toremifene and 4-hydroxytoremifene

A multicenter phase I pharmacokinetic study of a new triphenylethylene antiestrogen, toremifene, was examined in 70 patients with advanced breast cancer. Patients were randomized to receive single daily oral doses of either 10, 20, 40, 60, 200, or 400 mg for 8 weeks. Plasma toremifene and its major metabolites. N-desmethyltoremifene and 4-hydroxytoremifene, were determined weekly during therapy and at 0, 7, 14, and 21 days after the discontinuation of therapy. The time to reach steady-state plasma concentrations was between 1 and 5 weeks, with steady-state being achieved earlier (1-2 weeks) at daily doses of 200 and 400 mg. The time to peak concentration following oral doses of toremifene ranged from 1.5 to 4.5 h. The terminal half-life of elimination was 5.0, 6.0, and 5.0 days for toremifene, desmethyltoremifene, and 4-hydroxytoremifene, respectively. Plasma concentrations of 4-hydroxytoremifene were detectable only at high doses (200 and 400 mg/day) of toremifene. The results of this phase I pharmacokinetic study show that toremifene has metabolic and kinetic patterns that are similar to those previously reported with tamoxifen.

Topics: Administration, Oral; Breast Neoplasms; Chemical Phenomena; Chemistry; Chromatography, High Pressure Liquid; Drug Administration Schedule; Drug Evaluation; Estrogen Antagonists; Female; Half-Life; Humans; Multicenter Studies as Topic; Randomized Controlled Trials as Topic; Tamoxifen; Toremifene

Tamoxifen (Tam) is a selective estrogen receptor modulator used to inhibit breast tumor growth. Tam can be directly N-glucuronidated via the tertiary amine group or O-glucuronidated after cytochrome P450-mediated hydroxylation. In this study, the glucuronidation of Tam and its hydroxylated and/or chlorinated derivatives [4-hydroxytamoxifen (4OHTam), toremifene (Tor), and 4-hydroxytoremifene (4OHTor)] was examined using recombinant human UDP-glucuronosyltransferases (UGTs) from the 1A subfamily and human hepatic microsomes. Recombinant UGT1A4 catalyzed the formation of N-glucuronides of Tam and its derivatives and was the most active UGT enzyme toward these compounds. Therefore, it was hypothesized that single nucleotide polymorphisms (SNPs) in the promoter region of UGT1A4 have the ability to significantly decrease the glucuronidation rates of Tam metabolites in the human liver. In vitro activity of 64 genotyped human liver microsomes was used to determine the association between the UGT1A4 promoter and coding region SNPs and the glucuronidation rates of Tam, 4OHTam, Tor, and 4OHTor. Significant decreases in enzymatic activity were observed in microsomes for individuals heterozygous for -163G/A and -217T/G. These alterations in glucuronidation may lead to prolonged circulating half-lives and may potentially modify the effectiveness of these drugs in the treatment of breast cancer.

Topics: Genotype; Glucuronosyltransferase; Humans; Hydroxylation; Microsomes, Liver; Pharmacogenetics; Polymorphism, Single Nucleotide; Promoter Regions, Genetic; Tamoxifen; Toremifene

We investigated the in vitro metabolism and estrogenic and antiestrogenic activity of toremifene (TOR), tamoxifen (TAM) and their metabolites to better understand the potential effects of cytochrome P-450 2D6 (CYP2D6) status on the activity of these drugs in women with breast cancer. The plasma concentrations of TOR and its N-desmethyl (NDM) and 4-hydroxy (4-OH) metabolites during steady-state dosing with TOR were also determined. Unlike TOR, TAM and its NDM metabolite were extensively oxidized to 4-OH TAM and 4-OH-NDM TAM by CYP2D6, and the rate of metabolism was affected by CYP2D6 status. 4-OH-NDM TOR concentrations were not measurable at steady state in plasma of subjects taking 80 mg of TOR. Molecular modeling provided insight into the lack of 4-hydroxylation of TOR by CYP2D6. The 4-OH and 4-OH-NDM metabolites of TOR and TAM bound to estrogen receptor (ER) subtypes with fourfold to 30-fold greater affinity were 35- to 187-fold more efficient at antagonizing ER transactivation and had antiestrogenic potency that was up to 360-fold greater than their parent drugs. Our findings suggest that variations in CYP2D6 metabolic capacity may cause significant differences in plasma concentrations of active TAM metabolites (i.e., 4-OH TAM and 4-OH-NDM TAM) and contribute to variable pharmacologic activity. Unlike TAM, the clinical benefits in subjects taking TOR to treat metastatic breast cancer would not likely be subject to allelic variation in CYP2D6 status or affected by coadministration of CYP2D6-inhibiting medications.

