Search

Background: The role of hormones in focal nodular hyperplasia (FNH) has been investigated with conflicting results., Objective: The aim of this study was to evaluate oestrogen and progesterone receptor immunohistochemical expression in FNH and surrounding normal liver (control material)., Methods: Biopsy materials from FNH and control tissue were investigated using an immunostainer. Receptor expression was graded as the proportion score (percentage of nuclear staining) and oestrogen receptor intensity score., Results: Study material included tissue from 11 resected FNH lesions and two core biopsies in 13 patients (two male). Twelve samples showed oestrogen receptor expression. The percentage of nuclear oestrogen receptor staining was <33% in eight FNH biopsies, 34-66% in two FNH biopsies, and >67% in both core biopsies. The better staining in core biopsies relates to limitations of the staining technique imposed by the fibrous nature of larger resected FNH. Control samples from surrounding tissue were available for nine of the resected specimens and all showed oestrogen receptor expression. Progesterone receptor expression was negligible in FNH and control samples., Conclusions: By contrast with previous studies, the majority of FNH and surrounding liver in this cohort demonstrated oestrogen receptor nuclear staining. The implications of this for continued oral contraceptive use in women of reproductive age with FNH remain uncertain given the lack of consistent reported growth response to oestrogen stimulation or withdrawal., (© 2015 International Hepato-Pancreato-Biliary Association.)

Background: Pancreatic cancer (PC) has the worst survival of all periampullary cancers. This may relate to histopathological differences between pancreatic cancers and other periampullary cancers. Our aim was to examine the distribution and histopathologic features of pancreatic, ampullary, biliary and duodenal cancers resected with a pancreaticoduodenectomy (PD) and to examine local trends of periampullary cancers resected with a PD., Methods: A retrospective review of PD between January 2000 and December 2012 at a public metropolitan database was performed. The institutional ethics committee approved this study., Results: There were 142 PDs during the study period, of which 70 cases were pre-2010 and 72 post-2010, corresponding to a recent increase in the number of cases. Of the 142 cases, 116 were for periampullary cancers. There were also proportionately more PD for PC (26/60, 43% pre-2010 vs 39/56, 70% post-2010, P = 0.005). There were 65/116 (56%) pancreatic, 29/116 (25%), ampullary, 17/116 (15%) biliary and 5/116 (4%) duodenal cancers. Nodal involvement occurred more frequently in PC (78%) compared to ampullary (59%), biliary (47%) and duodenal cancers (20%), P = 0.002. Perineural invasion was also more frequent in PC (74%) compared to ampullary (34%), biliary (59%) and duodenal cancers (20%), P = 0.002. Microvascular invasion was seen in 57% pancreatic, 38% ampullary, 41% biliary and 20% duodenal cancers, P = 0.222. Overall, clear margins (R0) were achieved in fewer PC 41/65 (63%) compared to ampullary 27/29 (93%; P = 0.003) and biliary cancers 16/17 (94%; P = 0.014)., Conclusions: This study highlights that almost half of PD was performed for cancers other than PC, mainly ampullary and biliary cancers. The volume of PD has increased in recent years with an increased proportion being for PC. PC had higher rates of nodal and perineural invasion compared to ampullary, biliary and duodenal cancers.

Mice are resistant to aflatoxin hepatotoxicity, primarily due to high expression of glutathione S-transferases (GSTs), and in particular the GSTA3 subunit. Nuclear factor erythroid 2 related factor 2 (Nrf2) signaling, which controls a broad-based cytoprotective response, was activated either genetically or pharmacologically in an attempt to rescue GSTA3 knockout mice from aflatoxin genotoxicity. Genetic activation of Nrf2 signaling was attained in a GSTA3: hepatocyte-specific Keap1 double knockout (DKO) mouse whereas pharmacologic activation of Nrf2 was achieved through pretreatment of mice with the triterpenoid 1-[2-cyano-3-,12-dioxoleana-1,9(11)-dien-28-oyl] imidazole (CDDO-Im) prior to aflatoxin B1 exposure. Following oral treatment with aflatoxin, urine was collected from mice for 24 h and hepatic and urinary aflatoxin metabolites then quantified using isotope dilution-mass spectrometry. Although Nrf2 was successfully activated genetically and pharmacologically, neither means affected the response of GSTA3 knockout mice to chemical insult with aflatoxin. Hepatic aflatoxin B1-N(7)-guanine levels were elevated 120-fold in GSTA3 knockout mice compared with wild-type and levels were not attenuated by the interventions. This lack of effect was mirrored in the urinary excretion of aflatoxin B1-N(7)-guanine. By contrast, urinary excretion of aflatoxin B1-N-acetylcysteine was >200-fold higher in wild-type mice compared with the single GSTA3 knockout or DKO mouse. The inability to rescue GSTA3 knockout mice from aflatoxin genotoxicity through the Nrf2 transcriptional program indicates that Gsta3 is unilaterally responsible for the detoxication of aflatoxin in mice.

