Biomedical subjects

F F Kadlubar

Publications and source records attributed to F F Kadlubar.

At least 19 recordsLinked to original sources

PubMed2001

Frequency of CYP2A6 gene deletion and its relation to risk of lung and esophageal cancer in the Chinese population.

Cytochrome P450 2A6 (CYP2A6) plays an important role in the oxidation of nicotine and in the activation of tobacco-related carcinogens, such as N-nitrosodimethylamine, N-nitrosodiethylamine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone. It has been suggested that individuals with defective CYP2A6 alleles are at a lower risk of becoming smokers and of developing lung and other tobacco-related cancers. We examined the relationship between the CYP2A6 gene deletion and susceptibility to lung and esophageal cancer in a Chinese population via a hospital-based case-control study. The CYP2A6 gene deletion was determined by a PCR-based approach in 326 healthy controls, 149 patients with esophageal squamous-cell carcinoma and 151 patients with lung cancer. The allele frequency of the CYP2A6*4 deletion was 8.6% among controls compared with 8.4% among cases with esophageal squamous-cell carcinoma (p = 0.29) or 13.2% among cases with lung cancer (p < 0.01). Individuals who harbored at least one CYP2A6*4 deletion allele were at a 2-fold increased risk of developing lung cancer (95% confidence interval [CI] = 1.2-3.2) compared with those without a defective CYP2A6 allele. This effect was mainly limited to squamous-cell carcinoma and to non-smokers, although a joint effect of CYP2A6 deletion and tobacco smoking on lung cancer risk was observed among heavy smokers. The overall risk of esophageal cancer did not appear to be associated with this CYP2A6 genetic polymorphism (odds ratio [OR] = 1.2, 95% CI = 0.7-2.1). However, stratified analysis suggested an excess risk with borderline significance (OR = 2.1; 95% CI = 1.0-4.5) related to the CYP2A6*4 allele among non-smokers. The distribution of CYP2A6 genotype frequency was not significantly different (p = 0.40) between smokers (n = 174) and non-smokers (n = 152) in this study population. Our results demonstrate that the CYP2A6 gene deletion is associated with an increased risk of lung and esophageal cancer but not with a reduced tendency to smoke.

Aged↗

PubMed2001

Carcinogen substrate specificity of human COX-1 and COX-2.

The activation of carcinogenic aromatic and heterocyclic amines and benzo[a]pyrene-7,8-diol to intracellular electrophiles by prostaglandin H synthase (COX) is well documented for ovine sources of this enzyme. Here, the arachidonic acid-dependent activation of substrates by human (h)COX-1 and-2 is examined, utilizing recombinant enzymes. The COX-dependent activation of benzidine (BZ), 4-aminobiphenyl, (+)benzo[a]pyrene-7,8-diol, (+)benzo[a]pyrene-7,8-diol, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3-methylimidazo [4,5-f]quinoline (IQ), 2-amino-1-methyl-6-phenylimidazo [4,5-b] pyridine (PhIP), and 4,4'-methylenebis(2-chloroaniline) (MOCA) is assessed by means of COX-catalyzed, covalent DNA binding. The hCOX isozymes activated all substrates tested, activation varied from barely detectable for IQ (0.76 and 1.52 pmol bound/mg DNA for COX-1 and -2, respectively) to a high of 65 and 117 pmol bound/mg DNA for COX-1 and -2, respectively, for the activation of MOCA. BZ, which is an excellent peroxidase substrate, did not exhibit high DNA binding levels in hCOX assays and this phenomenon was found to be due to high levels of binding to protein, which effectively competed with the DNA for binding in the assay. The demonstrated ability of the COX enzymes to activate a variety of environmental and dietary carcinogens indicates a potential role for COX in the activation pathway of aromatic and heterocyclic amines and polycyclic hydrocarbons at extra-hepatic sites during early or late stages of carcinogenesis.

Animals↗

PubMed2001

In vitro human tissue models in risk assessment: report of a consensus-building workshop.

