Paired Tissue and Circulating Tumor DNA Profiling Reveals Novel ESR1 Amplifications and CDK4/6 Inhibitor Resistance Mechanisms in Hormone Receptor-Positive/HER2-Negative Metastatic Breast Cancers.

With the availability of new treatment options, such as selective estrogen receptor degraders, cyclin-dependent kinase 4/6 inhibitors (CDK4/6i), and PIK3CA inhibitors for hormone receptor-positive (HR+)/human epidermal growth factor receptor 2-negative (HER2-) advanced and metastatic breast cancers, the identification of ESR1, PIK3CA, and biomarkers associated with CDK4/6i resistance has become increasingly critical. Liquid biopsy has emerged as a convenient and valuable tool for molecular profiling and disease monitoring. In this study, we evaluate real-world genomic profiling in both tumor and liquid biopsies to determine how the complementary use of both specimen types optimizes the identification of resistance biomarkers. We performed a real-world analysis using comprehensive genomic profiling (CGP) of tumor tissue and liquid biopsy (circulating tumor DNA [ctDNA]) samples from patients with HR+/HER2- advanced or metastatic BC. Forty-five patients with ctDNA results were analyzed, including 19 with matched tumor tissue CGP results. The most frequently altered genes were TP53, PIK3CA, and ESR1. ESR1 alterations were detected in 33% (15/45) of HR+ BC patients in ctDNA testing. In the 19 tissue CGP and ctDNA pairs, ESR1 missense mutations were found in 53% (n = 10/19) of cases, increasing to 79% (n = 15/19) when including ESR1 amplifications identified in tissue only (n = 5/19, 26%). In our study, ESR1 missense mutations and ESR1 amplifications were mutually exclusive. Copy number variations (CNVs) frequently detected included CCND1 (n = 7/19, 37%) and FGFR1 (n = 6/19, 32%) that are associated with CDK4/6i resistance and co-occurring with ESR1 amplifications. Those copy number amplifications were detected in tissue only. PIK3CA variants showed high concordance between tissue and ctDNA (91%), whereas 67% of TP53 variants were detected exclusively in ctDNA. ctDNA effectively detected ESR1 and PIK3CA mutations relevant to targeted therapies, whereas it failed to identify a subset of patients with ESR1 amplification, as well as other CNVs potentially mediating CDK4/6i resistance. TP53 mutations observed at high frequency in ctDNA without tissue confirmation may reflect clonal hematopoiesis, and their prognostic significance requires careful interpretation. As a liquid biopsy is valuable for disease and treatment monitoring, understanding its strengths and limitations is essential. Tissue CGP remains a complementary tool for establishing baseline tumor profiles and enhancing CNV detection in comprehensive molecular characterization.