Abscess formation of the falciform ligament is incredibly rare and perplexing when encountered for the first time. It is reported to occur in the setting of cholecystitis and cholangitis, but the pathophysiology is poorly understood.In this case report, we present a 73-year-old man with falciform ligament abscess following cholangitis from an obstructive ampullary carcinoma. The patient was referred to the Royal Adelaide Hospital from a country hospital, with progressive jaundice, anorexia and nausea. Prior to transfer, he deteriorated with cholangitis, dehydration and renal failure. On arrival, his abdomen was exquisitely tender along the course of the falciform ligament. His blood tests revealed an elevated white cell count of 14.9 x 10(3)/μl, bilirubin of 291 μmol/l and creatinine of 347 μmol/l. His CA 19-9 was markedly elevated at 35,000 kU/l. A non-contrast computed tomography (CT) demonstrated gross biliary dilatation and a collection tracking along the path of the falciform ligament to the umbilicus. The patient was commenced on intravenous antibiotics and underwent an urgent endoscopic retrograde cholangiopancreatogram (ERCP) with sphincterotomy and biliary stent drainage. Cholangiogram revealed a grossly dilated biliary tree, with abrupt transition at the ampulla, which on biopsy confirmed an obstructing ampullary carcinoma. Following ERCP, his jaundice and abdominal tenderness resolved. He was optimized over 4 weeks for an elective pancreaticoduodenectomy. At operation, we found abscess transformation of the falciform ligament. Copious amounts of pus and necrotic material was drained. Part of the round ligament was resected along the undersurface of the liver. Histology showed that there was prominent histiocytic inflammation with granular acellular eosinophilic components. The patient recovered slowly but uneventfully. A contrast CT scan undertaken 2 weeks post-operatively (approximately 7 weeks after the initial CT) revealed left portal venous thrombosis, which was likely to be a delayed discovery and was managed conservatively. We present this patient's operative images and radiographic findings, which may explain the pathophysiology behind this rare complication. We hypothesize that cholangitis, with secondary portal pyaemia and tracking via the paraumbilical veins, can cause infectious seeding of the falciform ligament, with consequent abscess formation.

The synthetic oleanolic triterpenoid 1-[2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oyl]imidazole (CDDO-Imidazolide or CDDO-Im) is an extremely potent activator of Nrf2 signaling. In cells undergoing adipogenesis, CDDO-Im prevents lipid accumulation in an Nrf2-dependent manner. However, in vivo evidence for effects of CDDO-Im on obesity is lacking. The goals of these studies were to determine if CDDO-Im can prevent high-fat diet-induced obesogenesis in the mouse, and to elucidate the molecular target of drug action. Wild-type and Nrf2-disrupted C57BL/6J female mice were dosed 3 times per week with 30 micromol/kg CDDO-Im or vehicle by oral gavage, during 95 days of access to a control diet or a high-fat diet. Body weights, organ weights, hepatic fat accumulation and gene expression were measured. Treatment with CDDO-Im effectively prevented high-fat diet-induced increases in body weight, adipose mass, and hepatic lipid accumulation in wild-type mice but not in Nrf2-disrupted mice. Wild-type mice on a high-fat diet and treated with CDDO-Im exhibited higher oxygen consumption and energy expenditure than vehicle-treated mice, while food intake was lower in CDDO-Im-treated than vehicle-treated mice. Levels of gene transcripts for fatty acid synthesis enzymes were downregulated after CDDO-Im treatment in the liver of wild-type mice. This inhibitory effect of CDDO-Im on lipogenic gene expression was significantly reduced in Nrf2-disrupted mice. The results indicate that CDDO-Im is an exceedingly potent agent for preventing obesity, and identify the Nrf2 pathway as a novel target for management of obesogenesis.

Loss of NF-E2-related factor 2 (Nrf2) signaling increases susceptibility to acute toxicity, inflammation and carcinogenesis in mice due to the inability to mount adaptive responses. In contrast, disruption of Keap1 (a cytoplasmic modifier of Nrf2 turnover) protects against these stresses in mice, although inactivating mutations in Keap1 have been identified recently in some human cancers. Global characterization of Nrf2 activation is important to exploit this pathway for chemoprevention in healthy, yet at-risk individuals and also to elucidate the consequences of hijacking the pathway in Keap1-mutant human cancers. Liver-targeted conditional Keap1-null, Albumin-Cre:Keap1((flox/-)) (CKO) mice provide a model of genetic activation of Nrf2 signaling. By coupling global gene expression analysis of CKO mice with analysis of pharmacologic activation using the synthetic oleanane triterpenoid 1-[2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oyl]imidazole (CDDO-Im), we are able to gain insight into pathways affected by Nrf2 activation. CDDO-Im is an extremely potent activator of Nrf2 signaling. CKO mice were used to identify genes modulated by genetic activation of Nrf2 signaling. The CKO response was compared with hepatic global gene expression changes in wild-type mice treated with CDDO-Im at a maximal Nrf2 activating dose. The results show that genetic and pharmacologic activation of Nrf2 signaling modulates pathways beyond detoxication and cytoprotection, with the largest cluster of genes associated with lipid metabolism. Genetic activation of Nrf2 results in much larger numbers of detoxication and lipid metabolism gene changes. Additionally, analysis of pharmacologic activation suggests that Nrf2 is the primary mediator of CDDO-Im activity, though other cell-signaling targets are also modulated following an oral dose of 30 micromol/kg.