Advances in the technology of human cell and tissue culture and the increasing availability of human tissue for laboratory studies have led to the increased use of in vitro human tissue models in toxicology and pharmacodynamics studies and in quantitative modeling of metabolism, pharmacokinetic behavior, and transport. In recognition of the potential importance of such models in toxicological risk assessment, the Society of Toxicology sponsored a workshop to evaluate the current status of human cell and tissue models and to develop consensus recommendations on the use of such models to improve the scientific basis of risk assessment. This report summarizes the evaluation by invited experts and workshop attendees of the current status of such models for prediction of human metabolism and identification of drug-drug interactions, prediction of human toxicities, and quantitative modeling of pharmacokinetic and pharmaco-toxicodynamic behavior. Consensus recommendations for the application and improvement of current models are presented.

Cell Culture Techniques↗

PubMed2001

In situ hybridization and immunohistochemical analysis of cytochrome P450 1B1 expression in human normal tissues.

Cytochrome P450 1B1 (CYP1B1) is a recently cloned dioxin-inducible form of the cytochrome P450 supergene family of xenobiotic-metabolizing enzymes. CYP1B1 is constitutively expressed mainly in extrahepatic tissues and is inducible by aryl hydrocarbon receptor ligands. Human CYP1B1 is involved in activation of chemically diverse human procarcinogens, including polycyclic aromatic hydrocarbons and some aromatic amines, as well as the endogenous hormone 17 beta-estradiol. The metabolism of 17 beta-estradiol by CYP1B1 forms 4-hydroxyestradiol, a product believed to be important in estrogen-induced carcinogenesis. Although the distribution of CYP1B1 mRNA and protein in a number of human normal tissues has been well documented, neither the cells expressing CYP1B1 in individual tissue nor the intracellular localization of the enzyme has been thoroughly characterized. In this study, using nonradioactive in situ hybridization and immunohistochemistry, we examined the cellular localization of CYP1B1 mRNA and protein in a range of human normal tissues. CYP1B1 mRNA and protein were expressed in most samples of parenchymal and stromal tissue from brain, kidney, prostate, breast, cervix, uterus, ovary, and lymph nodes. In most tissues, CYP1B1 immunostaining was nuclear. However, in tubule cells of kidney and secretory cells of mammary gland, immunoreactivity for CYP1B1 protein was found in both nucleus and cytoplasm. This study demonstrates for the first time the nuclear localization of CYP1B1 protein. Moreover, the constitutive expression and wide distribution of CYP1B1 mRNA and protein in many human normal tissues suggest functional roles for CYP1B1 in the bioactivation of xenobiotic procarcinogens and endogenous substrates such as estrogens. (J Histochem Cytochem 49:229-236, 2001)

Aryl Hydrocarbon Hydroxylases↗

PubMed2000

Association between survival after treatment for breast cancer and glutathione S-transferase P1 Ile105Val polymorphism.

A glutathione S-transferase (GST) P1 polymorphism results in an amino acid substitution, Ile105Val; the Val-containing enzyme has reduced activity toward alkylating agents. Cancer patients with the variant enzyme may differ in removal of treatment agents and in outcomes of therapy. We evaluated survival according to GSTP1 genotype among women (n = 240) treated for breast cancer. Women with the low-activity Val/Val genotype had better survival. Compared with Ile/Ile, hazard ratios for overall survival were 0.8 (95% confidence interval, 0.5-1.3) for Ile/Val and 0.3 (95% confidence interval, 0.1-1.0) for Val/Val (P for trend = 0.04). Inherited metabolic variability may influence treatment outcomes.

Adult↗

PubMed2000

Expression of hGSTP1 alleles in human lung and catalytic activity of the native protein variants towards 1-chloro-2,4-dinitrobenzene, 4-vinylpyridine and (+)-anti benzo[a]pyrene-7,8-diol-9,10-oxide.

