Assessment of selegiline and trilostane combined therapy efficacy for canine pituitary-dependent hypercortisolism treatment: A pilot randomized clinical trial.

Canine pituitary-dependent hypercortisolism (PDH) management with trilostane usually demands lifelong therapy. The greater the dose needed, the greater the risk of side effects. Selegiline therapy has been previously described but not commonly used for PDH treatment. The present work aimed to assess the efficacy of selegiline and trilostane combined therapy for canine PDH treatment. Fifteen client-owned dogs diagnosed with spontaneous PDH were enrolled. The patients were treated with trilostane (Tri group, n = 8, initial dose of 0.5 mg/kg, PO, q12h), or with trilostane and selegiline (Tri + Sel group, n = 7, initial trilostane dose of 0.5 mg/kg, PO, q12h and selegiline 1 mg/kg, PO, q24h). Dogs underwent clinical examination, serum biochemical analysis, urinalysis, abdominal ultrasound, and eACTH and post-ACTH cortisol measurements on treatment days zero (D0), 30 (D30), 90 (D90), and 180 (D180). There was a lack of adverse effects due to the combined therapy. Both groups showed a similar clinical response and lower post-ACTH cortisol levels at the study's end. There was no significant difference in trilostane dosage at D180 between groups. There was no documented increase in either right or left adrenal gland thickness in the Tri + Sel group in contrast with patients in the Tri group. However, there was no statistical difference between the groups regarding eACTH at D0 and D180. Patients in the Tri + Sel group achieved better serum triglycerides control at the end of the study. The association of selegiline with trilostane might be a feasible therapy for canine PDH; however, its eventual advantages need larger studies.

Cortisol Concentrations in Well-Regulated Dogs with Hyperadrenocorticism Treated with Trilostane.

BACKGROUND: There are no clear treatment guidelines for dogs with clinically well-regulated hyperadrenocorticism in which serum cortisol concentrations before and after an ACTH stimulation test performed 3-6 hours after trilostane administration are < 2.0 µg/dL. OBJECTIVE: To determine if serum cortisol concentrations measured before (Pre1) and after (Post1) ACTH stimulation at 3-6 hours after trilostane administration are significantly lower than cortisol concentrations measured before (Pre2) and after (Post2) ACTH stimulation 9-12 hours after trilostane administration, in a specific population of dogs with clinically well-regulated hyperadrenocorticism and Pre1 and Post1 <2 µg/dL. ANIMALS: Thirteen client-owned dogs with clinically well-regulated hyperadrenocorticism and Pre1 and Post1 serum cortisol concentrations <2.0 µg/dL 3-6 hours after trilostane administration. METHODS: Prospective study. Dogs had a second ACTH stimulation test performed 9-12 hours after trilostane administration, on the same day of the first ACTH stimulation test. Cortisol concentrations before and after ACTH stimulation were compared using a paired t-test. RESULTS: Cortisol concentrations before (1.4 ± 0.3 µg/dL) and after the first stimulation (1.5 ± 0.3 µg/dL, mean ± SD) were significantly lower than cortisol concentration before the second stimulation (3.3 ± 1.6 µg/dL, P = .0012 each). Cortisol concentration before the first stimulation was also significantly lower than cortisol concentration after the second stimulation (5.3 ± 2.4 µg/dL, P = .0001). CONCLUSIONS AND CLINICAL IMPORTANCE: In dogs with clinically well-regulated, trilostane-treated, hyperadrenocorticism, and cortisol concentrations <2 µg/dL before and after the first stimulation, a second ACTH stimulation test performed 9-12 hours after treatment can result in higher cortisol concentrations that could support continued trilostane treatment.

Trilostane therapy for treatment of spontaneous hyperadrenocorticism in cats: 15 cases (2004-2012).

