Transfer Learning-Based Quantification of Mitotic Activity and Its Prognostic and Molecular Associations in Lung...

Background: Automated assessment of mitotic activity may provide a reproducible marker of tumor proliferation, but computational mitosis recognition remains challenging because of cross-domain variability. We developed a transfer-learning framework to quantify mitotic activity in lung squamous cell carcinoma (LUSC) and investigated its clinical and molecular associations. Methods: A ResNet-34 classifier was trained using MIDOG source-domain data and adapted to TCGA-LUSC diagnostic whole-slide images. The target cohort included 478 patients and 512 whole-slide images. Tumor regions and mitotic figures were annotated by two pathologists, and 128 × 128-pixel patches at 40× resolution were analyzed. After identifying patient overlap in the initial patch-level partition, we implemented patient-level stratified five-fold cross-validation to obtain leakage-mitigated performance estimates and out-of-fold predictions. Tumor-area mitosis proportion (TAMP) was defined as the proportion of evaluable tumor-region patches classified as mitosis-positive. Prognostic associations were evaluated using multivariable Cox proportional-hazards regression adjusted for age, sex, and pathological stage. Results: The classifier achieved an AUC of 0.972 (95% CI, 0.965-0.980) on the internal MIDOG test set. Direct application to TCGA-LUSC yielded an AUC of 0.81, whereas target-domain fine-tuning produced an initial patch-level AUC of 0.93. Patient tracing identified overlap in the original partitions, with 142 patients (29.7%) contributing patches to multiple subsets, including 67 patients (14.0%) represented in both training and test sets. Leakage-mitigated patient-level cross-validation yielded a mean AUC of 0.88 (95% CI, 0.86-0.90) and PR-AUC of 0.24. A patient-disjoint WSI holdout evaluation yielded an AUC of 0.87 (95% CI, 0.84-0.90) and PR-AUC of 0.22. High TAMP was associated with poorer overall survival (log-rank p=0.017; HR = 1.73, 95% CI, 1.12-2.68) and remained significant after adjustment for age, sex, and pathological stage (adjusted HR = 1.62, 95% CI, 1.03-2.54; p=0.036). High-TAMP tumors showed enrichment of proliferation- and growth-related transcriptional programs, whereas immune, genomic, and predicted drug-response associations were modest and hypothesis-generating. Conclusions: TAMP represents an image-derived measure of mitotic activity associated with survival and molecular characteristics in TCGA-LUSC. The findings support further investigation of computational mitotic phenotyping, but external multicenter validation and prospective clinical assessment are required before clinical translation.