The human glutathione S-transferase (GST) P1 alleles coding for Val(105) (hGSTP1*B and/or P1*C) are over- represented in lung cancer patients. However, the corresponding recombinant Val(105) protein variants tend to show higher catalytic activity than the Ile(105) variants towards bay-region diol epoxides that are thought to be etiological agents in lung cancer. We have examined 29 normal human lung samples with respect to several factors that could confound relationships between hGSTP1 allele type and cancer susceptibility, namely, inter-individual and allele-specific variation of hGSTP1 expression, and differences between the catalytic properties of the native and recombinant hGSTP1-1 variant protein products. hGSTP1 expression varied 7-fold among individuals but was independent of hGSTP1*A, P1*B or P1*C allele type. hGST subunits A1, A2, M1 and M3 were minor components, similarly variable in expression. Despite this variability of expression, the levels of hGSTP1 expression linearly correlated with those of the next most highly expressed GST, hGSTM3, even though the genes for these GSTs are on different chromosomes. Differences between the native protein variants, using 1-chloro-2,4-dinitrobenzene and (+)-anti-benzo[a]pyrene diolepoxide as substrates, were more marked than those between the recombinant variants. However, the order of differential catalytic specificity was the same for native and recombinant variants. Neither the expression of the hGSTP1 alleles nor the catalytic properties of the protein variants appears to provide a simple mechanistic rationale for the observed over-representation of the hGSTP1*B and/or 1*C alleles in lung cancer.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

PubMed2000

Quantitative analysis of interindividual variation of glutathione S-transferase expression in human pancreas and the ambiguity of correlating genotype with phenotype.

Analysis of glutathione S-transferases (GSTs) of the alpha, mu, and pi classes by reverse-phase high-performance liquid chromatography and electrospray-ionization mass spectrometry in 43 samples of normal human pancreas demonstrated a wide variation in expression of subunits P1, A1, A2, A4, M1, M2, and M3 and the presence of a novel form designated GST "A5." GSTA2 consisted of three forms that were differentially expressed between individuals in a manner consistent with allelic polymorphism at the hGSTA2 locus. Expression, in terms of microg GST subunit/mg cytosolic protein, varied by 6-15-fold for subunits P1, A2, and M3 and 17-30-fold in the case of GSTs A1 and M2. Less consistently expressed were GSTs M1a, M1b, A4, and A5. Among these, GSTM1 expression (excluding M1-null samples) varied 12-fold between samples, whereas GST A4 and A5 expression varied approximately 50-100-fold between samples, well beyond the range of other subunits, suggesting that their expression is highly inducible. Linear correlations (P < 0.001-0.003) existed between levels of the most consistently expressed GST, GSTP1, and total GSTs, GSTA2 and M3, and in GSTM1-positive samples, between GSTM1, M3, and P1. The correlation between GST subunits P1 and M3 was bimodal according to M1 genotype, reflecting the presence of the regulatory element in hGSTM3*B that is linked with the hGSTM1*A genotype. It is concluded that although a degree of regulation of expression of GSTs occurs in human pancreas, the variability of phenotype is high and might obscure the effects of genetic polymorphisms on individual cancer susceptibility. Interindividual variation of GST expression is, therefore, a factor that should be taken account of in epidemiological studies.

Adolescent↗

PubMed2000

Metabolic activation capacity of neonatal mice in relation to the neonatal mouse tumorigenicity bioassay.

The neonatal mouse tumorigenicity bioassay is a well-developed animal model that has recently been recommended as an alternative tumorigenicity bioassay by the International Conference on Harmonization (ICH) for Technical Requirements for the Registration of Pharmaceuticals for Human Use. There are sufficient data to conclude that this animal model is highly sensitive to genotoxic chemical carcinogens that exert their tumorigenicity through mechanisms involving the formation of covalently bound exogenous DNA adducts that lead to mutation. On the other hand, it is not sensitive to chemical carcinogens that exert tumorigenicity through a secondary mechanism. The metabolizing enzymes present in the neonatal mouse, particularly the cytochromes P450, are critical factors in determining the tumorigenic potency of a chemical tested in this bioassay. However, compared to the metabolizing enzymes of the adult mouse and rat, the study of the metabolizing enzymes in neonatal mouse tissues has been relatively limited.

Animals↗

PubMed2000

Human CYP1B1 Leu432Val gene polymorphism: ethnic distribution in African-Americans, Caucasians and Chinese; oestradiol hydroxylase activity; and distribution in prostate cancer cases and controls.