Background: Results of liver transplantation have improved significantly over the last three decades. Hospital volume has been cited as an important outcome factor. The South Australian (SA) programme commenced in 1992 as the fourth centre in Australia., Methods: Patient and graft survival curves were computed from the Australian and New Zealand (ANZ) Liver Transplant Registry. SA results were compared with the rest of ANZ. All recipients of a cadaveric liver from the start of the SA programme in 1992 until the 100th case in 2005 were included and compared with recipients from ANZ in the same period., Results: In SA, 100 recipients received 104 grafts (average 8.0/year). In the same period 1438 recipients received 1530 grafts in ANZ (mean 23.4/year per unit). The median follow up was 6.2 years. Patient and graft survival curves were similar (P = 0.51, log-rank test.) Survival rates at 1, 5 and 10 years were 0.93, 0.83, 0.61 and 0.88, 0.80, 0.72 for SA and ANZ, respectively (P = NS). There was trend towards a higher mortality rate of recurrent hepatitis C in SA patients when compared with the rest of ANZ (4.0 vs 1.3% respectively, P = 0.06). Recipient variables, such as age and primary liver disease did not differ between SA and ANZ. For the SA patients the biliary anastomotic stricture rate was 20%, primary non-function rate was 2%, hepatic artery thrombosis 1% and portal vein thrombosis 1%., Conclusion: Good results are possible in a small-volume unit for cadaveric adult liver transplant.

Eight new side-chain allylic, benzylic, and propargylic ether analogs of the natural hormone calcitriol have been rationally designed and easily synthesized. Three of these 23-oxa ether analogs lacking the typical side-chain OH group are more antiproliferative in vitro and desirably less calcemic in vivo than the natural hormone. One of these three 23-oxa analogs has transcriptional potency almost as high as that of calcitriol, even though it binds to the human vitamin D receptor only about 1% as well as calcitriol.

Chronic inflammation has been associated with increased risk of developing cancer. The transcription factor NF-E2-related factor 2 (Nrf2) controls the expression of numerous antioxidative enzymes that have been shown to attenuate acute inflammation. The present study investigated the role of Nrf2 genotype in modulating inflammation-promoted colorectal tumorigenesis. Nrf2 wild-type (WT) and Nrf2-deficient (N0) mice were administered a single dose of azoxymethane followed by a 1-week dose of drinking water with or without 1% dextran sulfate sodium (DSS). Aberrant crypt foci were counted 3 weeks after the cessation of DSS treatment. DSS treatment significantly increased numbers of aberrant crypt foci in N0 mice, but not WT mice. The extent of inflammation over the course of DSS treatment was analyzed in both genotypes. Histological analysis of colon sections revealed that N0 mice had markedly increased inflammation and mucosal damage when compared to WT mice beginning on Day 6 of DSS treatment. Although similar levels of inflammatory and oxidative damage biomarkers were evident in colons from WT and N0 mice at the start of DSS treatment, increased colonic proinflammatory cytokine mRNA transcript levels, myeloperoxidase activity and 3-nitrotyrosine immunoreactivity were observed on Day 6 of DSS treatment in N0 mice, but not WT mice. Additionally, DSS treatment resulted in increased lipid peroxidation and loss of aconitase activity in N0 mice, but not WT mice, reflecting increased oxidative damage in colons from N0 mice. Taken together, these results clearly illustrate the role of Nrf2 in regulating an adaptive response that protects against early-phase inflammation-mediated tumorigenesis., (Copyright (c) 2007 Wiley-Liss, Inc.)

Novel fluorinated sulfone analogs of the hormone 1alpha,25-dihydroxyvitamin D(3) have been designed and synthesized in order to study the biological effects of fluorine incorporation at the terminus of the C,D-ring side chain. Although biologically active 26,27-hexafluorinated 1alpha,25-dihydroxyvitamin D(3) analogs have been synthesized previously, this investigation reports the first successful fluorinated series in which trifluoromethyl sulfone analogs present a favorable biological profile. This study shows that two new analogs featuring incorporation of a synthetically simple single trifluoromethyl sulfone group have significantly increased antiproliferative activity while causing desirably low in vivo calciuria relative to that of calcitriol. Incorporation of additional fluorines, as in a perfluorobutyl analog, results in a loss of antiproliferative activity.

A dozen 24-sulfoximine analogues of the hormone 1alpha,25-dihydroxyvitamin D(3) were prepared, differing not only at the stereogenic sulfoximine stereocenter but also at the A-ring. Although these sulfoximines were not active transcriptionally and were only very weakly antiproliferative, some of them are powerful hydroxylase enzyme inhibitors. Specifically, 24-(S)-NH phenyl sulfoximine 3a is an extremely potent CYP24 inhibitor (IC(50) = 7.4 nM) having low calcemic activity. In addition, this compound shows high selectivity toward the CYP24 enzyme in comparison to CYP27A1 (IC(50) > 1000 nM) and CYP27B (IC(50) = 554 nM).

The new 24-phenylsulfone 4a, a low-calcemic analog of the natural hormone 1alpha,25-dihydroxyvitamin D(3), is a potent (IC(50) = 28nM) and highly selective inhibitor of the human 24-hydroxylase enzyme CYP24.

