Discovery of a small molecule that selectively destabilizes Cryptochrome 1 and enhances life span in p53 knockout mice.

Cryptochromes are negative transcriptional regulators of the circadian clock in mammals. It is not clear how reducing the level of endogenous CRY1 in mammals will affect circadian rhythm and the relation of such a decrease with apoptosis. Here, we discovered a molecule (M47) that destabilizes Cryptochrome 1 (CRY1) both in vitro and in vivo. The M47 selectively enhanced the degradation rate of CRY1 by increasing its ubiquitination and resulted in increasing the circadian period length of U2OS Bmal1-dLuc cells. In addition, subcellular fractionation studies from mice liver indicated that M47 increased degradation of the CRY1 in the nucleus. Furthermore, M47-mediated CRY1 reduction enhanced oxaliplatin-induced apoptosis in Ras-transformed p53 null fibroblast cells. Systemic repetitive administration of M47 increased the median lifespan of p53-/- mice by ~25%. Collectively our data suggest that M47 is a promising molecule to treat forms of cancer depending on the p53 mutation.

Escitalopram co-prescription in anastrozole-treated breast cancer patients.

OBJECTIVE: The purpose of the study was to evaluate the impact of escitalopram co-prescription on plasma anastrozole levels in post-menopausal breast cancer patients. METHODS: A total of 24 post-menopausal operated breast cancer patients co-prescribed with escitalopram and anastrozole were included. Blood samples were collected, before and 1-month after the onset of escitalopram to analyze plasma anastrozole and estradiol levels. RESULTS: No significant difference was noted in basal plasma anastrozole levels with respect to age, body mass index (BMI), tumor stage, previous antineoplastic treatments, concomitant medications, and serum estradiol levels. Overall, 17 patients completed the 1-month escitalopram treatment, while 7 patients discontinued escitalopram within the 1st week of the treatment. Basal anastrozole levels of 24 patients were 26.1±2.4 ng/mL. Among 17 patients who continued 1-month escitalopram treatment was associated with significant increase in plasma anastrozole levels (24.5±2.3 ng/mL to 32.2±3.2 ng/mL, p<0.05). Notably, 1-month escitalopram use was associated with significant increase in plasma anastrozole levels only in the subgroup of obese (BMI >29 kg/m2) patients (23.1±2.8 to 35.9±4.7 ng/mL, p<0.01), while no such interaction was noted among non-obese patients. The estradiol levels of the patients were below ≤10 pg/mL in 75% of patients and no change occurred after escitalopram administration. CONCLUSION: Escitalopram co-prescription resulted in significant increase in plasma anastrozole levels without affecting the serum estradiol levels. Our findings emphasize the need for close monitoring in case of concomitant use of anastrozole and escitalopram, especially in obese patients and the potential role of therapeutic drug monitoring.

Food effect on bioavailability of modified-release trimetazidine tablets.

This study aimed to investigate a food effect on the bio-availability of modified-release (MR) trimetazidine tablets in 36 healthy volunteers. Trimetazidine, an anti-ischemic drug, protects the myocardial cell from the harmful effects of ischemia. The authors investigated the effect of being under a fasting or fed state at the time of drug intake on the bioavailability of trimetazidine 35-mg MR tablets in a randomized, open-label, crossover, 2-arm, 4-period, 2-sequence bioequivalence study design with a 14-day washout period. Plasma concentration of trimetazidine was assayed in timed samples with a validated high- performance liquid chromatography/mass selective detector that had a lower limit of quantification of 2.5 ng/mL. Test and reference formulations gave a mean trimetazidine C(max) of 63.26 ng/mL and 69.18 ng/mL for the fasting state and 64.19 ng/mL and 63.11 ng/mL for the fed state, respectively. The AUC(0-tlast) mean of trimetazidine was 726.31 ng·h/mL and 733.01 ng·h/mL for the fasting state and 706.40 ng·h/mL and 691.40 ng·h/mL for the fed state for test/reference formulations. There were no significant differences in pharmacokinetic parameters between the 2 formulations and the fasting/fed states. The authors showed that there is no food effect and no need for a 4-period study to evaluate the bioequivalence of trimetazidine MR tablets.

Analysis of the Wnt/B-catenin/TCF4 pathway using SAGE, genome-wide microarray and promoter analysis: Identification of BRI3 and HSF2 as novel targets.

