Transvaginal Ultrasound-Guided Core Biopsy-Experiences in a Comprehensive Cancer Centre.

In this paper, we report our experience of transvaginal ultrasound (TVUS)-guided core biopsies involving 303 patients referred to the gynaecological ultrasound unit of our national comprehensive cancer centre. Adequate histologic specimens were obtained in 299 patients (98.7%). The most common sites of biopsy sampling were the adnexa (29.7%), the vaginal stump or wall (13.5%), the uterus (11.6%) and the peritoneum (10.2%). Malignancy was confirmed in two-thirds of patients (201/303) and a primary malignancy was diagnosed in 111 of the 201 histologically verified malignant cases (55.2%). Interestingly, 23.9% (48/201) of malignant tumours were proven to have a non-gynaecological origin. Among them, gastrointestinal tumours occurred the most frequently (31/48 patients). Three abscesses were discovered following the biopsy procedure, resulting in a complication rate of 1%. In 94 (31%) patients, subsequent surgery allowed the comparison of the ultrasound-guided and surgically obtained histologic results. We found inaccuracy in 12 cases (12.8%), which is discussed in this paper in detail. Sensitivity, specificity, PPV and NPV to diagnose malignancy was 94.8%, 94.1%, 98.7% and 80.0%, respectively. This is the largest study reported to date about the efficacy and safety of TVUS-guided core biopsy in evaluating pelvic lesions giving rise to a suspicion of gynaecological cancer.

Serum estrone concentration, estrone sulfate/estrone ratio and BMI are associated with human epidermal growth factor receptor 2 and progesterone receptor status in postmenopausal primary breast cancer patients suffering invasive ductal carcinoma.

BACKGROUND: We investigated in postmenopausal women with primary breast cancer prior to surgical intervention whether, serum levels of different steroid hormones and hormonal precursors associated with tumor tissue estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) status. METHODS: We enrolled 1,042 patients suffering invasive ductal carcinoma undergoing surgical resection in the National Institute of Oncology, Hungary between 2003 and 2011. Serum parameters were measured by RIA/IRMA assays; tumor tissue ER, PR and HER2 status was assessed histologically. Patients were classified according to tumor receptor status. Case-case analysis subjects were categorized into four subgroups based on serum hormone concentrations in ER, PR and HER2 receptor-negative cases, respectively. RESULTS: Serum estrone sulfate and dehydroepiandrosterone sulfate levels correlated with each other and also with serum estrone and estradiol levels. According to case-case study the odds ratios in the highest quartile were 1.517 (p = 0.0305, Ptrend = 0.0394) for androstenedione, 1.495 (p = 0.0317, Ptrend < 0.0105) for estrone and 0.654 (p = 0.0273, Ptrend < 0.0151) for estrone sulfate/estrone ratio in PR+ vs. PR- tumors. Regarding HER2 status (HER2+ vs. HER2-), the odds ratios for estrone, estrone sulfate and estrone sulfate/estrone ratio were 0.530 (p = 0.0234, Ptrend = 0.0595), 2.438 (p = 0.0042, Ptrend < 0.0066) and 3.118 (p = 0.0001, Ptrend < 0.0001) in the highest quartile, respectively. Of note significantly increased BMI associates with PR+ and ER +/PR+ status while significantly decreased BMI was observed in HER2+ cases. CONCLUSIONS: Taken together, measurement of serum estrone and estrone sulfate concentrations prior to surgical intervention might support the individualization of regime in postmenopausal primary breast cancer patients.

Regulation of phase II biotransformation enzymes by steroid hormones.

