Formation and turnover of long-chain fatty acid esters of 5-androstene-3β,17β -diol in estrogen receptor positive and negative human mammary cancer cell lines in culture

Abstract

Microsomal preparations derived from bovine placenta cotyledons, previously investigated as a convenient source of fatty acyl coenzyme A: estradiol-17β-acyl transferase, have been shown to acylate other steroids bearing 3β- or 17β-hydroxyl groups. In the presence of 0.1 mM oleoyl CoA, the apparent K mvalues for dehydroepiandrosterone, testosterone, and 5-androstene-3β,17β-diol (Δ 5-DIOL) were 45,67, and 20 μM, respectively. Acylation of Δ 5-DIOL occurred at either the 3β- or 17β-positions to give monoesters. Testosterone, estradiol-17β, and Δ 5-DIOL acted as competitive inhibitors for the acylation of the 3β-hydroxyl group of dehydroepiandrosterone (K i values 71, 75, and 41 μM, respectively). Such data indicate that a single enzyme of wide substrate specificity may be involved in these acylation reactions. When estrogen receptor (ER) positive and negative human mammary cancer cell lines were incubated with 10 nM [ 3H]Δ 5-DIOL, intracellular accumulation of Δ 5-DIOL long-chain fatty acid esters occurred; rates being higher ( p<0.001) in ER negative cells (MDA-MB-231 and MDA-MB-330) compared to MCF-7 cells (ER positive), and higher ( P<0.005) in MDA-MB-231 cells compared to ZR-75-1 cells (ER positive). After exposure to 10 nM [ 3H]Δ 5-DIOL for 16 h, the total labeled steroid fatty acid fraction was composed predominately of Δ 5-DIOL-3β- and 17β-monoesters (∼85%), the remainder containing approximately equal amounts of Δ 5-DIOL-diesters and dehydroepi- androsterone-3β-esters. Subsequent transfer to medium lacking Δ 5-DIOL was accompanied by a breakdown of the labeled esters, which was more rapid in the ER positive cell lines. During this period, intracellular free Δ 5-DIOL levels rapidly declined in MDA-MB-330 cells but were maintained in MCF-7 cells, presumably by binding to ER. This behavior parallels that of estradiol-17β previously observed in these cell lines and further emphasizes the potential importance of the adrenal-derived estrogen Δ 5-DIOL in consideration of a hormone-based etiology of human breast cancer.