Effects of androgens, progesterone and their antagonists on the developmental competence of in vitro matured bovine oocytes. (25/384)

The aim of the present study was to determine whether androgens and progesterone influence the in vitro maturation of bovine oocytes as assessed by cleavage rates and competence to form blastocysts after in vitro fertilization. Bovine cumulus-oocyte complexes were cultured (n = 20 per drop) for 22-24 h at 38.5 degrees C in TCM-199 medium supplemented with 10% oestrous cow serum, eCG (2.5 iu ml(-1)) and a range of treatments that included aromatizable (testosterone; 100 nmol l(-1)) and non-aromatizable (dihydrotestosterone; 100 nmol l(-1)) androgens, an androgen antagonist (flutamide; 36 micromol l(-1)), progesterone (300 nmol l(-1)) and a progesterone antagonist (mifeprisone, RU486; 100 nmol l(-1)). Production of inhibin A, total alpha-subunit, activin A and follistatin by each group of cumulus-oocyte complexes was also measured, since inhibin-related peptides have been implicated as modulators of oocyte maturation and their production may be influenced by steroids and anti-steroids. Both testosterone and dihydrotestosterone increased oocyte cleavage rate (25%; P < 0.01) and dihydrotestosterone also increased (24%; P < 0.05) the proportion of oocytes that reached the >/= eight-cell stage. However, neither androgen affected blastocyst yield, or the proportion of blastocysts that hatched. The stimulatory effect of dihydrotestosterone on cleavage rate was reduced by flutamide but the anti-androgen had no effect when tested alone. Treatment with testosterone, but not dihydrotestosterone, decreased (P < 0.05) endogenous follistatin and increased (P < 0.05) the activin A:follistatin ratio in maturation medium. Concentrations of inhibin A, total alpha-subunit and activin A were not affected significantly by androgen or flutamide. Addition of progesterone or the anti-progestin mifepristone to cumulus-oocyte complexes had no effect on cleavage rate. However, progesterone reduced by approximately 40% (P < 0.05) the proportions of both total oocytes and cleaved oocytes that formed blastocysts. This effect was partially reversed by mifepristone. Neither progesterone nor mifepristone affected inhibin A, activin A or follistatin production. However, total alpha-subunit concentration was significantly greater in the progesterone-treated group than in the controls (50%; P < 0.05), indicating that the negative effect of progesterone on blastocyst yield may be mediated by increased inhibin alpha-subunit expression by cumulus cells.  (+info)

Phenacetin deacetylase activity in human liver microsomes: distribution, kinetics, and chemical inhibition and stimulation. (26/384)

Microsomal and cytosolic phenacetin deacetylase activities were examined in human liver and kidneys. Kinetic properties of the activities were also studied in human liver microsomes. Phenacetin deacetylase activity was predominantly localized in the liver microsomal fraction. The specific activities of phenacetin deacetylation in liver cytosol and in kidney microsomes and cytosol were all less than 5% of that in liver microsomes. In human liver microsomes, Eadie-Hofstee plots for phenacetin deacetylation were monophasic, indicating a single-enzyme catalytic reaction. The Michaelis-Menten parameters, K(m) and V(max), for the deacetylation were 4.7 mM and 5.54 nmol/min/mg of protein, respectively. The intrinsic clearance, calculated as V(max)/K(m), was 1.18 microl/min/mg of protein. Although the organophosphate bis(4-nitrophenyl)phosphoric acid markedly inhibited the reaction in human liver microsomes, the activity has a tolerance to the treatment of phenylmethylsulfonyl fluoride, a serine hydrolase inhibitor. Prazosin, a peripheral alpha(1)-adrenergic antagonist, noncompetitively inhibited the phenacetin deacetylation with a K(i) value of 19.0 microM. Flutamide, a nonsteroidal androgen receptor antagonist, stimulated the activity by up to 349%. This increase was accompanied by a decrease in the K(m) value and no change in the V(max) value, resulting in an increase in the intrinsic clearance by up to 700% of the control. These results suggest that the phenacetin deacetylase localized in human liver microsomes has not only a catalytic site but also a negative and/or positive modulation site or sites.  (+info)

