUROC = 89.4%). Longitudinal analyses demonstrated that biomarker changes closely tracked tumor burden and anticipated clinical and radiological progression, outperforming conventional tumor markers CEA and CA19-9. Stepwise optimization produced a reduced panel (four cell-free-miRNAs and five exo-miRNAs), EXONERATE-Tracking Response and Clinical Evolution (TRaCE), which retained high predictive accuracy (AUROC = 94.4%) while enabling more practical, repeated monitoring. EXONERATE-TRaCE values increased during the 3 months preceding the first clinical or radiological evidence of progression and reliably detected early progression in both left- and right-sided tumors, including patients with lung metastases, and identified resistance up to 3 months before radiological confirmation. CONCLUSION EXONERATE-TRaCE extends the predictive utility of the original EXONERATE assay into a practical monitoring tool. This liquid biopsy detects emerging anti-EGFR resistance earlier than conventional markers and may inform treatment monitoring and support timely clinical decision making in mCRC.