Performance of an optimized methylation-protein multi-cancer early detection (MCED) test classifier

Background: Multi-cancer early detection (MCED) tests can detect several cancer types and stages. We previously developed a methylation and protein (MP V1) MCED classifier with a target specificity of ≥98.0% (Gainullin, medRxiv, 2025.08.24.25334244). Herein, we describe a refined MP V2 classifier that was identified based on the evaluation of classifier model architectures that improved performance. Methods: Compared to MP V1, the MP V2 classifier was trained to achieve increased early-stage sensitivity at a case-control target specificity of ≥97.0%. MP V2 classifier architecture was developed using a training set (654 cancer; 2,373 non-cancer) partitioned into 5-fold cross validation and mini-holdout sets. Locked candidate models were compared using an independent mini-holdout test set (110 cancer; 509 non-cancer). MP V1 and MP V2 classifier performance were compared using a previously described test set (729 cancer; 2,434 non-cancer), and MP V2 performance was also evaluated in an independent clinical validation test set (324 cancer; 800 non-cancer) that was designed to more closely mimic the intended use population. Results: Compared to MP V1, MP V2 (specificity of 97.4%) demonstrated a 7.3% increase in overall sensitivity, with sensitivity increases of 7.6%, 9.2%, 8.3%, 8.0%, 3.8%, and 13.3% for stages I, II, stages I/II, III, IV, and unknown, respectively, in the test set (Table 1). In the independent validation set, MP V2 (specificity 97.4%; 95% CI: 96.0-98.3%) overall sensitivity was 41.4%, with sensitivities of 16.0%, 31.3%, 22.8%, 52.9%, and 83.1% for stages I, II, stages I/II, III and IV, respectively. Excluding breast and prostate cancers, MP V2 overall sensitivity was 55.6%, with sensitivities of 26.8%, 42.9%, 34.8%, 63.6%, and 89.3% for stages I, II, stages I/II, III, and IV, respectively, in the validation set. Conclusion: In a case-control setting, the MP V2 classifier offered improved sensitivity for early-stage cancers at a lower specificity target. Table 1. MP V1 and MP V2 classifier performance in the test set. a breast and prostate excluded. MP V1 Classifier Performance (95% CI) MP V2 Classifier Performance (95% CI) Measured Specificity (N=2,434) 98.5% (97.9-98.9) 97.4% (96.7-97.9) Overall Sensitivity, all cancers (N=729) 50.9% (47.3-54.5) 57.8% (54.1-61.3) Stage I (n=182) 15.4% (10.9-21.3) 21.4% (16.1-27.9) Stage II (n=163) 38.0% (30.9-45.7) 46.0% (38.5-53.7) Stage III (n=180) 67.8% (60.6-74.2) 76.1% (69.4-81.8) Stage IV (n=172) 85.5% (79.4-90.0) 89.5% (84.1-93.3) Unknown Stage (n=32) 37.5% (22.9-54.7) 50.0% (33.6-66.4) Stages I/II (n=345) 26.1% (21.7-31.0) 33.0% (28.3-38.2) Overall Sensitivity, (N=590) a 56.8% (52.8-60.7) 64.1% (60.1-67.8) Stage I (n=145) 17.2% (12.0-24.2) 24.8% (18.5-32.4) Stage II (n=109) 48.6% (39.4-57.9) 57.8% (48.4-66.6) Stage III (n=151) 73.5% (66.0-79.9) 81.5% (74.5-86.8) Stage IV (n=155) 86.5% (80.2-91.0) 90.3% (84.6-94.0) Unknown Stage (n=30) 40.0% (24.6-57.7) 53.3% (36.1-69.8) Stages I/II (n=254) 30.7% (25.4-36.6) 39.0% (33.2-45.1)