Delayed diagnosis of coeliac disease increases cancer risk

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Tamoxifen in treatment of hepatocellular carcinoma: a randomised controlled trial

Background Results from small randomised trials on tamoxifen in the treatment of hepatocellular carcinoma (HCC) are conflicting, We studied whether the addition of tamoxifen to best supportive care prolongs survival of patients with HCC. Methods Patients with any stage of HCC were eligible, irrespective of locoregional treatment. Randomisation was centralised, with a minimisation procedure accounting for centre, evidence of disease, and time from diagnosis. Patients were randomly allocated best supportive care alone or in addition to tamoxifen, Tamoxifen was given orally, 40 mg per day, from randomisation until death. Results 496 patients from 30 institutions were randomly allocated treatment from January, 1995, to January, 1997. Information was available for 477 patients. By Sept 15, 1997, 119 (50%) of 240 and 130 (55%) of 237 patients had died in the control and tamoxifen arms, respectively. Median survival was 16 months and 15 months (p=0.54), respectively, No differences were found within subgroups defined by prognostic variables. Relative hazard of death for patients receiving tamoxifen was 1.07 (95% CI 0.83-1.39). Interpretation Our findings show that tamoxifen is not effective in prolonging survival of patients with HCC

Regulation of bovine CYP3A28 promoter by pregnane X receptor (PXR) and constitutive androstane receptor (CAR).

INTRODUCTION The regulation of cytochrome P450 3A (CYP3A) has been extensively studied in human liver. In cattle, some information on the expression and modulation of CYP3As are now available, but molecular mechanisms involved in basal and xenobiotic- mediated gene regulation are still unknown. In this study, selected transcription factor binding sites were studied for their role in the basal and nuclear receptor-driven trans-activation of CYP3A28 promoter. MATERIALS AND METHODS Putative binding sites for transcription factors were predicted within the CYP3A28 promoter region (~10 kbp) through specific software. Fragments of the CYP3A28 promoter were amplified from bovine liver gDNA and cloned into pGL4.10 vector. Bovine nuclear receptor (bPXR, bCAR and bRXRa) fulllength cDNAs were amplified and inserted into pCI-neo expression vector. Deletion of ER6, HNF-1 and HNF-4 cis-elements in the proximal promoter (PP) construct, and mutagenesis of both ER6 and HNF-4 binding elements within the PP region, and of DR5 within the Fragment 3 (F3) were performed through inverse PCR and according to Fang et al. (2012), respectively. To identify the most important regions for gene activation, cotransfection studies in HepG2 cells were performed using all the promoter constructs, bPXR and its agonist SR12813, or constitutively active bCAR. To assay the role of specific binding sites on CYP3A28 regulation, C3A cells were then co-transfected with deleted or mutated plasmids, bPXR and SR12813 or rifampicin (RIF), and bCAR. Human 3A4-XREM- luc and PBREM-tk-luc were used as positive controls. RESULTS AND CONCLUSIONS CYP3A28 promoter screening revealed PP and PP-F3 as the most important elements for bPXR and bCAR activation. 3A4-XREMluc, the PP and PP-F3 constructs were significantly activated by bPXR and SR12813 (P < 0.05), but not by RIF. Deletion of ER6, HNF-1 and HNF-4 lead to a significant decrease of PP activation by bPXR and SR12813 (P < 0.01), while the mutated single elements ER6, HNF-4 and DR5 did not. Conversely, only PP-F3 was significantly activated by bCAR (P < 0.05). Thus, CYP3A28 PP contains interaction sites for bPXR trans-activation. ER6 and HNF-1 elements seem to contribute to SR12813 response. F3 fragment is involved in bPXR- and bCAR-mediated CYP3A28 regulation, but additional binding elements, apart from DR5, need to be investigated. ACKNOWLEDGEMENTS Project supported by University of Padova (CPDA109434, 60A08-4783/14)

Understanding the mechanism of cattle CYP3A: recent advances and remaining problems

Cytochrome P450 3A (CYP3A) has been extensively studied in humans, but its characterization in cattle is still incomplete. Comparatively, bovine CYP3A is a minor contributor to drug metabolism, despite its constitutive expression in liver, gut and other tissues. Several factors influence cattle CYP3A expression and function: species, breed, age, gender, induction and inhibition phenomena that may lead to drug-drug interactions (DDIs). Marker substrates are testosterone, ethylmorphine and macrolides, but luciferin IPA and midazolam have been recently used, too. Three CYP3A isoforms have been identified in cattle liver (CYP3A28, 3A38 and 3A48) but their tissue distribution and substrate specificities are currently unclear. In humans, CYP3A polymorphisms (SNPs) are sources of individual differences in disease risk and treatment response. Sequencing of CYP3A coding regions in Piedmontese cattle revealed 12 SNPs, whose relevance is now under investigation. At present, no data about intron and splicing variant SNPs are available, but some CYP3A28 SNPs have been associated with productive traits. Species-differences exist in CYP3A regulation, and similar data have been reported for cattle. Following sequencing and in silico analysis of CYP3A28 promoter (~10000 bp 5\ub4 upstream), clusters relevant for CYP3A28 proximal promoter transactivation, resembling those known for CYP3A4, have been identified. Functional studies are currently ongoing. Additional studies on cattle CYP3A are needed to: i) set up and validate in vitro tools useful for in-depth molecular studies; ii) clarify expression and regulation phenomena; iii) find out likely relationships with drug transporters; iv) avoid harmful DDIs. Grants: 2009ZE5HJP (MIUR); CPDA109434, 60A08-7517/13 (University of Padua)