Topics: Adult; Cytochrome P-450 CYP2D6; Humans; Male; Oxidation-Reduction; Selective Estrogen Receptor Modulators; Structure-Activity Relationship; Tamoxifen; Toremifene

Toremifene (TOR) is a selective estrogen receptor modulator used in adjuvant therapy for breast cancer and in clinical trials for prostate cancer prevention. The chemical structure of TOR differs from that of tamoxifen (TAM) by the presence of a chlorine atom in the ethyl side chain, resulting in a more favorable toxicity spectrum with TOR. In addition, some patients who fail on TAM therapy benefit from high-dose TOR therapy. Several studies have indicated that functional genetic variants in the TAM metabolic pathway influence response to therapy, but pharmacogenomic studies of patients treated with TOR are lacking. In this study, we examined individual variability in sulfation of 4-hydroxy TOR (4-OH TOR) (the active metabolite of TOR) in human liver cytosols from 104 subjects and found approximately 30-fold variation in activity. 4-OH TOR sulfation was significantly correlated (r = 0.98, P < 0.0001) with β-naphthol sulfation (diagnostic for SULT1A1) but not with 17β estradiol sulfation, a diagnostic substrate for SULT1E1(r = 0.09, P = 0.34). Examination of recombinant sulfotransferases (SULTs) revealed that SULT1A1 and SULT1E1 catalyzed 4-OH TOR sulfation, with apparent Km values of 2.6 and 6.4 μM and Vmax values of 8.5 and 5.5 nmol x min(-1) x mg protein(-1), respectively. 4-OH TOR sulfation was inhibited by 2,6-dichloro-4-nitrophenol (IC50 = 2.34 ± 0.19 μM), a specific inhibitor of SULT1A1. There was also a significant association between SULT1A1 genotypes and copy number and 4-OH TOR sulfation in human liver cytosols. These results indicate that variability in sulfation could contribute to response to TOR in the treatment of breast and prostate cancer.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Arylsulfotransferase; Child; Female; Genetic Variation; Humans; Liver; Male; Middle Aged; Pharmacogenetics; Protein Isoforms; Selective Estrogen Receptor Modulators; Tamoxifen; Toremifene; Young Adult

Tamoxifen remains the endocrine therapy of choice in the treatment of all stages of hormone-dependent breast cancer. However, tamoxifen has been shown to increase the risk of endometrial cancer which has stimulated research for new effective antiestrogens, such as droloxifene and toremifene. In this study, the potential for these compounds to cause cytotoxic effects was investigated. One potential cytotoxic mechanism could involve metabolism of droloxifene and toremifene to catechols, followed by oxidation to reactive o-quinones. Another cytotoxic pathway could involve the oxidation of 4-hydroxytoremifene to an electrophilic quinone methide. Comparison of the amounts of GSH conjugates formed from 4-hydroxytamoxifen, droloxifene, and 4-hydroxytoremifene suggested that 4-hydroxytoremifene is more effective at formation of a quinone methide. However, all three substrates formed similar amounts of o-quinones. Both the tamoxifen-o-quinone and toremifene-o-quinone reacted with deoxynucleosides to give corresponding adducts. However, the toremifene-o-quinone was shown to be considerably more reactive than the tamoxifen-o-quinone in terms of both kinetic data as well as the yield and type of deoxynucleoside adducts formed. Since thymidine formed the most abundant adducts with the toremifene-o-quinone, sufficient material was obtained for characterization by (1)H NMR, COSY-NMR, DEPT-NMR, and tandem mass spectrometry. Cytotoxicity studies with tamoxifen, droloxifene, 4-hydroxytamoxifen, 4-hydroxytoremifene, and their catechol metabolites were carried out in the human breast cancer cell lines S30 and MDA-MB-231. All of the metabolites tested showed cytotoxic effects that were similar to the parent antiestrogens which suggests that o-quinone formation from tamoxifen, droloxifene, and 4-hydroxytoremifene is unlikely to contribute to their cytotoxicity. However, the fact that the o-quinones formed adducts with deoxynucleosides in vitro implies that the o-quinone pathway might contribute to the genotoxicity of the antiestrogens in vivo.