The cancer chemopreventive actions of oltipraz (4-methyl-5-[2-pyrazinyl]-1,2-dithiole-3-thione) have been primarily associated with the induction of phase 2 detoxifying enzymes through transcriptional activation of the antioxidant response element (ARE) in the promoter regions of these genes. The transcription factor Nrf2 has been shown to bind to and activate AREs. Previously, we demonstrated that nrf2-deficient mice had low basal expression of phase 2 enzymes and were substantially more susceptible to benzo[a]pyrene (B[a]P)-induced neoplasia of the forestomach than wild-type. Moreover, loss of Nrf2 abrogated the chemopreventive action of oltipraz, when administered 48 h before B[a]P, an interval allowing maximal induction of many phase 2 enzymes. Oltipraz also inhibits some cytochrome P450s involved in the bioactivation of B[a]P. In the present study we observed that oltipraz had no protective effect on tumor burden in the forestomach of nrf2-deficient mice when administered 1 h before B[a]P, a timeline that selectively optimizes for possible inhibitory effects on cytochrome P450s. To evaluate the role of nrf2 genotype on B[a]P disposition, levels of B[a]P-DNA adducts were measured as tetrols released from DNA isolated from target (forestomach) and non-target tissues (liver) of wild-type and nrf2-deficient mice treated with either vehicle or oltipraz 1 or 48 h before B[a]P. Levels of B[a]P-DNA adducts in forestomach were significantly higher in nrf2-deficient compared with wild-type mice. Oltipraz treatment at 1 or 48 h before B[a]P had no protective effect on forestomach tetrol levels in nrf2-deficient mice, whereas a significant reduction was observed in wild-type mice treated with oltipraz 48 h, but not 1 h, before carcinogen. Combining all treatments and genotypes, there was a strong correlation (R(2) = 0.91) between levels of B[a]P-DNA adducts in forestomach and subsequent yield of tumors. In contrast to the results in forestomach, nrf2 genotype did not modify hepatic B[a]P-DNA adduct levels while both oltipraz treatments were protective, suggesting that Nrf2-independent mechanisms (e.g. P450 inhibition) for oltipraz can also occur in vivo in some tissues.

Gastric infection with Helicobacter pylori is a cosmopolitan problem, and is especially common in developing regions where there is also a high prevalence of gastric cancer. These infections are known to cause gastritis and peptic ulcers, and dramatically enhance the risk of gastric cancer. Eradication of this organism is an important medical goal that is complicated by the development of resistance to conventional antimicrobial agents and by the persistence of a low level reservoir of H. pylori within gastric epithelial cells. Moreover, economic and practical problems preclude widespread and intensive use of antibiotics in most developing regions. We have found that sulforaphane [(-)-1-isothiocyanato-(4R)-(methylsulfinyl)butane], an isothiocyanate abundant as its glucosinolate precursor in certain varieties of broccoli and broccoli sprouts, is a potent bacteriostatic agent against 3 reference strains and 45 clinical isolates of H. pylori [minimal inhibitory concentration (MIC) for 90% of the strains is

One of the major mechanisms of protection against carcinogenesis, mutagenesis, and other forms of toxicity mediated by carcinogens is the induction of enzymes involved in their metabolism, particularly phase 2 enzymes such as glutathione S-transferases (GSTs), UDP-glucuronosyl transferases, and quinone reductases. Animal studies indicate that induction of phase 2 enzymes is a sufficient condition for obtaining chemoprevention and can be achieved by administering any of a diverse array of naturally-occurring and synthetic chemopreventive agents. Indeed, monitoring of enzyme induction has led to the recognition or isolation of novel, potent chemopreventive agents such as 1,2-dithiole-3-thiones, terpenoids and the isothiocyanate sulforaphane. For example, oltipraz, a substituted 1,2-dithiole-3-thione originally developed as an antischistosomal agent, possesses chemopreventive activity against different classes of carcinogens targeting multiple organs. Mechanistic studies in rodent models for chemoprevention of aflatoxin B(1) (AFB(1))-induced hepatocarcinogenesis by oltipraz indicates that increased expression of phase 2 genes is of central importance, although inhibition of phase 1 activation of AFB(1) can also contribute to protection. Exposure of rodents to 1,2-dithiole-3-thiones triggers nuclear accumulation of the transcription factor Nrf2 and its enhanced binding to the "antioxidant response element" (ARE), leading to transcriptional activation of a score of genes involved in carcinogen detoxication and attenuation of oxidative stress. Nrf2-deficient mice fail to induce many of these genes in response to dithiolethiones; moreover, basal expression of these genes is typically repressed. To test the hypothesis that enzyme induction is a useful strategy for chemoprevention in humans, three key elements are necessary: a candidate agent, an at-risk population and modulatable intermediate endpoints. Towards this end, a placebo-controlled, double blind clinical trial of oltipraz was conducted in residents of Qidong, PR China who are exposed to dietary aflatoxins and who are at high risk for the development of liver cancer. Oltipraz significantly enhanced excretion of a phase 2 product, aflatoxin-mercapturic acid, a derivative of the aflatoxin-glutathione conjugate, in the urine of study participants administered 125 mg oltipraz by mouth daily. Administration of 500 mg oltipraz once a week led to a significant reduction in the excretion of the primary oxidative metabolite of AFB(1), AFM(1), when measured shortly after drug administration. While this study highlighted the general feasibility of inducing phase 2 enzymes in humans, a longer term intervention is addressing whether protective alterations in aflatoxin metabolism can be sustained for extended periods of time in this high-risk population.