BACKGROUND: Medical treatment with trilostane improves clinical signs, causes unclear insulin requirement changes, and variable survival times in cats. OBJECTIVES/HYPOTHESIS: To characterize the long-term efficacy of trilostane in treating cats with hyperadrenocorticism (HAC). ANIMALS: Fifteen client-owned cats with spontaneous HAC. METHODS: Multicenter descriptive retrospective study with a search performed on all medical records for cats diagnosed with spontaneous HAC. RESULTS: Clinical signs (13 of 15 cats) and ACTH stimulation testing results (13 of 15) improved with trilostane therapy. Diabetes mellitus was reported in 9/15 cases. Insulin requirements decreased by 36% within 2 months in 6/9 diabetic cats. Median survival time was 617 days for all cats (range 80-1,278 days). Complications included weight loss, urinary tract infections, chronic kidney disease, seizures, and recurrent pancreatitis. Hypocortisolemia was documented in 1 case. Cause of death occurred as a result of nonadrenal or nondiabetic illnesses (renal failure, seizures [caused by hypoglycemia or unknown]), or lymphoma. CONCLUSIONS AND CLINICAL IMPORTANCE: Trilostane ameliorates clinical signs of HAC in cats, is tolerated well in the long term, and can lead to improved regulation of diabetes.

Developmental programming: impact of prenatal testosterone excess on ovarian cell proliferation and apoptotic factors in sheep.

Prenatal testosterone (T) excess leads to reproductive dysfunctions in sheep, which include increased ovarian follicular recruitment and persistence. To test the hypothesis that follicular disruptions in T sheep stem from changes in the developmental ontogeny of ovarian proliferation and apoptotic factors, pregnant Suffolk sheep were injected twice weekly with T propionate or dihydrotestosterone propionate (DHT; a nonaromatizable androgen) from Days 30 to 90 of gestation. Changes in developmental expression of proliferating cell nuclear antigen (PCNA), BCL2, BAX, activated CASP3, and FAS/FASLG were determined at Fetal Days 90 and 140, 22 wk, 10 mo, and 21 mo of age by immunocytochemisty. Prenatal T treatment induced changes in expression of proliferative and apoptotic markers in a follicle-, age-, and steroid-specific manner. Changes in BAX were evident only during fetal life and PCNA, BCL2, and CASP3 only postnatally. Prenatal T and not DHT increased PCNA and decreased BCL2 in granulosa/theca cells of antral follicles at 10 and 21 mo but decreased CASP3 in granulosa/theca cells of antral follicles at 22 wk (prepubertal) and 10 and 21 mo. Both treatments decreased BAX immunostaining in granulosa cells of Fetal Day 90 primordial/primary follicles. Neither treatment affected FAS expression at any developmental time point in any follicular compartment. Effects on BAX appear to be programmed by androgenic actions and PCNA, BCL2, and CASP3 by estrogenic actions of T. Overall, the findings demonstrate that fetal exposure to excess T disrupts the ovarian proliferation/apoptosis balance, thus providing a basis for the follicular disruptions evidenced in these females.

Developmental programming: prenatal androgen excess disrupts ovarian steroid receptor balance.

Steroid hormones play an important role in reproduction and the receptors through which they signal change in a developmental time, follicle stage, and cell-specific manner. Disruption in steroid receptor expression affects follicle formation and differentiation. In this study, using prenatal testosterone (T) and dihydrotestosterone (DHT)-treated female sheep as model systems, we tested the hypothesis that prenatal androgen excess disrupts the developmental ontogeny of ovarian steroid receptor protein expression. Pregnant Suffolk ewes were injected twice weekly with T propionate or DHT propionate (a non-aromatizable androgen) in cottonseed oil from days 30 to 90 of gestation. Changes in ovarian estrogen receptors (ER; ESR1, ESR2), androgen receptor (AR) and progesterone receptor (PGR) proteins were determined at fetal (days 90 and 140), postpubertal (10 months), and adult (21 months; only prenatal T-treated sheep studied) ages by immunohistochemistry. Prenatal T and DHT treatment induced selective increase in AR but not ER or PGR expression in the stroma and granulosa cells of fetal days 90 and 140 ovaries. An increase in ESR1 and decrease in ESR2 immunostaining coupled with increased AR expression were evident in granulosa cells of antral follicles of 10- and 21-month-old prenatal T but not DHT-treated females (analyzed only at 10 months). These findings provide evidence that an early increase in ovarian AR is the first step in the altered ovarian developmental trajectory of prenatal T-treated females, and manifestations of postnatal ovarian dysfunction are likely facilitated via altered equilibrium of antral follicular granulosa cell ER/AR protein expression.