Cytochrome P4501B1 (CYP1B1) is involved in the activation of many carcinogens and in the metabolism of steroid hormones, including 17beta-oestradiol (E2) and testosterone. We report a significant difference in the allele frequencies of two point mutations in the coding region of the CYP1B1 gene among Caucasian (n = 189), African-American (n = 52) and Chinese (Linxian) (n = 109) populations. A (C to G) transversion at position 1666 in exon 3, which results in an amino acid substitution of Leu432 to Val, was present in African-Americans with an allele frequency for Va1432 of 0.75, in Caucasians of 0.43, and in Chinese of 0.17. A (C to T) transition at position 1719 in exon 3, with no amino acid change (Asp449), appeared to be closely linked with the Val432 variant. Results using human lung microsomal preparations from individuals with the CYP1B1Val/Val and CYP1B1Leu/Leu genotypes indicate that Val432 variant may be a high activity allele and thus may contribute to the interindividual differences in CYP1B1 activity. Because CYP1B1 is involved in hormone and carcinogen metabolism, and given the disparate rates of prostate cancer among ethnic groups, we also evaluated the association of the CYP1B1 Leu432Val polymorphism with prostate cancer risk in a pilot case-control study. Among Caucasians, 34% of men with cancer (n = 50) were homozygous for the Val432 polymorphism, while only 12% of matched control subjects (n = 50) had this genotype. These preliminary data indicate that genetic polymorphisms in CYP1B1 might play an important role in human prostate carcinogenesis.

Adult↗

PubMed2000

Relationship of phenol sulfotransferase activity (SULT1A1) genotype to sulfotransferase phenotype in platelet cytosol.

Sulfation catalysed by human cytosolic sulfotransferases is generally considered to be a detoxification mechanism. Recently, it has been demonstrated that sulfation of heterocyclic aromatic amines by human phenol sulfotransferase (SULT1A1) can result in a DNA binding species. Therefore, sulfation capacity has the potential to influence chemical carcinogenesis in humans. To date, one genetic polymorphism (Arg213His) has been identified that is associated with reduced platelet sulfotransferase activity. In this study, data on age, race, gender, SULT1A1 genotype and platelet SULT1A1 activity were available for 279 individuals. A simple colorimetric phenotyping assay, in conjunction with genotyping, was employed to demonstrate a significant correlation (r = 0.23, P < 0.01) of SULT1A1 genotype and platelet sulfotransferase activity towards 2-naphthol, a marker substrate for this enzyme. There was also a difference in mean sulfotransferase activity based on gender (1.28 nmol/min/mg, females; 0.94 nmol/min/mg, males, P = 0.001). DNA binding studies using recombinant SULT1A1*1 and SULT1A1*2 revealed that SULT1A1*1 catalysed N-hydroxy-aminobiphenyl (N-OH-ABP) DNA adduct formation with substantially greater efficiency (5.4 versus 0.4 pmol bound/mg DNA/20 min) than the SULT1A1*2 variant. A similar pattern was observed with 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5b]pyridine (N-OH-PhIP) (4.6 versus 1.8 pmol bound/mg DNA/20 min).

Age Factors↗

PubMed2000

Cancer therapy and polymorphisms of cytochromes P450.

Cytochrome P450 (CYP) enzymes are important in the metabolism of some endogenous compounds, environmental and dietary xenobiotics and many drugs. Many of these enzymes have genetic polymorphisms that produce significant changes in metabolic activity, however the function of other polymorphisms is unknown. Genetic polymorphisms have important influences on variability in human pharmacokinetics, including intra-individual differences in drug toxicity, drug interactions and response to chemotherapy. Other factors that influence drug metabolism include differences in enzyme expression due to differences in age, gender, smoking status, exposure to dietary or environmental xenobiotics or co-administration of other drugs. In addition, some xenobiotics and drugs can directly inhibit or induce the activity of CYPs. All of these factors can produce differences in metabolic capacities among individuals which can produce toxicity in some patients and sub-effective dosing in others. Maximum clinical benefit will require a more complete understanding of the influence of these polymorphisms on allele function and their interaction with inducers and inhibitors of enzyme expression or activity. This effort will permit the pharmacogenetic screening of patients before the administration of drugs and result in the identification of individuals who are prone to adverse reactions or poor response, resulting in more effective individualized therapy.

Antineoplastic Agents↗

PubMed2000

Characterization of an ATP-dependent pathway of activation for the heterocyclic amine carcinogen N-hydroxy-2-amino-3-methylimidazo[4, 5-f]quinoline.