The Wnt signaling pathway is involved in many differentiation events during embryonic development and can lead to tumor formation after aberrant activation of its components. beta-catenin, a cytoplasmic component, plays a major role in the transduction of canonical Wnt signaling. The aim of this study was to identify novel genes that are regulated by active beta-catenin/TCF signaling in hepatocellular carcinoma-derived Huh7 cells with high (transfected) and low beta-catenin/TCF activities. High TCF activity Huh7 cells led to earlier and larger tumor formation when xenografted into nude mice. SAGE (Serial Analysis of Gene Expression), genome-wide microarray and in silico promoter analysis were performed in parallel, to compare gene expression between low and high beta-catenin/TCF activity clones, and also those that had been rescued from the xenograft tumors. SAGE and genome-wide microarray data were compared and contrasted. BRI3 and HSF2 were identified as novel targets of Wnt/beta-catenin signaling after combined analysis and confirming experiments including qRT-PCR, ChIP, luciferase assay and lithium treatment.

Clinical study on the bioequivalence of two tablet formulations of flurbiprofen.

Flurbiprofen (CAS 5104-49-4) is a member of phenylalkanoic acid derivative group of nonsteroid anti-inflammatory drugs. It exhibits anti-inflammatory, analgesic and antipyretic activities. Two different tablets containing flurbiprofen (FLU) were investigated in 24 healthy volunteers to prove the bioequivalence between both treatments after single oral dose administrations. Fluroben 100 mg tablet and 100 mg tablet of the originator product were used as test and reference preparation respectively. The study was performed open label, randomized, two period cross-over design with 15 days wash out period. Blood samples were taken up to 24 hours for pharmacokinetic profiling. The plasma concentrations of flurbiprofen were determined with validated HPLC-UV method. Maximum plasma concentration (Cmax) of FLU 19,143.65 ng/ml and 19,164.22 ng/ml were found for test and reference formulation respectively. Areas under the plasma concentration time curve AUC(0-infinity), of 118 501.4 ng.h/ml and 111,339.8 ng.h/ml were calculated test and reference formulation respectively. Primary target parameters AUC (0-infinity) and Cmax, both of them were tested parametrically by analysis of variance (ANOVA); 90% confidence intervals were between 100.5%-111.18% for AUC(0-infinity), and 87.6%-115.0% for Cmax. All these values were within the acceptance range (80%-125%) for bioequivalence studies.

The effect of hydroxy metabolites of clarithromycin to the pharmacokinetic parameters, and determination of hydroxy metabolites ratio of clarithromycin.

Clarithromycin is a broad-spectrum macrolide antibacterial agent which is effective both in vitro and in vivo against the major pathogens responsible for respiratory tract infections. Clarithromycin's principal metabolite is 14-(R) hydroxyclarithromycin (14-OH-clarithromycin). The other metabolite, namely 14-(S) hydroxyclarithromycin is inactive. The purpose of this study was to show the hydroxylation of CLA at the 14 position to form the R and S epimers and to determine the metabolic ratio of 14ROHCLA/CLA and 14SOHCLA/CLA for understanding the metabolization. This study suggest that in healthy adults, the individual variations in therapeutic responses to clarithromycin can be assumed by taking the drug and its metabolites ratios. Clarithromycin and metabolites ratios increase during metabolization.

The effect of hydroxy metabolites of clarithromycin to the pharmacokinetic parameters, and determination of hydroxy metabolites ratio of clarithromycin.

Clarithromycin is a broad-spectrum macrolide antibacterial agent which is effective both in vitro and in vivo against the major pathogens responsible for respiratory tract infections. Clarithromycin's principal metabolite is 14-(R) hydroxyclarithromycin (14-OH-clarithromycin). The other metabolite, namely 14-(S) hydroxyclarithromycin is inactive. The purpose of this study was to show the hydroxylation of CLA at the 14 position to form the R and S epimers and to determine the metabolic ratio of 14ROHCLA/CLA and 14SOHCLA/CLA for understanding the metabolization. This study suggest that in healthy adults, the individual variations in therapeutic responses to clarithromycin can be assumed by taking the drug and its metabolites ratios. Clarithromycin and metabolites ratios increase during metabolization.

Purine nucleoside phosphorylase deficiency in a patient with spastic paraplegia and recurrent infections.

