At least 70% of breast cancers are classified as estrogen-receptor (ER) positive and consequently, patients with this disease receive some form of endocrine therapy as part of their adjuvant care. Whilst endocrine therapy reduces the risk of disease recurrence through inhibition of the ER signalling network, a third of all patients will experience drug resistance and disease relapse. Current second-line therapeutic strategies often include the sequential delivery of alternative endocrine therapies. However, responses are often short-lived, suggestive of an underlying mechanism of reduced endocrine sensitivity rather than drug-specific resistance. In experimental model systems, the rapid kinetics and reversibility of acquired drug resistance, combined with the absence of genetic mutations suggests a potential epigenetic basis for drug insensitivity. Here, we interrogate the epigenetic profile of multiple models of estrogen resistance to determine if there are common underlying DNA methylation aberrations that are associated with reduced endocrine response. Differential methylation analysis using Illumina 450K arrays was performed on endocrine sensitive and resistant breast cancer cells. Notably, we found that hypermethylation of estrogen-responsive enhancer regions are a frequent and significant feature of the resistant phenotypes. The methylation status of these regulatory regions was found to be a critical determinant in the prevention of ER binding and loss of endocrine sensitivity in human disease. Our data highlight the considerable potential for an enhanceosome DNA methylation prognostic signature that promises to stratify ER-positive breast cancer patients as responders to endocrine therapy.