Exclusive androgenic effect of dehydroepiandrosterone in sebaceous glands of rat skin. (27/384)

In order to analyze the hormonal effects of dehydroepiandrosterone (DHEA) in skin sebaceous glands, the precursor steroid was administered to ovariectomized (OVX) female Sprague-Dawley rats at a dose of 30 mg applied on the dorsal skin, twice daily, for 3, 6 and 12 months. In a parallel experiment, female OVX rats were treated with DHEA at the same daily percutaneous dose of 30 mg, alone or in combination with the antiandrogen Flutamide or the pure antiestrogen EM-800, for 12 months, in order to determine the androgenic and/or estrogenic components of DHEA action. Treatment of female OVX rats with DHEA resulted in a similar mild to moderate hyperplasia of the sebaceous glands of both dorsal (site of application) and ventral skin, as illustrated by an increase in the number and size of the acini. The above-indicated effects were observed at all time intervals studied, beginning at 3 months of treatment, and they were not further increased after longer term administration of DHEA (for 6 and 12 months). The addition of Flutamide to DHEA treatment completely prevented the DHEA-induced changes in the sebaceous glands, whereas the antiestrogen EM-800 had no effect. The present data indicate an exclusive androgenic stimulatory action of DHEA on the sebaceous glands, thus pointing out the importance of local intracrine DHEA transformation into androgens for skin anatomical integrity and function, while showing that estrogens, if active in rat skin, do not originate from DHEA.  (+info)

Efficacious chemoprevention of primary prostate cancer by flutamide in an autochthonous transgenic model. (28/384)

Although the etiology of prostate cancer is still not clear, family history, hormones, and age are thought to play a role in its initiation and progression. There is no cure for the advanced disease. Because prostate cancer initially develops as an androgen-dependent tumor, agents with antiandrogen activity have become the focus for chemoprevention of this disease. A pilot study was undertaken to test the efficacy of flutamide (an antiandrogen) in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model of prostate cancer. Three groups of mice received s.c. implantation of slow-release flutamide pellets: (a) low-dose flutamide group (6.6 mg/kg); (b) high-dose flutamide group (33 mg/kg); and (c) control placebo group. Efficacy was measured by the absence of palpable tumor formation. Prostate tissues/tumors were harvested for evaluation by molecular and histology techniques. The low-dose flutamide group did not differ significantly from the placebo group, in which palpable tumors initially presented at 17 weeks of age, and by 33 weeks, all of the animals developed palpable tumors. In the high-dose flutamide group, however, tumors did not appear until 24 weeks, a lag of 7 weeks, and by 34 weeks, 42% of the animals were still tumor free. The period of time at which 50% of the animals had tumors was 33 weeks in the high-dose flutamide group, 24.5 weeks in the low-dose flutamide group, and 24.5 weeks in the placebo group. The difference between the placebo and high-dose flutamide groups was statistically significant (log rank, P = 0.0036; Wilcoxon's statistical analysis, P = 0.0060). Tumors from high-dose flutamide-treated animals were more differentiated and retained much of the normal glandular architecture compared with those of the placebo group, whose tumors consisted of sheets of poorly differentiated cells. The expression of T antigen in the prostate tissues of flutamide-treated animals (at 10 weeks age) was lower than that in the comparable placebo-treated group. Flutamide had the ability to suppress T antigen-driven carcinogenesis, resulting in a significant decrease in the incidence of prostate cancer and an increase in the latency period of prostate cancer in TRAMP mice.  (+info)

Rapid androgen actions on calcium signaling in rat sertoli cells and two human prostatic cell lines: similar biphasic responses between 1 picomolar and 100 nanomolar concentrations. (29/384)

