Biomedical subjects

William R Harrington

Publications and source records attributed to William R Harrington.

4 recordsLinked to original sources

PubMed2006

Estrogen regulation of the glucuronidation enzyme UGT2B15 in estrogen receptor-positive breast cancer cells.

Estrogens and androgens influence many properties of breast cancer cells; hence, regulation of local estrogen and androgen levels by enzymes involved in steroid hormone biosynthesis and metabolism would impact signaling by these hormones in breast cancer cells. In this study, we show that the UDP-glucuronosyltransferase (UGT) enzyme UGT2B15, a member of the UGT family of phase II enzymes involved in the glucuronidation of steroids and xenobiotics, is a novel, estrogen-regulated gene in estrogen receptor (ER)-positive human breast cancer cells (MCF-7, BT474, T47D, and ZR-75). UGT2B15 is the only UGT2B enzyme up-regulated by estrogen, and marked estradiol stimulation of UGT2B15 mRNA levels is observed, in a time- and dose-dependent manner. UGT2B15 stimulation by estradiol is blocked by the antiestrogen ICI182,780, but not by the translational inhibitor cycloheximide, indicating that UGT2B15 is likely a primary transcriptional response mediated through the ER. UGT2B15 up-regulation is also evoked by other estrogens (propylpyrazoletriol, genistein) and by the androgen 5alpha-dihydrotestosterone working through the ER, but not by other steroid hormone receptor ligands. Western blot and immunocytochemical analyses with several UGT2B-specific antibodies we have designed and steroid glucuronidation assays indicate a large increase in both cellular UGT2B15 protein and enzyme activity after estrogen treatment. Due to the important role of UGT enzymes in forming conjugates between steroids and glucuronic acid, thereby inactivating them and targeting them for removal, the estrogen-induced up-regulation of UGT2B15 might have a significant moderating effect on estrogen and androgen concentrations, thereby reducing their signaling in breast cancer cells.

Androgens↗

PubMed2005

Estrogen dendrimer conjugates that preferentially activate extranuclear, nongenomic versus genomic pathways of estrogen action.

Estrogenic hormones are classically thought to exert their effects by binding to nuclear estrogen receptors and altering target gene transcription, but estrogens can also have nongenomic effects through rapid activation of membrane-initiated kinase cascades. The development of ligands that selectively activate only the nongenomic pathways would provide useful tools to investigate the significance of these pathways. We have prepared large, abiotic, nondegradable poly(amido)amine dendrimer macromolecules that are conjugated to multiple estrogen molecules through chemically robust linkages. Because of their charge and size, these estrogen-dendrimer conjugates (EDCs) remain outside the nucleus. They stimulate ERK, Shc, and Src phosphorylation in MCF-7 breast cancer cells at low concentrations, yet they are very ineffective in stimulating transcription of endogenous estrogen target genes, being approximately 10,000-fold less potent than estradiol in genomic actions. In contrast to estradiol, EDC was not effective in stimulating breast cancer cell proliferation. Because these EDC ligands activate nongenomic activity at concentrations at which they do not alter the transcription of estrogen target genes, they should be useful in studying extranuclear initiated pathways of estrogen action in a variety of target cells.

Adaptor Proteins, Signal Transducing↗

PubMed2003

Activities of estrogen receptor alpha- and beta-selective ligands at diverse estrogen responsive gene sites mediating transactivation or transrepression.

Estrogens exert their regulatory transcriptional effects, which can be stimulatory or repressive, at diverse gene sites via two estrogen receptors, ERalpha and ERbeta. Since these two ERs have different tissue distributions, ligands that have the capacity to selectively activate or inhibit these two ERs would be useful in elucidating the biology of these two receptors and might assist in the development of estrogen pharmaceuticals with improved tissue selectivity. We have developed several ligands that showed ERalpha or ERbeta selectivity at promoter-gene sites containing consensus estrogen response elements (EREs): ERalpha-selective agonist (propyl-pyrazole-triol (PPT)), ERalpha-selective antagonist (methyl-piperidino-pyrazole (MPP)), ERbeta-potency selective agonist (diarylpropionitrile (DPN)) and ERbeta-selective antagonist/ERalpha-agonist (R,R-tetrahydrochrysene (R,R-THC)). In this study, we have examined the activity of these compounds at a range of gene sites where ER stimulates gene expression through non-consensus EREs (complement C3), or multiple half-EREs (NHE-RF/EBP50), or by tethering to DNA via other proteins (TGF beta3 and progesterone receptor A/AP-1), and at gene sites where ER represses gene transcription (interleukin-6). At all of these genes, PPT showed full stimulation through ERalpha while displaying no agonism through ERbeta. MPP antagonized estradiol actions on gene transactivation and transrepression through ERalpha, with little or no effect on transcription mediated through ERbeta. DPN displayed subtype-selective agonism, being ca. 30-fold more potent through ERbeta. R,R-THC was a complete antagonist through ERbeta and displayed agonism through ERalpha, the level of which was promoter dependent. Because these ligands maintain their agonist or antagonist character and ER subtype-selectivity at gene sites of diverse nature, where estradiol is either stimulatory or inhibitory, these compounds should prove useful in elucidating the biological functions of ERalpha and ERbeta.

Cell Line, Tumor↗

PubMed2002

Antagonists selective for estrogen receptor alpha.

To develop compounds that are antagonists on ER(alpha), but not ER(beta), we have added basic side-chains typically found in nonsteroidal antiestrogens to pyrazole compounds that bind with much higher affinity to ER(alpha) than to ER(beta). In this way we have developed basic side-chain pyrazoles (BSC-pyrazoles) that are high affinity, potent, selective antagonists on ER(alpha). These BSC-pyrazoles are themselves inactive on ER(alpha) and ER(beta), and they antagonize E2 stimulation by ER(alpha) only. We investigated seven basic side-chain substituents on various alkyl-triaryl-substituted pyrazoles, and the most ER(alpha)-selective compound was methyl-piperidino-pyrazole (MPP). ER(alpha)-selective antagonism was observed on diverse reporter-promoter gene constructs containing estrogen response elements that are consensus, nonconsensus (pS2), or comprised of multiple half-estrogen response elements (NHERF/EBP50) and on genes in which ER works indirectly by tethering to other DNA-bound proteins (TGF(beta)3). In contrast to these BSC-pyrazoles, the antiestrogens trans-hydroxytamoxifen, raloxifene, and ICI 182,780 suppress E2 activity via both ER(alpha) and ER(beta). The most effective BSC-pyrazole, MPP, fully antagonized E2 stimulation of pS2 mRNA in MCF-7 breast cancer cells, consistent with the fact that these cells contain almost exclusively ER(alpha). These compounds should be useful in studying the biological functions of ER(alpha) and ER(beta) and in selectively blocking responses that are mediated through ER(alpha).

Estrogen Antagonists↗