Phase II biotransformation enzymes conjugate xenobiotics using small, organic donor molecules, such as glutathione, UDP-glucuronic acid, S-adenosyl-L-methionine, acetyl coenzyme A and amino acids (primarily glycine). These reactions generally resulted in detoxification by the loss of pharmacological activity and by quickening the elimination of xenobiotics from the body, however Bioactivation is also known to be occurred. Historically, it was placed more emphasis on research of phase I than of phase II enzymes. Nevertheless, it is well known that conjugation enzymes play an important role in drug and toxicant disposition since they can dramatically alter pharmacokinetics and therefore therapeutic efficacy and toxicity of drugs. In this context surprisingly, the exact regulation mechanism of phase II conjugation enzymes expression is not fully understood. However, available experimental data suggest that several transcriptional factors are involved in this process. In the current review, we characterize and summarize our knowledge about regulation of the most important phase II enzymes, such as UDP-glucuronosyltransferases (UGTs), sulfotransferases (SULTs), glutathione S-transferases (GSTs), arylamine N-acetyltransferases (NATs), catechol O-methyltransferase (COMT) and thiopurine S-methyltransferase (TPMT) by different steroid hormones.

Vibrational and chiroptical spectroscopic characterization of gamma-turn model cyclic tetrapeptides containing two beta-Ala residues.

The optical spectroscopic characterization of gamma-turns in solution is uncertain and their distinction from beta-turns is often difficult. This work reports systematic ECD and vibrational circular dichroism (VCD) spectroscopic studies on gamma-turn model cyclic tetrapeptides cyclo(Ala-beta-Ala-Pro-beta-Ala) (1), cyclo(Pro-beta-Ala-Pro-beta-Ala) (2) and cyclo(Ala-beta-Ala-Ala-beta-Ala) (3). Conformational analysis performed at the 6-31G(d)/B3LYP level of theory using an adequate PCM solvent model predicted one predominant conformer for 1-3, featuring two inverse gamma-turns. The ECD spectra in ACN of 1 and 2 are characterized by a negative n-->pi* band near 230 nm and a positive pi-->pi* band below 200 nm with a long wavelength shoulder. The ECD spectra in TFE of 1-3 show similar spectra with blue-shifted bands. The VCD spectra in ACN-d(3) of 1 and 2 show a +/-/+/- amide I sign pattern resulting from four uncoupled vibrations in the case of 1 and a sequence of two positive couplets in the case of 2. A -/+/+/- amide I VCD pattern was measured for 3 in TFE-d(2). All three peptides give a positive couplet or couplet-like feature (+/-) in the amide II region. VCD spectroscopy, in agreement with theoretical calculations revealed that low frequency amide I vibrations (at approximately 1630 cm(-1) or below) are indicative of a C(7) H-bonded inverse gamma-turns with Pro in position 2, while gamma-turns encompassing Ala absorb at higher frequency (above 1645 cm(-1)).

Drug interaction potential of 2-((3,4-dichlorophenethyl)(propyl)amino)-1-(pyridin-3-yl)ethanol (LK-935), the novel nonstatin-type cholesterol-lowering agent.

The widely prescribed lipid-lowering statins are considered to be relatively safe drugs. However, the risk of severe myopathy and drug interactions as a consequence of statin therapy provides a challenge for development of novel cholesterol-lowering agents, targeting enzymes other than HMG-CoA reductase. The novel pyridylethanol-(phenylethyl)amine derivative, (2-((3,4-dichlorophenethyl)(propyl)-amino)-1-(pyridin-3-yl)ethanol (LK-935), blocking lanosterol 14alpha-demethylase, was demonstrated to efficiently reduce cholesterol biosynthesis. The drug interaction potential of LK-935 was investigated and compared with that of atorvastatin and rosuvastatin in primary human hepatocytes. Clear evidence was provided for the induction of CYP3A4 by LK-935. LK-935 was proved to be a potent human pregnane X receptor (hPXR) activator as a prerequisite for the transcriptional activation of CYP3A4 gene; however, the rapid metabolism of LK-935 in primary hepatocytes prevented maximal CYP3A4 induction. Therefore, the induction of CYP3A4 by LK-935 may be prone to mild or negligible drug interactions. However, because CYP3A4 and also CYP2C9 play a significant role in LK-935 metabolism, the inhibition of these cytochromes P450 by coadministered drugs may lead to some increase in the LK-935 concentration required for the potent induction of CYP3A4. Rosuvastatin was found to increase human constitutive androstane receptor (hCAR)-mediated transcription of CYP3A4, CYP2C9, and CYP2B6 genes, predicting the consequent potential for drug interactions with several coadministered drugs. Activation of hCAR and hPXR by atorvastatin and the subsequent induction of not only CYP2B6 and CYP3A4 but also of CYP2C9 present an additional target by which atorvastatin, a widely used cholesterol-lowering drug, can modify the kinetics of numerous drugs.