Chemometric and chemoinformatic analyses of anabolic and androgenic activities of testosterone and dihydrotestosterone analogues.

Predictive quantitative structure-activity relationship (QSAR) models of anabolic and androgenic activities for the testosterone and dihydrotestosterone steroid analogues were obtained by means of multiple linear regression using quantum and physicochemical molecular descriptors (MD) as well as a genetic algorithm for the selection of the best subset of variables. Quantitative models found for describing the anabolic (androgenic) activity are significant from a statistical point of view: R(2) of 0.84 (0.72 and 0.70). A leave-one-out cross-validation procedure revealed that the regression models had a fairly good predictability [q(2) of 0.80 (0.60 and 0.59)]. In addition, other QSAR models were developed to predict anabolic/androgenic (A/A) ratios and the best regression equation explains 68% of the variance for the experimental values of AA ratio and has a rather adequate q(2) of 0.51. External validation, by using test sets, was also used in each experiment in order to evaluate the predictive power of the obtained models. The result shows that these QSARs have quite good predictive abilities (R(2) of 0.90, 0.72 (0.55), and 0.53) for anabolic activity, androgenic activity, and A/A ratios, respectively. Last, a Williams plot was used in order to define the domain of applicability of the models as a squared area within +/-2 band for residuals and a leverage threshold of h=0.16. No apparent outliers were detected and the models can be used with high accuracy in this applicability domain. MDs included in our QSAR models allow the structural interpretation of the biological process, evidencing the main role of the shape of molecules, hydrophobicity, and electronic properties. Attempts were made to include lipophilicity (octanol-water partition coefficient (logP)) and electronic (hardness (eta)) values of the whole molecules in the multivariate relations. It was found from the study that the logP of molecules has positive contribution to the anabolic and androgenic activities and high values of eta produce unfavorable effects. The found MDs can also be efficiently used in similarity studies based on cluster analysis. Our model for the anabolic/androgenic ratio (expressed by weight of levator ani muscle, LA, and seminal vesicle, SV, in mice) predicts that the 2-aminomethylene-17alpha-methyl-17beta-hydroxy-5alpha-androstan-3-one (43) compound is the most potent anabolic steroid, and the 17alpha-methyl-2beta,17beta-dihydroxy-5alpha-androstane (31) compound is the least potent one of this series. The approach described in this report is an alternative for the discovery and optimization of leading anabolic compounds among steroids and analogues. It also gives an important role to electron exchange terms of molecular interactions to this kind of steroid activity.

Effects of castration and hormone replacement on male sexual behavior and pattern of expression in the brain of sex-steroid receptors in BALB/c AnN mice.

Little is known about the hormonal regulation of sexual behavior and about the pattern of expression in the brain of sex-steroid receptors in the BALB/c AnN strain of mice (Mus musculus). In this study, 8-week old male BALB/c AnN mice were castrated and the temporal course of decline of sexual behavior was studied, as well as the effects of daily treatment with either testosterone propionate (TP), estradiol benzoate (EB), or dihydrotestosterone propionate (PDHT). Castration resulted in rapid decline of sexual behavior, in both control or vehicle-treated mice. TP maintained full sexual behavior of castrated mice, while PDHT or EB did not have this effect. The expression of ER-alpha dropped nearly 50% after castration, and this pattern remained in TP or PDHT-treated mice, while EB increased the ER-alpha mRNA levels to almost the same values as in intact control mice. The same pattern was found when ER-beta mRNA levels were analyzed. The expression of the PR-A/B gene in the different brain regions in intact mice and after castration, or among the differently treated mice, showed significant differences between normal and castrated mice at all times in all brain regions studied, with the exception of the frontal cortex. Castration reduced the expression of AR by 10-fold, as compared to intact control mice, while TP or PDHT treatment returned its expression to the same levels as in intact control mice, in all brain areas studied. The changes are more prominent in POA-HIP than in HYP and CF. These results demonstrated a rapid decline of sexual behavior in this strain of mice after castration, and show that only TP was able to maintain male sexual behavior, with no correlation with the pattern of expression of sex hormone receptors in specific areas of the mouse brain.