2-amino-3-methylimidazo[4,5-f]quinoline (IQ) is one of several mutagenic and carcinogenic heterocyclic amines formed during the cooking process of protein-rich foods. These compounds are highly mutagenic and have been shown to produce tumours in various tissues in rodents and non-human primates. Metabolic activation of IQ is a two-step process involving N-hydroxylation by CYP1A2 followed by esterification to a more reactive species capable of forming adducts with DNA. To date, acetylation and sulphation have been proposed as important pathways in the formation of N-hydroxy esters. In this study we have demonstrated the presence of an ATP-dependent activation pathway for N-hydroxy-IQ (N-OH-IQ) leading to DNA adduct formation measured by covalent binding of [(3)H]N-OH-IQ to DNA. ATP-dependent DNA binding of N-OH-IQ was greatest in the cytosolic fraction of rat liver, although significant activity was also seen in colon, pancreas and lung. ATP was able to activate N-OH-IQ almost 10 times faster than N-hydroxy-2-amino-1-methyl-6-phenylimidazo[4, 5-b]pyridine (7.7 +/- 0.3 and 0.9 +/- 0.1 pmol/mg protein/min, respectively). Using reported intracellular concentrations of cofactor, the ability of ATP to support DNA binding was similar to that seen with 3'-phosphoadenosine 5'-phosphosulphate and approximately 50% of that seen with acetyl coenzyme A (AcCoA). In addition to DNA binding, HPLC analysis of the reaction mixtures using ATP as co-factor showed the presence of two stable, polar metabolites. With AcCoA, only one metabolite was seen. The kinase inhibitors genistein, tyrphostin A25 and rottlerin significantly inhibited both DNA binding and metabolite formation with ATP. However, inhibition was unlikely to be due to effects on enzyme activity since the broad spectrum kinase inhibitor staurosporine had no effect and the inactive analogue of genistein, daidzein, was as potent as genistein. The effects of genistein and daidzein, which are naturally occurring isoflavones from soy and other food products, on DNA adduct formation may potentially be useful in the prevention of heterocyclic amine-induced carcinogenesis.

Acetylation↗

PubMed2000

Impact of genetic polymorphisms in cytochrome P450 2E1 and glutathione S-transferases M1, T1, and P1 on susceptibility to esophageal cancer among high-risk individuals in China.

Esophageal cancer, which is prevalent in China, is believed to be induced by environmental carcinogens such as nitrosamines and other agents. The disproportionate geographical distribution of this cancer among individuals suggests a role for gene-environment interactions in developing the disease. We have shown in our preliminary study that a genetic polymorphism in cytochrome P450 2E1 (CYP2E1) that is known to activate nitrosamines may be a susceptibility factor involved in the early events leading to the development of esophageal cancer (Lin et al., Cancer Epidemiol. Biomark. Prev., 7: 1013-1018, 1998). This relatively larger study was conducted to compare the results with our previous findings. One hundred and fifty cases with esophageal cancer, 146 cases with esophageal dysplasia, and 150 normal controls were residents of Linxian, China, a high-risk area. Genomic DNA samples were assayed for restriction fragment length polymorphisms in the CYP2E1 and GSTP1 loci by PCR amplification followed by digestion with RsaI and Alw26I, respectively. Deletion of the GSTM1 and GSTT1 genes was detected by multiplex PCR. The distribution of CYP2E1 c1/c1 allele frequency was found to be significantly different between controls (44.0%) and cases with cancer (71.3%) or cases with dysplasia (70.6%; P < 0.0001). Individuals having the c1/c1 genotype were at a 3.1-fold [95% confidence interval (CI), 2.4-3.9] increased risk of developing dysplasia and a 3.2-fold (95% CI, 2.5-4.1) increased risk of developing squamous cell carcinoma of the esophagus. Although polymorphisms in the GSTT1 and GSTP1 were not significantly different between cases with cancer or cases with dysplasia and controls, the frequency of the GSTM1 non-null (+/+ and +/0) genotypes appeared to be overrepresented in cases with cancer compared with controls (odds ratio, 2.3; 95% CI, 1.8-3.0). Furthermore, a joint effect of the CYP2E1 c1/c1 genotype and GSTM1 non-null genotype on the cancer risk was observed, showing an odds ratio of 8.5 (95% CI, 3.7-19.9). These results demonstrate that CYP2E1 and perhaps GSTM1 are genetic determinants in the development of squamous cell carcinoma of the esophagus.