Topics: Animals; Antineoplastic Agents; Benzoquinones; Breast Neoplasms; Cell Survival; Deoxyribonucleosides; DNA Adducts; Female; Glutathione; Indolequinones; Indoles; Microsomes, Liver; Quinones; Rats; Rats, Sprague-Dawley; Spectrometry, Mass, Electrospray Ionization; Tamoxifen; Toremifene; Tumor Cells, Cultured

Tamoxifen is widely prescribed for the treatment of hormone-dependent breast cancer, and it has recently been approved by the Food and Drug Administration for the chemoprevention of this disease. However, long-term usage of tamoxifen has been linked to increased risk of developing endometrial cancer in women. One of the suggested pathways leading to the potential toxicity of tamoxifen involves its oxidative metabolism to 4-hydroxytamoxifen, which may be further oxidized to an electrophilic quinone methide. The resulting quinone methide has the potential to alkylate DNA and may initiate the carcinogenic process. To further probe the chemical reactivity and toxicity of such an electrophilic species, we have prepared the 4-hydroxytamoxifen quinone methide chemically and enzymatically, examined its reactivity under physiological conditions, and quantified its reactivity with GSH. Interestingly, this quinone methide is unusually stable; its half-life under physiological conditions is approximately 3 h, and its half-life in the presence of GSH is approximately 4 min. The reaction between 4-hydroxytamoxifen quinone methide and GSH appears to be a reversible process because the quinone methide GSH conjugates slowly decompose over time, regenerating the quinone methide as indicated by LC/MS/MS data. The tamoxifen GSH conjugates were detected in microsomal incubations with 4-hydroxytamoxifen; however, none were observed in breast cancer cell lines (MCF-7) perhaps because very little quinone methides is formed. Toremifene, which is a chlorinated analogue of tamoxifen, undergoes similar oxidative metabolism to give 4-hydroxytoremifene, which is further oxidized to the corresponding quinone methide. The toremifene quinone methide has a half-life of approximately 1 h under physiological conditions, and its rate of reaction in the presence of excess GSH is approximately 6 min. More detailed analyses have indicated that the 4-hydroxytoremifene quinone methide reacts with two molecules of GSH and loses chlorine to give the corresponding di-GSH conjugates. The reaction mechanism likely involves an episulfonium ion intermediate which may contribute to the potential cytotoxic effects of toremifene. Similar to what was observed with 4-hydroxytamoxifen, 4-hydroxytoremifene was metabolized to di-GSH conjugates in microsomal incubations at about 3 times the rate of 4-hydroxytamoxifen, although no conjugates were detected with MCF-7 cells. Finally, these data suggest that quinone

Topics: Animals; Antineoplastic Agents, Hormonal; Breast Neoplasms; Cytochrome P-450 Enzyme System; Estrogen Receptor Modulators; Female; Glutathione; Humans; Hydroxylation; Indolequinones; Indoles; Mass Spectrometry; Oxidation-Reduction; Quinones; Rats; Rats, Sprague-Dawley; Tamoxifen; Toremifene; Tumor Cells, Cultured

Tamoxifen and its analogues 4-hydroxytamoxifen, toremifene, 4-hydroxytoremifene, clomifene and droloxifene were tested for clastogenic effects in a human lymphoblastoid cell line (MCL-5) expressing elevated native CYP1A1 and containing transfected CYP1A2, CYP2A6, CYP2E1 and CYP3A4 and epoxide hydrolase and in a cell line containing only the viral vector (Ho1). MCL-5 or Ho1 cells were incubated with 4-hydroxytamoxifen, 4-hydroxytoremifene, clomifene or droloxifene and the incidence of micronuclei estimated. With MCL-5 cells there was an increase in micronuclei with 4-hydroxytamoxifen, 4-hydroxytoremifene and clomifene but not with droloxifene. With Ho1 cells only 4-hydroxytamoxifen and 4-hydroxytoremifene caused an increase in micronuclei. MCL-5 cells were incubated with tamoxifen, 4-hydroxytamoxifen, toremifene, droloxifene, clomifene or diethylstilbestrol (0.25-10 microg/ml) for 48 h and subjected to 3 h treatment with vinblastine (0.25 microg/ml) to arrest cells in metaphase. The incidence of cells with chromosomal numerical aberrations (aneuploidy) was increased in cells treated with tamoxifen, 4-hydroxytamoxifen, toremifene, clomifene and diethylstilbestrol but not droloxifene. The frequency of cells with structural abnormalities (excluding gaps) was increased in cells treated with tamoxifen and toremifene but not 4-hydroxytamoxifen, clomifene, droloxifene or diethylstilbestrol. The clastogenic activities of tamoxifen (35 mg/kg), toremifene (36.3 mg/kg), droloxifene (35.2 mg/kg) and diethylstilbestrol (25 mg/kg) were compared in groups of four female Wistar rats. Each chemical was dissolved in glycerol formal, administered as a single dose by gavage and hepatocytes isolated by collagenase perfusion 24 h later. The cells were cultured in the presence of epidermal growth factor (40 ng/ml) for 48 h, colchicine (10 microg/ml) being added for the final 3 h of incubation. At least 100 chromosomal spreads were examined from each animal for the presence of numerical and structural abnormalities. The incidences of aneuploidy following treatment were: tamoxifen 81%, toremifene 46%, droloxifene 9.6%, diethylstilbestrol 45.7%, vehicle control 5.3%. The incidences of chromosomal structural abnormalities excluding gaps were: tamoxifen 4.3%, toremifene 0.8%, droloxifene 0.5%, diethylstilbestrol 0.8%, control 0.5%. The incidence of chromosomal structural aberrations excluding gaps in the treated animals was not statistically significantly different from controls exce

