Assessing Endocrine Resistance: Monitoring Circulating ESR1 mutations in Irosustat-Treated ER positive Breast Cancer

Purpose: We aimed to investigate the prevalence and spectrum of ESR1 mutations alongside cell-free DNA (cfDNA) dynamics in patients with estrogen receptor-positive metastatic breast cancer recruited to the phase II IRIS study who had progressed on first-line aromatase inhibitor (AI) therapy and then continued their AI in combination with Irusostat (40mg), an irreversible steroid sulfatase inhibitor. Methods: cfDNA was isolated from 96 serial plasma samples from 24 patients, alongside primary tumour DNA (n = 16), and analysed by next-generation sequencing using a custom-designed mutation panel on the Illumina NovaSeq platform. Results: Thirteen of 16 tumour DNA samples harboured at least one somatic mutation across nine genes. Twenty one of 24 patients (88%) had at least one somatic mutation in cfDNA (248 total mutations across 10 genes). Circulating tumour DNA ESR1 mutations (ct ESR1 m) were the most prevalent, present in 16 patients (76%) with both stable (SD) and progressive disease (PD), showing no clear association with disease progression. Eleven patients had polyclonal ct ESR1 m within the ligand-binding domain, six at baseline, while five harboured a single ct ESR1 m variant. Five other patients acquired polyclonal mutations over treatment. Conclusion: Analysis of serial plasma samples revealed frequent detection of polyclonal ct ESR1 m in patients recruited to the IRIS study with both SD and PD. These findings underscore the challenge of targeting a single ESR1 mutation and emphasise the need for careful patient selection, specifically those with wild-type ESR1 , in trials investigating sequential estrogen-lowering therapies.