Abstract P5-09-16: Contribution of estrone-sulfate to cell proliferation in aromatase inhibitor resistance of estrogen receptor positive breast cancer
Authors: T Higuchi, M Endo, T Gohno, T Hanamura, T Niwa, Y Yamaguchi, J Horiguchi, I Takeyoshi, S-I Hayashi
Journal: Cancer Research (2013)

Abstract [Background] Estrogen receptor (ER) positive breast cancer was composed of about 70% of clinical breast cancer and therapy for the ER positive breast cancer have progressed, but some patients recur after endocrine therapy including aromatase inhibitors (AIs). The switch of proliferation drive to human epidermal growth factor receptor 2 was reported as one of AI resistance mechanisms, but ER status of many recurrent patients after AI treatment was maintained or slightly decreased. Moreover, plasma concentration of estrone-sulfate (E1S) is mostly unchanged after menopause and treatment of AI. Thus, we established AI resistant model with conserved ER expression and its activity from MCF-7 cell, and found that E1S contributed to proliferation of the AI-resistant cells. [Material and Method] We introduced plasmids carrying ERE-green fluorescent protein (GFP) gene to MCF-7 (E10), and then aromatase gene was additionally introduced to E10 (E10arom). E10arom was cultured in estrogen-depleted medium including testosterone and letrozole, monitoring GFP as ER activity for about three months. We finally established ER positive Letrozole-Resistant cell lines (LR cells). In addition, to confirm whether E1S-dependent growth in LR cells could be reproduced in ER positive breast cancer cell line, we also established human steroid sulfatase (STS) gene-transfected MCF-7 (MCF-STS) and organic anion transporter peptide (OATP) 4A1 gene-transfected MCF-7 (MCF-4A1). ER activity of these cell lines was examined by ERE-luciferase reporter assay, and response to each steroid and drug was investigated by cell proliferation assay. Quantitative PCR was used for comparing expression of mRNA in these cell lines. [Results] mRNA expression of STS and four OATPs including OATP4A1 increased in LR cells comparing to E10arom. Moreover, there was no change in mRNA expressions of efflux drug transporters, aromatase-metaboic enzyme and enzymes related with production for estrogenic androgen. ERE-luciferase assay with E1S showed higher ER activity of LR cells than E10arom, and LR cells were more proliferated by E1S in dose-dependent manner than the parental cell. STS inhibitor alone and its combination with letrozole were inhibitory to proliferation of LR cells. LR cells were also inhibited by selective estrogen receptor modulator (SERM) and downregulator (SERD). In proliferation assay of MCF-STS and MCF-4A1, we observed that these cell lines also proliferated by E1S in dose-dependent manner as LR cells did. [Discussion] Since ER status of recurrent patients after AI treatment was mostly unchanged, our model could be more practical for analysis of AI resistance in ER positive breast cancer. These results showed that combined induction of STS and OATP contributed to estradiol (E2) production from E1S in LR cells, suggesting that analysis of E1S as alternative E2 productive pathway needed investigation of both STS and transporter of E1S. The combination of STS inhibitor with letrozole was effective to LR cells, and SERM and SERD were also effective to these cells, suggesting that combined therapy of STS inhibitor with AI or switching AI to SERM or SERD was beneficial for AI-resistant breast cancer. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P5-09-16.