Induction of phase 2 enzymes, which neutralize reactive electrophiles and act as indirect antioxidants, appears to be an effective means for achieving protection against a variety of carcinogens in animals and humans. Transcriptional control of the expression of these enzymes is mediated, at least in part, through the antioxidant response element (ARE) found in the regulatory regions of their genes. The transcription factor Nrf2, which binds to the ARE, appears to be essential for the induction of prototypical phase 2 enzymes such as glutathione S-transferases (GSTs) and NAD(P)H:quinone oxidoreductase (NQO1). Constitutive hepatic and gastric activities of GST and NQO1 were reduced by 50-80% in nrf2-deficient mice compared with wild-type mice. Moreover, the 2- to 5-fold induction of these enzymes in wild-type mice by the chemoprotective agent oltipraz, which is currently in clinical trials, was almost completely abrogated in the nrf2-deficient mice. In parallel with the enzymatic changes, nrf2-deficient mice had a significantly higher burden of gastric neoplasia after treatment with benzo[a]pyrene than did wild-type mice. Oltipraz significantly reduced multiplicity of gastric neoplasia in wild-type mice by 55%, but had no effect on tumor burden in nrf2-deficient mice. Thus, Nrf2 plays a central role in the regulation of constitutive and inducible expression of phase 2 enzymes in vivo and dramatically influences susceptibility to carcinogenesis. Moreover, the total loss of anticarcinogenic efficacy of oltipraz in the nrf2-disrupted mice highlights the prime importance of elevated phase 2 gene expression in chemoprotection by this and similar enzyme inducers.

New C,D-ring side-chain-modified sulfone 4a, with natural 1alpha, 3beta-hydroxyl groups but lacking the 25-hydroxyl group characteristic of the natural hormone 1alpha,25-dihydroxyvitamin D(3) (1), has been prepared and characterized. Novel synthetic features include: (1) chemoselective oxidation of only a primary silyl ether in a primary-secondary bis-silyl ether intermediate and (2) smooth reductive etherification without interference by a neighboring sulfonyl group. Sulfone 4a, but not its 1beta, 3alpha-diastereomer 4b, is powerfully antiproliferative and transcriptionally active in vitro but desirably noncalcemic in vivo. Although sulfone 4a, designed to resemble Leo Pharmaceutical Co.'s KH-1060 (3), is recognized by catabolic enzymes, the selective biological profile of sulfone 4a is likely not due to its metabolites that are formed in only minor amounts.

Development of vitamin D analogs (deltanoids) as chemopreventive agents requires separation of desirable antiproliferative and pro-differentiating activities from the undesirable calcemic activity also found in the hormone calcitriol (1 alpha, 25-dihydroxyvitamin D(3)). Therefore, several conceptually new deltanoids were synthesized with modifications to the 1alpha- and/or 25-hydroxyl groups, positions traditionally considered essential for stimulating biological responses. In this study, 1 beta-hydroxymethyl-3-epi-25-hydroxyvitamin D(3), a non-calcemic CH(2) homolog of the natural hormone with antiproliferative activity in vitro, was ineffective as an inhibitor of 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced induction of ornithine decarboxylase activity in mouse epidermis. However, a hybrid analog incorporating not only the calcemia-ablating 1 beta-hydroxymethyl alteration, but potentiating C,D ring 16-unsaturation and side chain 24,24-fluorination and 26, 27-homologation was found to be as effective as calcitriol. Several non-calcemic 24- or 25-t-butyl sulfones, some containing side chain fluorination but all lacking the 25-hydroxyl group, were also shown to be active in this assay. Three sulfones and the 1 beta-hydroxymethyl hybrid were evaluated as inhibitors of multistage carcinogenesis in mouse skin. Female CD-1 mice were initiated with a single dose of 7,12-dimethylbenz[a]anthracene and then promoted twice weekly for 20 weeks with TPA. Deltanoids were applied topically 30 min before TPA. Unlike calcitriol, none of the atypical deltanoids affected body weight gain in these animals. Minimal effects on urinary calcium excretion were observed following chronic treatment with these analogs. All deltanoids inhibited the incidence and multiplicity of papilloma formation, with the hybrid analog showing the greatest efficacy. With this deltanoid, tumor incidence was significantly reduced by 28% and tumor multiplicity by 63%. These results, coupled with the rich chemical diversity available in side chain sulfur-containing deltanoids, particularly when combined with A ring modifications such as 1 beta-hydroxylalkyl groups, provide important new advances in the fundamental understanding of chemical structure-biological activity relationships as well as more potent and safe vitamin D analogs for cancer chemoprevention and other medicinal uses.

Studies in animals and humans have established serum aflatoxin-albumin adducts as biomarkers of exposure to aflatoxin B1 (AFB1), a food-borne hepatocarcinogen. To assess the utility of measurements of aflatoxin-albumin adducts to predict risk of hepatocellular carcinoma (HCC), 123 male F344 rats were dosed with 20 microg of AFB1 daily for 5 weeks after randomization into three groups: no intervention; delayed-transient (500 ppm of oltipraz, weeks 2 and 3 relative to AFB1); or persistent (500 ppm oltipraz, weeks -1 to 5). Serial blood samples were collected from each animal at weekly intervals throughout aflatoxin B1 exposure and assayed for levels of aflatoxin-albumin by radioimmune assay. Area under the curve (AUC) values for aflatoxin-albumin adducts decreased 20 and 39% in the delayed-transient and persistent oltipraz intervention groups, respectively, as compared to no intervention. Similarly, the total incidence of HCC dropped from 83 to 60% (P = 0.03) and 48% (P < 0.01) in these groups. Tumor multiplicity was also reduced in the two oltipraz intervention groups, whereas time to HCC was increased. Mononuclear cell leukemia, a common neoplasm in F344 rats, was seen in 39% of the control animals, whereas the two oltipraz interventions reduced incidence to 18% (P = 0.05) and 13% (P = 0.01), respectively. Overall, a significant association was seen between biomarker AUC and risk of HCC (P = 0.01). However, when the predictive value of aflatoxin-albumin adducts was assessed within treatment groups, there was no association between AUC and risk of HCC (P = 0.56). Thus, aflatoxin-albumin adducts can be useful for monitoring population-based changes induced by interventions, such as in chemoprevention trials, but have limited utility in identifying individuals destined to develop HCC. As a consequence, the use of this biomarker in quantitative risk assessment should be pursued cautiously.