Investigation of the CYP2C9 induction profile in human hepatocytes by combining experimental and modelling approaches.

The goal of the present review is to characterise the induction profile of CYP2C9, a member of the cytochrome P450 superfamily. Since the mechanism of CYP2C9 induction is fairly complex, with parallel processes triggered by various inducers, an evaluation of the experimental results is often a great challenge. At least three nuclear receptors, the glucocrticoid receptor (GR), the pregnane X receptor (PXR) and the constitutive androstane receptor (CAR), are known to mediate the CYP2C9 gene induction in man. However, mathematical modelling and simulation can provide an appropriate tool for the interpretation of CYP2C9 regulatory mechanisms. As an example, we present modelling and simulation approaches of the CYP2C9 gene expression in human hepatocytes treated with well-known CYP2C9 inducers: the steroid hormone precursor dehydroepiandrosterone (DHEA) and the synthetic glucocorticoid dexamethasone (DXM). The results of the analysis suggest that in addition to the potent function of GR and the further involvement of PXR and CAR activated by DXM or DHEA, an additional factor might play a role in CYP2C9 regulation by DHEA. The novel potential candidate for DHEA action in CYP2C9 induction is likely to be the estrogen receptor. Additionally, the balance of DHEA sulphation-desulphation processes should also be considered in any description of DHEA-induced CYP2C9 profiles.

[Crosstalk between cholesterol homeostasis and drug metabolism]. / A koleszterinhomeosztázis és a gyógyszer-metabolizmus kapcsolata.

Nowadays cardiovascular diseases are among the major causes of mortality in the developed world. High cholesterol level in blood and atherosclerosis play the main role in progression of cardiovascular diseases. Reducing serum cholesterol level has been shown to avoid deleterious effects, whereas in serious diseases it improves the outcome. A widely used, statin-based therapy induces a decrease in de novo cholesterol biosynthesis in the liver. Another possibility for lowering serum cholesterol level is to block the uptake of dietary cholesterol from intestine (e.g. ezetimibe). Coadministration of statins and cholesterol uptake inhibitors provides an efficient therapeutical strategy. Several therapeutic agents, reducing serum cholesterol level, are able to regulate the expression of not only cholesterogenic enzymes, but of the major drug metabolizing enzymes, cytochromes P450. The crosstalk between cholesterol homeostasis and drug metabolism is mediated by nuclear receptors, activating target genes in response of endogenous and exogenous ligands. Better understanding of the crosstalk between cholesterol homeostasis and drug metabolism is essential for developing an adequate strategy in therapy and in novel drug development.

Drug-induced liver graft toxicity caused by cytochrome P450 poor metabolism.

