Abstract PO3-07-04: Prediction of PAM50 molecular subtypes from H&E-stained breast cancer specimens using tumor microenvironment features and additive multiple instance learning models
Authors: Guramare Maria , Javed Syed Ashar , Kirkup Christian , Juyal Dinkar , Brosnan-Cashman Jacqueline , Mountain Victoria , Leung Ryan , Rahsepar Bahar , Abel John , Taylor-Weiner Amaro , Conway Jake 
Published: May 2024

Abstract
Background: PAM50, a 50-gene signature, classifies breast cancers into one of five subtypes (basal, luminal A, luminal B, HER2-enriched, and normal-like), revealing information about underlying tumor biology, and has emerged as a key prognostic indicator influencing treatment decisions. There is growing interest in bridging the gap between expression-based metrics and histopathology, where immunohistochemistry (IHC) and sequencing-based approaches have been proposed for this purpose. However, hematoxylin and eosin (H&E)-stained slides are ubiquitously utilized by pathologists for cancer diagnosis, while IHC and sequencing-based approaches require additional tissue and specialized processing and/or analysis. Here, we describe a computer vision-based approach to predict PAM50 classification using H&E-stained whole slide images (WSIs).
Methods: We obtained expression-based PAM50 subtype labels and corresponding H&E-stained WSIs for 961 breast carcinomas from the TCGA BRCA cohort. We used two separate machine learning (ML) approaches to predict PAM50 subtypes from WSIs. In the first approach, we deployed previously trained PathExplore models to extract quantitative human-interpretable features (HIFs) that summarize the TME. We subsequently trained random forest classification models on these HIFs to predict PAM50 subtypes. For the second approach, we developed additive multiple instance learning (aMIL) models. Additionally, we explored the effects of PAM50 subtype labeling and aggregation strategies beyond the 5-class approach. Our 3-class approach combines Luminal A and B, as seen in IHC efforts to increase agreement with PAM50 assays, while excluding Normal, a category containing few and heterogeneous samples. We also performed binary classification for each subtype in the 3-class model (e.g. luminal vs. other). Slides were split into training (60%), validation (20%), and test (20%) sets, stratified by PAM50 labels, and model performance was assessed using the area under the receiver operator curve (AUROC) metric on the held-out test set, using a one vs. rest approach for multi-class models. To establish a baseline for PAM50 prediction, we developed random forest classification models using only clinical covariates (tumor stage, histologic grade, histological subtype, and BRCA1/2 status).
Results: We compared the performance of our two ML models (HIF and aMIL) to that of the baseline model, and we report the AUROC values in Table 1. These models both performed well in predicting Basal, Luminal A, Luminal B, and Luminal (A+B), while the model performance was less strong for predictions of the HER2 and Normal classifications. The three-class model showed improved performance of predicting Luminal classifications relative to the five-class model that separates Luminal A and B. Although simplifying classification problems to a binary use case typically provides improved performance, this phenomenon was not observed for any of the PAM50 subtypes.
Conclusions: These results demonstrate that AI-powered digital pathology can accurately and reproducibly perform molecular-based classification tasks, such as predicting PAM50 classifications, using WSIs, suggesting a more efficient path toward clinically relevant breast cancer characterization.
Table 1. Performance of all models in predicting PAM50 molecular subtypes. AUROC values are shown. Shaded cells represent the best test-set performance for each class (row).
Citation Format: Maria Guramare, Syed Ashar Javed, Christian Kirkup, Dinkar Juyal, Jacqueline Brosnan-Cashman, Victoria Mountain, Ryan Leung, Bahar Rahsepar, John Abel, Amaro Taylor-Weiner, Jake Conway. Prediction of PAM50 molecular subtypes from H&E-stained breast cancer specimens using tumor microenvironment features and additive multiple instance learning models [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-07-04.