Targeted NGS identifies activating ESR1mutations in ctDNA and CTCs of endocrine-resistant breast cancers.. Acquired activating ESR1 mutations in cell-free DNA and circulating tumor cells in estrogen receptor positive metastatic breast cancer

Acquired activating mutations in ESR1, the gene that encodes estrogen receptor-alpha (ERα) have recently been shown to drive resistance to endocrine therapy. Detection of these mutations in circulating tumor DNA (ctDNA) and circulating tumor cells (CTCs) would represent a new method for monitoring response to endocrine therapy and emergence of resistance. Through targeted next-generation sequencing, we detected 3 activating mutations in ESR1 (8.3%), 8 PIK3CA (16.7%) and 2 TP53 (4.2%) in ctDNA of 48 patients with ERα positive metastatic breast cancer who were receiving systemic therapy. Two other patients acquired mutations in ctDNA during therapy in ESR1 and PIK3CA. Of those individuals with >100 CTCs, one had an ESR1 p.D538G mutation, which matched the ctDNA. Lastly, 1 primary patient thought to have metastatic disease, but who was clear by scans had the ESR1 p.D538G mutation at 1.3% in ctDNA. Since these mutations predict for a constitutive ERα activity, implying a mutant ERα which is active even in the presence of low or absent levels of estradiol or anti-estrogens, both ctDNA and CTC liquid biopsies have potential for detecting emergence of resistance to systemic hormonal therapy.