UNLABELLED: What is already known about this subject. The activity of drug-metabolizing enzymes, primarily cytochrome P450 enzymes, can determine a patient's response to a drug. Therapeutic failure or drug toxicity in the postoperative period after liver transplantation is influenced by the drug metabolizing capacity of the graft. Dose adjustment or selection of an alternative drug, which is not a substrate for the polymorphic enzyme may prevent the development of side-effects in recipients of poor metabolizer liver grafts. What this study adds. A validated analytical system with metabolomic tools has been developed to estimate the drug-metabolizing capacity of transplanted liver, which allows the prediction of potential poor metabolizer phenotypes of donors and facilitates the improvement of individual recipient therapy. In the test of drug-metabolizing status, one of the liver grafts was found to be a CYP2C9 poor metabolizer, while the other was a CYP2C19 poor metabolizer. Rationalization of the medication resulted in the recovery of both the grafts and the recipients within 1 week. AIMS: The drug-metabolizing capacity of transplanted liver highly influences drug efficacy or toxicity, particularly in the early postoperative period. The aim of our study was to predict therapeutic failures or severe adverse drug reactions by phenotyping for cytochrome P450 (P450) polymorphism resulting in reduced or no activity of the key drug-metabolizing enzymes. METHODS: A validated analytical system with metabolomic tools has been developed for estimation of the drug-metabolizing capacity of transplanted liver, which allows the prediction of potential poor metabolizer phenotypes of donors and facilitates improvement of the individual recipient therapy. RESULTS: Of the 109 liver donors in Hungary, the frequency of poor metabolizers was found to be 0.92%, 5.5% and 8.3% for CYP2C9, CYP2C19 and CYP2D6, respectively. In the present study, two liver grafts transplanted in paediatric recipients were reported to be poor metabolizer phenotypes. The liver grafts presented normal function in the early postoperative days; 2 weeks after transplantation, however, increasing liver enzymes were detected. Histological investigation of a liver biopsy suggested drug toxicity. The test of drug metabolizing status showed one of the liver grafts to be a CYP2C9 poor metabolizer, and the other was found to be a CYP2C19 poor metabolizer. Rationalization of the medication resulted in the recovery of both the grafts and the recipients within 1 week. CONCLUSIONS: Prospective investigation of the P450 status may lead to the optimization of drug choice and/or dose for a more effective therapy, avoid serious adverse effects, and decrease medical costs. Phenotyping donor livers and tailored medication can contribute to the improvement of graft and recipient survival.

Limited applicability of 7-methoxy-4-trifluoromethylcoumarin as a CYP2C9-selective substrate.

Fluorometric substrates selective for various cytochrome P450 isoforms (P450s) have great advantages in in vitro enzyme inhibition and induction studies because they are highly sensitive and suitable for rapid screening. 7-Methoxy-4-trifluoromethylcoumarin (MFC) has been reported as a CYP2C9-selective substrate. The present study investigated the relative catalytic selectivity of several human P450s in the O-demethylation of MFC and the applicability of MFC as a probe substrate for CYP2C9. The CYP2C9-selectivity in liver microsomes was not supported by the correlation analysis within a series of microsomes from individual donors or by studies using chemical inhibitors. MFC O-demethylation of microsomes did not correlate with tolbutamide 4-hydroxylation, the classical CYP2C9-marker activity, suggesting the possible participation of some of the other P450s. Results of inhibition studies using model P450 inhibitors also brought the CYP2C9-selectivity of MFC O-demethylation into question. In microsomes containing cDNA-expressed individual P450s, CYP2B6 and CYP2E1 seemed to be the most active in the O-demethylation of MFC. Our results support the participation of several P450 enzymes (CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2E1 and CYP3A4) in MFC O-dealkylation. Therefore, MFC cannot be considered a suitable probe substrate in models that express several P450s, such as liver microsomes or primary hepatocytes. Moreover, MFC is a more potent fluorogenic substrate for CYP2B6 and CYP2E1 than for CYP2C9 in microsomes containing cDNA-expressed P450s.

Dehydroepiandrosterone induces human CYP2B6 through the constitutive androstane receptor.