Purine nucleoside phosphorylase deficiency is a rare autosomal recessive immunodeficiency disease. The characteristic features of the disease include severe T cell immune defects with recurrent infections, a failure to thrive, and progressive neurological findings. To date, 35 cases of purine nucleosidase phosphorylase deficiency have been reported worldwide. A 2-year-old female patient was hospitalized due to recurrent infections starting from 6 months and a fever that had continued for a month. The parents were first cousins. Physical examination showed a failure to thrive, herpetic lesions around the lips, painful lesions on the tongue and the buccal mucosa, lung infection, and spastic paraparesis in the lower extremities. She had motor and mental retardation. Laboratory tests revealed lymphopenia; low CD3, CD4, and CD8 counts; normal immunoglobulin levels; low uric acid; and very low purine nucleoside phosphorylase enzyme activity (1.4 nmol/h/mg; normal range, 490-1530). DNA sequencing of the purine nucleosidase phosphorylase gene revealed a missense homozygous mutation, a G to A transition at exon 4 position 64 (349G>A transition), which led to a substitution of alanine by threonine at codon 117 (Ala117Thr). Both parents were heterozygous for the mutation. This is the second purine nucleosidase phosphorylase deficient case to have been presented and carrying this mutation worldwide. Various antibiotics, antifungal drugs, and intravenous immunoglobulin were used to treat the infections during her 3 months. This form of treatment proved to be unresponsive, resulting in her subsequent death at 26 months of age.

Investigation of serum leptin levels in professional male football players and healthy sedentary males.

OBJECTIVE: In the present study we aimed at investigating leptin levels in professional male athletes who have been exercising regularly for a long time and leptin levels in healthy sedentary males. METHODS: The study included 10 male professional football players and 17 healthy sedentary males. The relations between groups in terms of leptin levels, Max VO2 levels, blood lactic acid levels before and after exercise and effort durations were investigated. RESULTS: It was found in the study that although BMI of professional male athletes was higher than that of the healthy sedentary males, leptin levels of the former were significantly lower (p<0.01), while VO2Max levels (p<0.05) and test periods (p<0.01) were significantly higher than those in the latter. As for lactic acid levels after exercise and between groups, these were also higher in athletes, but the difference was not statistically significant (p>0.05). CONCLUSION: Leptin levels of those who exercised regularly were found lower than the levels in healthy males. Although the increase in serum leptin levels is in direct proportion with BMI in general, the major determinant of serum leptin level is the body fat rate. As regular exercising reduces body fat rate, it also reduces serum leptin levels.

The relation between serum leptin levels and max VO2 in male patients with type I diabetes and healthy sedentary males.

This study aimed at investigating leptin levels in male diabetes type I patients who were on insulin treatment and also healthy sedentary males. The study included 10 male type I diabetes patients and 17 healthy sedentary males. Leptin levels of type I diabetes patients and healthy sedentary males with body mass index (BMI) over 25 kg/m2 were evaluated separately. The relation between serum leptin, max VO2, blood lactic acid levels before and after exercise, and effort durations of participants were investigated. At the end of the tests, no difference was found between leptin levels, max VO2 values, lactic acid values before exercise, and test durations of male type I diabetes patients and healthy sedentary males (p > .05), whereas lactic acid levels after exercise were found to be lower in healthy sedentary males (p < .05). Leptin levels in the group with BMI above 25 kg/m2 were higher than those in the group with BMI below 25 kg/m2 (p < .001). It was also seen that max VO2 values and test durations were higher in the group with BMI below 25 kg/m2 (p < .05). In conclusion, leptin levels of male type I diabetes patients are close to those of healthy sedentary males. The increase in leptin levels in both groups is in proportion to the BMI of individuals.

Gene doping in sports.

Gene or cell doping is defined by the World Anti-Doping Agency (WADA) as "the non-therapeutic use of genes, genetic elements and/or cells that have the capacity to enhance athletic performance". New research in genetics and genomics will be used not only to diagnose and treat disease, but also to attempt to enhance human performance. In recent years, gene therapy has shown progress and positive results that have highlighted the potential misuse of this technology and the debate of 'gene doping'. Gene therapies developed for the treatment of diseases such as anaemia (the gene for erythropoietin), muscular dystrophy (the gene for insulin-like growth factor-1) and peripheral vascular diseases (the gene for vascular endothelial growth factor) are potential doping methods. With progress in gene technology, many other genes with this potential will be discovered. For this reason, it is important to develop timely legal regulations and to research the field of gene doping in order to develop methods of detection. To protect the health of athletes and to ensure equal competitive conditions, the International Olympic Committee, WADA and International Sports Federations have accepted performance-enhancing substances and methods as being doping, and have forbidden them. Nevertheless, the desire to win causes athletes to misuse these drugs and methods. This paper reviews the current status of gene doping and candidate performance enhancement genes, and also the use of gene therapy in sports medicine and ethics of genetic enhancement.