Topics: Aneuploidy; Animals; Anticarcinogenic Agents; Cell Line; Cell Nucleus; Clomiphene; Female; Humans; Liver; Lymphocytes; Micronucleus Tests; Rats; Rats, Wistar; Tamoxifen; Toremifene; Transfection

The ability of the anti-oestrogens tamoxifen, toremifene and their 4-hydroxy and N-desmethyl metabolites to modify doxorubicin (dox) toxicity to intrinsically resistant and multidrug resistant cell lines was compared, using human breast and lung cancer, and Chinese hamster ovary cell lines. The anti-oestrogens significantly enhanced dox toxicity to multidrug resistant, P-glycoprotein-positive cell lines, but did not affect toxicity to intrinsically resistant, P-glycoprotein-negative cells. Modification was observed at clinically achievable anti-oestrogen concentrations. Toremifene and tamoxifen would therefore appear to be good candidates for in vivo studies as MDR modulating agents in selected patients with P-glycoprotein-positive tumours.

Topics: Animals; Antibodies, Monoclonal; ATP Binding Cassette Transporter, Subfamily B, Member 1; Breast Neoplasms; Carrier Proteins; Cell Division; CHO Cells; Cricetinae; Doxorubicin; Drug Interactions; Drug Resistance; Drug Screening Assays, Antitumor; Epitopes; Estrogen Antagonists; Humans; Lung Neoplasms; Membrane Glycoproteins; Tamoxifen; Toremifene; Tumor Cells, Cultured

Toremifene, a new antiestrogenic antitumor compound, has several biologically active metabolites. The hormonal effects of the main metabolites resemble those of unchanged toremifene. The main metabolite in humans, N-demethyltoremifene, is bound to estrogen receptors (ER), inhibits the growth of MCF-7 cells, and exerts an antiestrogenic effect similar to that of toremifene. However, its antitumor effect in vivo against dimethylbenz(a)anthracene (DMBA)-induced rat mammary cancers is weaker than that of toremifene. Didemethyltoremifene has antiestrogenic actions in mouse and rat uterus at high doses. 4-Hydroxytoremifene is bound to ER with higher affinity and inhibits MCF-7 growth at concentrations lower than those of toremifene. It has a weaker intrinsic estrogenic effect than does toremifene. The efficacy of 4-hydroxytoremifene against DMBA-induced cancers is weak except at very high doses. Oxidations of N-demethylated metabolites to (deamino)hydroxylated compounds and carboxylic acids are the detoxification routes of toremifene. (deaminohydroxy)Toremifene has only weak hormonal actions at high doses and carboxylated metabolites have no estrogenic/antiestrogenic effects. The antitumor effect of toremifene in vivo is mainly due to unchanged toremifene, but hormonal effects (which may have a role in antitumor actions) are partly attributable to metabolites N-demethyltoremifene, didemethyltoremifene, (deaminohydroxy)toremifene, 4-hydroxy-N-demethyltoremifene, and 4-hydroxytoremifene, which have pharmacological properties similar to those of toremifene.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Antineoplastic Agents; Cell Division; Mammary Neoplasms, Experimental; Mice; Rats; Receptors, Estrogen; Tamoxifen; Toremifene; Tumor Cells, Cultured