The Australian National Liver Transplantation Unit carried out 251 multiple organ-retrieval procedures between January 1986 and December 1994: 53% were in the Sydney metropolitan area, 22% in country New South Wales and the Australian Capital Territory, 21% interstate and 4% in New Zealand; mean total procurement times were, respectively, 4 hours 57 minutes, 6 hours 58 minutes, 9 hours 20 minutes and 12 hours 20 minutes. Donor operation times steadily decreased over the nine years. One-year patient and graft survival rates were 77.1% and 70.3%, respectively; primary non-function occurred in only one case. The importance of coordination of donor organ procurement and efficient organ retrieval to the success of transplantation programs is discussed.

The validation process for biomarkers to be used for monitoring the efficacy of preventive interventions in humans includes assessments of whether levels of the biomarker can be modulated in experimental models. From this perspective, the influence of two intervention protocols with the chemopreventive agent oltipraz on rates of formation and disappearance of aflatoxin-albumin adducts has been evaluated in rats chronically exposed to aflatoxin B1. Male F344 rats were treated daily with 20 micrograms aflatoxin B1, p.o. for 35 days. The first strategy employed a standard, long-term intervention in which basal AIN-76A diet was supplemented with 0.05% oltipraz beginning one week before AFB1 treatment and continuing throughout the period of carcinogen exposure. In this setting, treatment with oltipraz reduced the rate of formation of aflatoxin-albumin adducts such that steady-state levels were lowered by > 50% from control values of 400 pmol aflatoxin adducts/mg albumin. The time-course for reaching respective steady-state levels was unchanged, with or without oltipraz intervention. The second intervention strategy utilized a delayed, transient protocol in which oltipraz was fed for 2 weeks beginning 1 week after AFB1 dosing began and ending 2 weeks before AFB1 dosing was completed. This second strategy, which models clinical interventions in chronically exposed individuals, produced a steady decline in aflatoxin-albumin adduct levels that approached a 50% reduction by the end of the AFB1 exposure period. Development of smooth curve functions allowed for the estimation of the ratio of effects between the non-intervention and intervention groups as well as the simultaneous 90% confidence intervals for the aflatoxin-albumin adduct levels. These analyses indicated that long-term intervention with oltipraz produced a statistically significant reduction in levels of the aflatoxin-albumin biomarker at all times throughout aflatoxin exposure. By contrast, a statistically significant decrease in biomarker levels was not seen in the delayed, transient intervention protocol until the ninth day of the intervention. However, once achieved, significant differences from the control group were maintained for the remainder of the aflatoxin exposure period. These changes in aflatoxin biomarker levels are consistent with the cancer chemopreventive outcomes of these intervention protocols in rats. Collectively, these results support the utility of measuring this biomarker as a means for assessing the efficacy or chemopreventive interventions in individuals at high risk for aflatoxin exposure and development of hepatocellular carcinoma.

Metabolism of the skin tumor promoter butylated hydroxy-toluene hydroperoxide (2,6-di-tert-butyl-4-hydroperoxyl-4-methyl-2,5-cyclohexadienone; BHTOOH) to reactive intermediates is required for tumor promotion by this compound. In particular, an electrophilic quinone methide is known to mediate both in vivo tumor promotion as well as in vitro cytotoxicity by BHTOOH. In the present study, the role of this reactive intermediate in the induction of ornithine decarboxylase (ODC), a gene strongly associated with tumor promotion, was investigated in cultured keratinocytes. BHTOOH stimulates a time-dependent increase in ODC enzyme activity, paralleled by ODC mRNA induction, suggesting transcriptional regulation of ODC by BHTOOH. Depletion of intracellular glutathione caused a 5-fold potentiation of keratinocyte sensitivity to BHTOOH. Concordantly, ODC induction by BHTOOH could be completely inhibited by soluble thiol compounds. These results suggest that ODC induction is mediated by a thiol-reactive metabolite of BHTOOH. The iron-specific chelator desferal blocked ODC induction by BHTOOH, indicating that formation of this intermediate is iron-dependent. Substitution of the 4-methyl group of BHTOOH with alkyl groups of incrementally larger size is known to reduce accordingly quinone methide production; comparative study of these BHTOOH analogs demonstrated a corresponding loss of potency for ODC induction, indicating that BHT-quinone methide mediates the in vitro induction of ODC by BHTOOH. Finally, kinase inhibitor studies suggested a role for protein kinase C in the induction of ODC by BHTOOH. Taken together, these results provide insight into the cellular mechanisms through which the reactive electrophile BHT-quinone methide can mediate alterations in gene expression, such as occur in tumor promotion in vivo.