Androgen-induced calcium fluxes and gap junctional intercellular communication (GJIC) were studied in three different cell types. A transient (2-3 min duration) increase in intracellular calcium levels was observed within 20-30 sec of androgen addition, which was followed by a plateau phase with steroid concentrations higher than 1 nM. The kinetics of the calcium responses were similar in immature rat Sertoli cells, which contain normal nuclear receptors; the human prostatic tumor cell line, LNCaP, which contains a mutated nuclear receptor; and the human prostatic cell line, PC3, which does not contain a nuclear receptor. The human A431 tumor cell line did not respond to androgens. Concentrations of testosterone and the synthetic androgen, R1881, between 1-1000 pM induced transient calcium increases with ED(50) values near 1 pM and 1 nM, whereas dihydrotestosterone (DHT) was not active at these concentrations. At concentrations higher than 1 nM, testosterone, R1881, and DHT were equipotent in stimulating an increase in calcium that lasted for more than 10 min, with ED(50) values between 5 and 20 nM. Testosterone covalently bound to albumin was also active, whereas 11 related androstane compounds as well as progesterone and estradiol-17beta were inactive at 1000 nM. The calcium response induced by the three androgens (10 nM) was abolished in all cell types by hydroxyflutamide (1000 nM) and finasteride (1000 nM), but not by cyproterone acetate (1000 nM). The calcium response was also abolished in the absence of extracellular calcium and strongly inhibited by the presence of verapamil. Exposure of the responsive cells to brief (150-sec) pulses of androgens generated calcium responses that were similar to those after continuous exposure. After exposure of Sertoli cells for only 30 sec to 100 nM testosterone, the calcium response lasted for at least 50 min. Although nuclear binding of androgens could be demonstrated, there was no evidence for tight binding to the plasma membrane under similar conditions. When protein synthesis was inhibited, an enhancement of GJIC between rat Sertoli cells, but not between LNCaP cells or PC3 cells, was observed within 15 min of the addition of 10 nM testosterone. Because nuclear androgens are not present in PC3 cells and many functional properties of the responsive system are different from the nuclear receptor in all three cell types, we postulate the existence of an alternative cell surface receptor system with biphasic response characteristics (high and low affinity). The calcium signals are probably coupled to the regulation of gap junctional efficiency between Sertoli cells. The low-affinity receptors may convey complementary androgen signals at elevated local levels such as in the testis, when nuclear receptors are (over)saturated.  (+info)

Flutamide-hydroxypropy-beta-chiyclodextrin complex: formulation, physical characterization, and absorption studies using the Caco-2 in vitro model. (30/384)

PURPOSE: The objective of this research was to formulate flutamide (FLT) in hydroxypropyl-beta-cyclodextrin (HPbetaCyD), and to investigate FLT transcellular permeation from the complex using the Caco-2 monolayer in vitro model. METHODS: Classical solubility data were used to derive thermodynamic parameters which, together with Differential Scanning Calorimetry (DSC), (1)H-NMR and (19)F-NMR, were used to characterize and derive stability constants for the FLT-HPbetaCyD complex. The Caco-2 cell line was used to examine the role of HPbetaCyD on the passage of FLT across cell monolayers in vitro. RESULTS: The solubility of FLT in water (1.46 mmol/L) increased almost 170 times (to 243.45 mmol/L) in the presence of 50% (w/v) HPbetaCyD. Solubility data for FLT in aqueous HPbetaCyD were used to derive thermodynamic parameters (DeltaG degrees at 298 K = -3.48, DeltaH degrees = 2.85, DeltaS degrees at 298 K = 21.24). The solubility of FLT in HPbetaCyD increased proportionally with an increase in temperature. The FLT-HPbetaCyD complex had an A(L)-type (DSC) isotherm, consistent with a linear increase in FLT solubility and unchanged stoichiometry. The DSC of free FLT and HPbetaCyD showed endothermic peaks at 110 degrees C and 300 degrees C, respectively. FLT-HPbetaCyD did not display a free-FLT endothermic response, but exhibited broadening of the endothermic peak in the HPbetaCyD region. (19)F- and (1)H-NMR chemical shifts of FLT moved upfield as a function of its increased solubility in the presence of HPbetaCyD. The FLT-HPbetaCyD stability constant, K(s) (1:1) was estimated to be 356 M(-1 )and 357 M(-1), from thermodynamic and (19)F NMR data, respectively. The apical-to-basal permeability coefficient (P(eff) = 4.75 x 10(-5) cm.s(-1)) for FLT across Caco-2 cell monolayers at 37; C increased as HPbetaCyD concentrations were reduced, indicative of transepithelial passage via passive diffusion of available free FLT in solution. Studies in the presence and absence of Ca(2+ )ruled out a significant paracellular transport component. CONCLUSIONS: FLT-HPbetaCyD is a relatively stable, 1:1 inclusion complex. Formation of this complex substantially increases the water solubility of FLT, but HPbetaCyD, except in high dilution, reduces transcellular passage of FLT in the Caco-2 cell in vitro model.  (+info)

