USE OF FINE-NEEDLE ASPIRATES FOR THE IDENTIFICATION OF MUTATIONS WITHIN THE ESTROGEN RECEPTOR-a HORMONE-BINDING DOMAIN IN TAMOXIFEN-RESISTANT HUMAN BREAST CARCINOMAS
1Department of Endocrinology and Diabetes, Sir Charles Gairdner Hospital, Nedlands, Australia 2PathCentre, Queen Elizabeth II Medical Centre, Nedlands, Australia 3Mount Medical Centre, Perth, Australia 4Department of Medical Oncology, Sir Charles Gairdner Hospital, Nedlands, Australia 5Department of Pharmacology, University of Western Australia, Queen Elizabeth II Medical Centre, Nedlands, Western Australia 6009, Australia
Abstract. We investigated the hypothesis that mutations within estrogen receptor-a (ERa) may be one of the mechanisms of breast carcinoma progression to a hormone-independent phenotype. Fine-needle aspirations were collected from patients with tamoxifen resistant metastatic breast carcinomas. Total RNA was isolated and the entire ERa hormone-binding domain (HBD) amplified by RT-PCR using 4 pairs of overlapping primers. PCR products were screened for mutations by single-stranded conformational polymorphism (SSCP) analysis and DNA sequencing. Although no mutations were detected by SSCP analysis, two silent polymorphisms were identified by automated DNA sequencing at codons 538 [GAC - GAT (Asp)] and 594 [ACA - ACG (Thr)]. The codon 538 variant was detected in tumour specimens at a low frequency whereas the codon 594 variant was detected in patients and healthy controls at a similar frequency. Exon 7-deleted variant was also detected in all tumour specimens. Fine-needle aspirations, in conjunction with RT-PCR, could be a rapid and reliable method for the mutational analysis of the ERa HBD. However, mutations within the ERa HBD do not appear to represent a significant mechanism in the development of antiestrogen resistance.