Objective: To determine factors important to clinical success in micromanipulation-assisted in vitro fertilization (IVF)., Design: Procedures invoked in two separate series of micromanipulation-assisted IVF cycles, one unsuccessful (series I) and the other successful (series II), were compared in an effort to identify changes that led to clinical success., Setting: University-based IVF clinic., Patients: In both IVF series involving micromanipulation, patients consisted of infertile couples who fit any of five categories of male-factor related infertility. The female patients underwent controlled hyperstimulation for oocyte retrieval and the oocytes were inseminated normally or were subjected either to partial zona dissection or subzonal sperm insertion to assist fertilization. Results in all groups were compared between the two patient series., Results: In the successful series II, a noticeable improvement in fertilization rate and embryo quality was observed compared with series I. A significant increase in the percentage of patients reaching embryo transfer, the pregnancy rate per transfer, and the pregnancy rate per retrieval were noted in series II; a 25% ongoing pregnancy rate per retrieval was observed overall in this successful group, with "ongoing" defined as manifestation of at least a fetal sac on ultrasound with no detectable problems. Patients with a mixed transfer of embryos derived from manipulated and normally inseminated oocytes had a 75% rate of pregnancy per transfer in series II. Differences between the two series could not be attributed to patient selection or biases in selection of oocytes relegated to micromanipulation. However, oocyte handling, micromanipulation, and culture protocols differed significantly between the two series in that temperature and pH of oocytes was better controlled, and micromanipulation time was minimized in series II., Conclusion: Success in micromanipulation depends on maintenance of the oocyte in a stable and supportive environment throughout the micromanipulation procedure. It is also important to minimize trauma to the eggs by performing micromanipulation rapidly and with minimal distortion of the egg. Patients with a poor fertilization rate in standard IVF may experience a substantial increase in the likelihood of pregnancy when micromanipulation-assisted fertilization is performed on some eggs.

Benzoyl peroxide (BzPO) is both a tumor promoter and progressor in mouse skin; however, BzPO is neither an initiator nor a complete carcinogen in this tissue. Although not mutagenic, BzPO has been observed to produce strand breaks in DNA of exposed cells. These actions are presumed to be mediated by free-radical derivatives of BzPO. Previous studies suggested that the metabolism of BzPO in keratinocytes proceeds via the initial cleavage of the peroxide bond, yielding benzoyloxy radicals which, in turn, can either fragment to form phenyl radicals and carbon dioxide or abstract H atoms from biomolecules to yield benzoic acid. Benzoic acid is the major stable metabolite of BzPO produced by keratinocytes. In the present study we have investigated the role of BzPO and its metabolites in the generation of strand scissions in a cell-free system using phi X-174 plasmid DNA. In this system BzPO produced DNA damage that was dose-dependent over a concentration range of 0.1-1 mM and required the presence of copper but not other transition metals. By contrast, benoic acid did not produce DNA damage in this system, either in the presence or in the absence of copper. The inclusion of spin trapping agents, such as N-tert-butyl-alpha-phenylnitrone (PBN), 3,5-dibromo-4-nitrosobenzenesulfonate, and nitrosobenzene, in incubations was found to significantly reduce the extent of DNA damage generated via the copper-mediated activation of BzPO. Electron paramagnetic resonance spectroscopy studies suggested that the primary radical trapped by PBN following copper-mediated decomposition of BzPO was the benzoyloxy radical.(ABSTRACT TRUNCATED AT 250 WORDS)

During a 4 and a half-year experience, 283 patients were referred to the Australian National Liver Transplant Unit. Sixty (21%) were children. The major causes of liver failure in the adults were chronic active hepatitis (27%), primary biliary cirrhosis (13%), primary sclerosing cholangitis (12%), fulminant hepatic failure (9%), alcoholic cirrhosis (9%), and malignancy (9%). In the children they were biliary atresia (43%) and inborn errors of metabolism (18%). One hundred and ninety-seven (69%) were accepted for liver transplantation. Of 111 (39%) accepted for early transplantation, 18 (16%) died before a donor became available. There were 319 possible organ donors. Ninety patients (24 of them children) received 100 grafts. Sixty-three (70%) patients survived. For all recipients, 1-year survival was 73%. Two-, 3- and 4-year survivals were 67%. One- to 4-year survivals for adults with benign conditions were 76%, contrasting with results for those with hepatic malignancy (20% 1-year survival). Children weighing greater than 8 kg did well whether they received whole grafts (80% 1- to 4-year survival) or reduced grafts (75% 1- to 4-year survival). Infants weighing less than 8 kg who received reduced adult grafts did significantly worse (20% 1- to 4-year survival). All 8 (100%) patients with fulminant hepatic failure who received grafts survived, including 3 who received ABO-incompatible grafts, though 2 of these subsequently required retransplantation. Rehabilitation of survivors was excellent with 91% of adults and 94% of children pursuing normal activities. Only 2 (2%) grafts failed with primary nonfunction, both in infants because of infarction. Graft survival was significantly worse (p less than 0.01) in patients with a positive direct crossmatch test against the donor. We calculate that the need for liver transplantation in Australia is approximately 7 per million of population per year. Increased donor offers are required to avoid deaths of patients on the waiting list. Reduced-size adult livers are successful for children and have alleviated considerably the critical shortage of pediatric donor livers. Liver transplantation is a highly satisfactory treatment for patients with benign liver disorders but not for those with malignant conditions. Patients with fulminant hepatic failure not responding to conservative therapy should be treated by liver transplantation. In this and other urgent circumstances, an ABO-incompatible liver may be lifesaving though retransplantation with an ABO-compatible liver may subsequently be required. The results of liver transplantation for nonmalignant conditions have improved steadily with clinical experience, with 1- and 2-year patient survivals during the past 28 months of 85%.