A prospective randomized multicenter study of chlormadinone acetate versus flutamide in total androgen blockade for prostate cancer. (31/384)

BACKGROUND: A randomized multicenter study was conducted to investigate the efficacy of total androgen blockade (TAB) for patients with previously untreated prostate cancer using the steroidal anti-androgen chlormadinone acetate (CMA) and the non-steroidal anti-androgen flutamide. We also compared the liver dysfunction in these two arms. METHODS: From November 1995 to October 1997, 71 patients were registered into this study and 70 of them were eligible. RESULTS: There was no significant difference in the efficacy of TAB between CMA and flutamide at 24 weeks. The testosterone and prostate-specific antigen (PSA) levels in patients administered flutamide (Group II) increased significantly 3 days after the first dose of LH-RH analog, whereas no such increase was observed in patients administered CMA (Group I), indicating that CMA prevented the flare-up. Parameters of liver function, serum GOT and GPT levels, which were normal at the baseline, became abnormal in 30.0% and 35.3%, respectively, of patients in Group II. These figures were significantly higher than the corresponding figures of 6.3% and 12.5%, respectively, in Group I. When the degree of change in each of these parameters was analyzed, both GOT and GPT levels showed a significantly greater increase in Group II than in Group I. CONCLUSION: These results indicate that attention must be paid to changes in liver function during the administration of flutamide in patients with prostate cancer even if their baseline liver function is normal. It is also suggested that CMA may be better tolerated from the viewpoint of the drug effects on liver function.  (+info)

Flutamide versus prednisone in patients with prostate cancer symptomatically progressing after androgen-ablative therapy: a phase III study of the European organization for research and treatment of cancer genitourinary group. (32/384)

PURPOSE: Time to progression (TTP), overall survival, and quality of life (QL) were compared in patients with hormone-resistant prostate cancer (HRPC) treated with prednisone (5 mg orally, four times a day) or flutamide (250 mg orally, three times a day). PATIENTS AND METHODS: Symptomatic patients were randomized to receive either prednisone (101 patients) or flutamide (100 patients). Subjective response was assessed based on performance status, the use of analgesics, and the need to apply alternative palliative treatment. Prostate-specific antigen (PSA)-based biochemical response (>or= 50% reduction of baseline PSA) was recorded. At baseline and at 6-week intervals during follow-up, patients completed the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire C-30. RESULTS: There was no difference between the groups in median TTP (prednisone, 3.4 months; flutamide, 2.3 months) or overall survival (prednisone, 10.6 months; flutamide, 11.2 months). In the prednisone group, 56% of the patients experienced a subjective response, compared with 45% in the flutamide group (P: = .18). The median response duration was 4.8 months for prednisone and 4.2 months for flutamide. A biochemical response was observed in 21% and 23% of the prednisone and flutamide groups, respectively. Gastrointestinal toxicity was the reason for trial discontinuation in seven patients receiving flutamide and two patients receiving prednisone. The QL assessment parameters favored the use of prednisone with statistically significant differences in pain, fatigue, role functioning, appetite loss, gastrointestinal distress, and overall QL. CONCLUSION: In symptomatic HRPC, treatment with prednisone or flutamide leads to similar rates of TTP and overall survival and no difference in subjective or biochemical response. The QL results favor the use of low-cost prednisone in patients with HRPC.  (+info)