Single-Cell RNA Sequencing Identifies Molecular Biomarkers Predicting Response to CDK4/6 Inhibition in Metastatic HR+/HER2- Breast Cancer

Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i), combined with endocrine therapy, represent the standard treatment for metastatic HR+/HER2- breast cancer (mBC). Despite their established efficacy, intrinsic resistance affects approximately one-third of patients, posing a significant challenge and underscoring the importance of identifying reliable predictive biomarkers. This study utilizes single-cell RNA sequencing (scRNAseq) of metastatic site biopsies to unveil cellular and molecular biomarkers predictive of CDK4/6i response. Through this analysis, we have identified and validated the tumor-specific molecular biomarkers across two independent mBC patient cohorts using bulk RNAseq data from baseline fresh biopsies or FFPE slides of HR+/HER2- mBC patients prior to CDK4/6i treatment. Additionally, the data indicate that baseline predictors may include tumor-infiltrating cytotoxic NK and T lymphocytes, suggesting the potential utility of checkpoint inhibitors in CDK4/6i progressors. These potentially actionable insights underscore the importance of optimizing therapeutic strategies based on microenvironment-specific changes observed following disease progression. Overall design: Metastatic tumor biopsies were obtained from HR+/HER2- BC patients pre-CDK4/6i treatment at baseline (BL) and/or at disease progression. BL samples were from CDK4/6i responders (n = 8, median progression-free survival [mPFS] = 17 months), while progressors were categorized as early-progressors (EP, n = 3, mPFS = 3 months) and late-progressors (LP, n = 8, mPFS = 10 months). Metastatic sites included liver (10), pleural effusions (6), ascites (2), and bone (1). InferCNV distinguished tumor cells, and functional analysis utilized the Molecular Signatures Database. Signature panel genes were validated in two independent cohorts (MDACC n = 89, Korean n = 61) with RNAseq data from baseline biopsies from HR+/HER2- mBC patients pre-CDK4/6i treatment.