In an attempt to dissociate the chemotherapeutic from the carcinogenic properties of the antischistosomal and antitrypanosomal nitrovinylfuran trans-5-amino-3-[2-(5-nitro-2-furyl)vinyl]1,2,4-oxadiazole [(SQ18506) CAS: 28754-68-9; (E)-5-amino-3-(2-(5-nitro-2-furyl)vinyl)-1,2,4-oxadiazole], potential inhibitors of carcinogenesis were administered to female outbred CD-1 mice before and during exposure to SQ18506. The compounds tested were ascorbic acid, etretinate, butylated hydroxyanisole (BHA), cysteamine hydrochloride, cysteine hydrochloride, dimercaprol, disulfiram, 1,4-dithiothreitol, reduced glutathione, and spermidine phosphate. The primary types of tumors observed were squamous cell carcinomas of the stomach and thymic and nonthymic lymphomas. BHA reduced the incidence of malignant tumors to control levels, whereas cysteine hydrochloride, spermidine phosphate, and disulfiram reduced the incidence of chemically induced tumors by 42, 34, and 32%, respectively. Although cysteamine hydrochloride and disulfiram had no or only a modest effect on the overall incidence of tumors, the data suggested possible tissue-specific anticarcinogenic properties for these agents. Of the 8 antioxidants tested, only 1 had marked anticarcinogenic properties against SQ18506. These data indicate that antioxidant properties alone cannot account for the anticarcinogenic activity of the compounds tested. Coadministration of the anticarcinogen BHA with SQ18506 also blocked the chemotherapeutic effects of this agent on female CD-1 mice infected with Schistosoma mansoni.

1,2-Dithiol-3-thiones, reported constituents of cruciferous vegetables, are five-membered cyclic sulfur-containing compounds with antioxidant, chemotherapeutic, and chemoprotective activities. The effects of dietary administration of a substituted 1,2-dithiol-3-thione, oltipraz [5-(2-pyrazinyl)-4-methyl-1,2-dithiol-3-thione], a potent antischistosomal agent, on aflatoxin B1 (AFB1) metabolism, DNA adduct formation, and hepatic tumorigenesis were examined in male F344 rats. Rats were fed graded doses of oltipraz (0.01-0.1%) for 4 wk. During the second and third wk of oltipraz feeding rats were gavaged with 250 micrograms of AFB1/kg five times a wk. Rats were finally restored to control diet 1 wk after cessation of AFB1 dosing. At 4 months focal areas of hepatocellular alteration were identified and quantitated by staining sections of liver for gamma-glutamyl transpeptidase activity. Treatment with oltipraz at all doses reduced by greater than 90% the volume of liver occupied by gamma-glutamyl transpeptidase-positive foci. Levels of AFB1 bound to hepatic DNA were reduced between 40 and 80% in animals fed increasing doses of dietary oltipraz (0.01-0.1%) for 1 wk prior to a single exposure to AFB1. Feeding of the higher levels of oltipraz led to marked increases in the specific activity of glutathione S-transferases, presumably serving to facilitate the detoxication of the ultimate electrophilic form of AFB1, the 8,9-oxide. At low dietary concentrations of oltipraz (0.01%), the only inductive effects seen were on the activities of selected cytochrome P-450 monooxygenases. Therefore, the protection afforded by oltipraz may be due to both the enhancement of electrophile detoxication pathways as well as modified oxidative metabolism of AFB1. In in vitro metabolism studies with hepatic post-mitochondrial supernatant, low-dose oltipraz pretreatment facilitated the oxidative production of aflatoxins P1 and Q1, but not M1, from AFB1. High-dose (0.1%) oltipraz pretreatment enhanced the primary metabolism of AFB1 to aflatoxins P1, M1, and Q1 as well as the formation of chloroform-insoluble metabolites. Feeding studies with a series of 1,2-dithiol-3-thione and 1,2-dithiol-3-one derivatives of oltipraz demonstrated that the inductive activity for cytochrome P-450-dependent monooxygenases and electrophile detoxication enzymes, such as glutathione S-transferases, could be readily separated by minor modifications of the 1,2-dithiol-3-thione structure. The unsubstituted 1,2-dithiol-3-thione nucleus strongly induced electrophile detoxication enzymes, but not the monooxygenases, and was the most effective inhibitor of the binding of AFB1 to hepatic DNA in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)

The antischistosomal and antitrypanosomal drug trans-5-amino-3-[2-(5-nitro-2-furyl)vinyl]-1,2,4-oxadiazole [(SQ18506) CAS: 28754-68-9; (E)-amino-3-(2-(5-nitro-2-furyl)-vinyl)-1,2,4-oxadiazole] was carcinogenic for both male and female CD-1 mice when it was administered either in the diet or by gastric intubation. Dose-dependent increases in tumors of the forestomach and lymphatic tissues were observed in all groups receiving SQ18506 including mice infected with Schistosoma mansoni. The predominant tumor observed was squamous cell carcinoma of the forestomach. The presence or absence of schistosome infection did not appear to alter the incidence or distribution of tumors at comparable doses of SQ18506. The incidence of bladder tumors was positively correlated with the dose in gastric intubation studies and inversely correlated with the dose in dietary studies. The carcinogen N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (CAS: 24554-26-5) was fed to male and female CD-1 mice in the diet as a positive control. The predominant tumor observed in these groups was transitional cell carcinoma of the bladder. These data indicate that SQ18506 is unsuitable for use in the treatment of parasitic diseases.