Purpose: We undertook this study to determine the prevalence of estrogen receptor (ER) α (ESR1) mutations throughout the natural history of hormone-dependent breast cancer and to delineate the functional roles of the most commonly detected alterations.

Experimental Design: We studied a total of 249 tumor specimens from 208 patients. The specimens include 134 ER-positive (ER⁺/HER2⁻) and, as controls, 115 ER-negative (ER⁻) tumors. The ER⁺ samples consist of 58 primary breast cancers and 76 metastatic samples. All tumors were sequenced to high unique coverage using next-generation sequencing targeting the coding sequence of the estrogen receptor and an additional 182 cancer-related genes.

Results: Recurring somatic mutations in codons 537 and 538 within the ligand-binding domain of ER were detected in ER⁺ metastatic disease. Overall, the frequency of these mutations was 12% [9/76; 95% confidence interval (CI), 6%–21%] in metastatic tumors and in a subgroup of patients who received an average of 7 lines of treatment the frequency was 20% (5/25; 95% CI, 7%–41%). These mutations were not detected in primary or treatment-naïve ER⁺ cancer or in any stage of ER⁻ disease. Functional studies in cell line models demonstrate that these mutations render estrogen receptor constitutive activity and confer partial resistance to currently available endocrine treatments.

Conclusions: In this study, we show evidence for the temporal selection of functional ESR1 mutations as potential drivers of endocrine resistance during the progression of ER⁺ breast cancer. Clin Cancer Res; 20(7); 1757–67. ©2014 AACR.