Effects of androgens and antiandrogens on the quantitative immunohistochemistry of gonadotrope cells in prepubertal male rats.

In the male rat, androgens are involved in the feedback regulation of gonadotropin synthesis and secretion. Specific androgen-receptor blockade by the nonsteroidal antiandrogens, flutamide and Casodex, has proven to be a valid tool for studying androgen effects in vivo. The aim of the present study was to investigate the effect of antiandrogen administration at the pituitary level by evaluating the changes in gonadotropes through quantitative immunohistochemistry, and by comparing these alterations with the effect of androgen deprivation by castration either with or without subsequent androgen replacement. Male Sprague-Dawley rats (23 days old) were randomly divided into 5 groups for the following treatments: (a) controls; (b) flutamide-injected (10 mg/rat/day in a gelatin vehicle); (c) Casodex-injected (10 mg/rat/day in an oil vehicle); (d) castrated, and (e) castrated and dihydrotestosterone propionate-replaced (40 microg/rat/day in an oil vehicle). Groups were then sacrificed after 10 days of maintenance under each condition. Pituitaries were fixed in Bouin's fluid and embedded in paraffin. Serial sections (4 micrometer) were obtained at different levels and immunostained by means of the primary murine monoclonal antibodies anti-FSH and anti-LH and a peroxidase-mediated EnVision System (Dako). Measurements of volume density (VD) and individual mean cell area were made by means of an image-analysis system (Imaging Technology, Optimas). Serum FSH and LH levels were determined by radioimmunoassay (RIA). Serum gonadotropin levels, VD, and mean cell area increased significantly in the flutamide-treated, Casodex-treated, and castrated groups (p < 0.05). Androgen replacement in the castrated rats, however, reduced VD, mean cell area, and serum gonadotropins to levels comparable to those of controls. We conclude that either androgen blockade by antiandrogens or castration produce an enhancement in the gonadotrope cell population in prepubertal rats, as shown by an increase in both VD and mean cell area, as well as an elevation in FSH- and LH-immunoreactive cells. These observations correlate well with the changes found in the levels of circulating gonadotropins as measured by RIA.

Treatment of persistent pubertal gynecomastia with dihydrotestosterone heptanoate.

Four boys with persistent pubertal gynecomastia were given intramuscular dihydrotestosterone heptanoate (DHT-hp) at 2 to 4-week intervals for 16 weeks. By the end of treatment, breast size in all four boys had decreased 67% to 78%. Initial plasma levels of gonadotropins, estradiol, testosterone, and dihydrotestosterone (DHT) were normal. Mean plasma DHT concentration rose with the injections of DHT-hp, and remained elevated throughout the treatment period. Estradiol, LH, FSH, and testosterone decreased during treatment, as did 24-hour urinary LH and FSH. No regrowth of breast tissue was observed 6 to 15 months after treatment, although hormone concentrations had returned to near pretreatment values by 2 months after the last injection. DHT-hp has potential to be an effective medical therapy for persistent pubertal gynecomastia.

[Androgenic therapy and hepatocellular carcinoma. Report of a case]. / Terapia androgénica y carcinoma hepatocelular. Informe de un caso.

A 65 years old, female patient with acquired aplastic anemia secondary to frequent exposure to hair dye. While on treatment with anabolic steroids hormone became jaundiced and developed hepatomegaly eight months later. During laparotomy the liver was enlarged, hard, with multiple whitish nodules on its surgace but was otherwise normal. Liver biopsy showed hepatocellular carcinoma, there were not cirrhosis niether hemochromatosis. A review of the related literature was done and discussed on the experimental and clinical evidences that suggested that androgens may play same role on the etiology of liver cancer.