Dehydroepiandrosterone (DHEA), the major precursor of androgens and estrogens, has several beneficial effects on the immune system, on memory function, and in modulating the effects of diabetes, obesity, and chemical carcinogenesis. Treatment of rats with DHEA influences expression of cytochrome P450 (P450) genes, including peroxisome proliferator-activated receptor alpha (PPAR alpha)- and pregnane X receptor (PXR)-mediated induction of CYP4As and CYP3A23, and suppression of CYP2C11. DHEA treatment elevated the expression and activities of CYP3A4, CYP2C9, CYP2C19, and CYP2B6 in primary cultures of human hepatocytes. Induction of CYP3A4 in human hepatocytes was consistent with studies in rats, but induction of CYP2Cs was unexpected. The role of PXR in this response was studied in transient transfection assays. DHEA activated hPXR in a concentration-dependent manner. Because CYP2B6 induction by DHEA in human hepatocytes might involve either PXR or constitutive androstane receptor (CAR) activation, we performed experiments in primary hepatocytes from CAR knockout mice and observed that CAR was required for maximal induction of Cyp2b10 by DHEA. Furthermore, CAR-mediated Cyp2b10 induction by DHEA was inhibited by the inverse agonist of CAR, androstanol (5 alpha-androstan-3 alpha-ol). Further evidence for CAR activation was provided by cytoplasmic/nuclear transfer of CAR upon DHEA treatment. Elucidation of CAR activation and subsequent induction of CYP2B6 by DHEA presented an additional mechanism by which the sterol can modify the expression of P450s. The effect of DHEA on the activation of the xenosensors PPAR alpha, PXR, and CAR, and the consequent potential for adverse drug/toxicant interactions should be considered in humans treated with this nutriceutical agent.

Biotransformation of deramciclane in primary hepatocytes of rat, mouse, rabbit, dog, and human.

The metabolic fate of deramciclane [(1R,2S,4R)-(-)-2-phenyl-2-(2'-dimethylamino-ethoxy)-1,7,7-trimethyl-bicyclo[2.2.1]heptane], a new anxiolytic drug candidate, has been determined in rat, mouse, rabbit, dog, and human hepatocytes. Rat and rabbit cells were the most active, whereas the rate of metabolism was quite slow in human hepatocytes. During biotransformation, deramciclane underwent side chain modification and oxidation at several positions of the molecule. The side chain modification led to the formation of N-desmethyl deramciclane and phenylborneol. The oxidation of deramciclane resulted in several hydroxy-, carboxy-, and N-oxide derivatives. The hydroxylation took place at primary or secondary carbons of the camphor ring as well as at the side chain; furthermore, dihydroxylated derivatives were also found. The side chain-modified metabolites were also oxidized to hydroxy- or carboxy-derivatives. Conjugation of phase I metabolites, as a route of elimination, was also observed in rat, rabbit, and dog hepatocytes. Although there were some species differences in biotransformation of deramciclane, it was concluded that phase I metabolism in human liver cells seemed to be similar to the metabolism in the hepatocytes isolated from rat. With careful approach, the rat model may be considered to be predictive for human metabolism of deramciclane.

Role of CYP2E1 in deramciclane metabolism.

The aim of our study was to identify the form(s) of cytochrome P450 responsible for the metabolism of deramciclane, a new anxiolytic drug candidate. The main routes of biotransformation in hepatic microsomes were side chain modification (N-demethylation or total side chain cleavage) and hydroxylation at several points of the molecule. Although several cytochrome P450 forms were involved in the metabolism, the role of CYP2E1 should be emphasized, since it catalyzed almost all steps. Production of deramciclane metabolites was significantly inhibited by diethyl-dithiocarbamate and was elevated in liver microsomes of isoniazid-treated rats. Furthermore, cDNA-expressed rat CYP2E1 generated the metabolites formed by side chain modification and hydroxylation. Neither deramciclane nor its primary metabolite, N-desmethyl deramciclane were able to influence directly the activity of CYP2E1. However, during the biotransformation, one or more metabolites must have been formed which were potent inhibitors of CYP2E1.

The effect of synthetic glucocorticoid, dexamethasone on CYP1A1 inducibility in adult rat and human hepatocytes.

Glucocorticoids act synergistically with polycyclic aromatic hydrocarbons in increasing mRNA and protein levels of CYP1A1 in rat liver. The action of dexamethasone to modify CYP1A1 expression has been investigated in adult human hepatocytes. The effect of dexamethasone on the induction of CYP1A1 by 3-methylcholanthrene is different in rat and human liver cells. Dexamethasone potentiates the induction of CYP1A1 about 3- to 4-fold in rat cells. In human hepatocytes, it reduces CYP1A1 induction by 50-60% at enzyme protein level, while it does not have an effect on CYP1A1 mRNA amount.