Carcinoma, Squamous Cell↗

PubMed2000

In vitro bioactivation of N-hydroxy-2-amino-alpha-carboline.

2-Amino-alpha-carboline (A alpha C) is a mutagenic and carcinogenic heterocyclic amine present in foods cooked at high temperature and in cigarette smoke. The mutagenic activity of A alpha C is dependent upon metabolic activation to N-hydroxy-A alpha C (N-OH-A alpha C); however, the metabolism of N-OH-A alpha C has not been studied. We have synthesized 2-nitro-alpha-carboline and N-OH-A alpha C and have examined in vitro bioactivation of N-OH-A alpha C by human and rodent liver cytosolic sulfotransferase(s) and acetyltransferase(s) and by recombinant human N-acetyltransferases, NAT1 and NAT2. The sulfotransferase-dependent bioactivation of N-OH-A alpha C by human liver cytosol exhibited large inter-individual variation (0.5-75, n = 14) and was significantly higher than bioactivation of N-hydroxy-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP). Correlation and inhibition studies suggested that the isoform of sulfotransferase primarily responsible for bioactivation of N-OH-A alpha C in human liver cytosol is SULT1A1. O-Acetyltransferase-dependent bioactivation of N-OH-A alpha C by human liver cytosol also exhibited large inter-individual variation (16-192, n = 18). In contrast to other N-hydroxy heterocyclic amines, which are primarily substrates only for NAT2, both NAT1 and NAT2 catalyzed bioactivation of N-OH-A alpha C. The rate of bioactivation of N-OH-A alpha C by both NAT1 and NAT2 was significantly higher than that for N-OH-PhIP. In rat and mouse liver cytosols, the level of sulfotransferase-dependent bioactivation of N-OH-A alpha C was similar to the level in the high sulfotransferase activity human liver cytosol. The level of O-acetyltransferase-dependent bioactivation of N-OH-A alpha C in rat liver cytosol was also comparable with that in the high acetyltransferase activity human liver cytosol. However, the level of O-acetyltransferase-dependent bioactivation of N-OH-A alpha C in mouse liver cytosol was comparable with that in the low acetyltransferase activity human liver cytosol. In contrast to N-OH-PhIP, bioactivation of N-OH-A alpha C was not inhibited by glutathione S-transferase activity; however, DNA binding of N-acetoxy-A alpha C was inhibited 20% in the presence of GSH. These results suggest that bioactivation of N-OH-A alpha C may be a significant source of DNA damage in human tissues after dietary exposure to AalphaC and that the relative contribution of each pathway to bioactivation or detoxification of N-OH-A alpha C differs significantly from other N-hydroxy heterocyclic or aromatic amines.

Acetylation↗

PubMed2000

A simple colorimetric assay for phenotyping the major human thermostable phenol sulfotransferase (SULT1A1) using platelet cytosols.

A thermostable phenol sulfotransferase, SULT1A1, has been implicated in numerous detoxification and bioactivation pathways; however, little is known regarding its endogenous function or its putative role in mediating risk for human environmental disease. A simple endpoint colorimetric assay is described that can be used for rapid phenotyping of SULT1A1 activity in human populations. The assay utilizes a microtiter-plate format and relatively small amounts of platelet cytosol-derived enzyme. The enzyme catalyzes the synthesis of 2-naphthylsulfate from 2-naphthol and 5'-phosphoadenosine 3'-phosphosulfate (PAPS), whereas addition of p-nitrophenyl sulfate to the assay contributes to an effective PAPS-regenerating system. In contrast to other sulfotransferase assay methods, 3'-phosphoadenosine 5'-phosphate (PAP) does not accumulate during the incubation to interfere with enzyme activity, but instead serves as a cofactor to cause the removal of sulfate from p-nitrophenyl sulfate to regenerate PAPS. This reaction concomitantly results in generation of p-nitrophenol that can be quantified colorimetrically at 405 nm (epsilon = 18,200 M(-1)) to give an indirect measure of sulfotransferase activity. Using platelet enzyme preparations from adult human subjects, sulfation rates of two prototypical thermostable phenol sulfotransferase substrates (2-naphthol and p-nitrophenol) and one thermolabile phenol sulfotransferase substrate (dopamine) were determined using standard radiochemical protocols. These data were then compared with results from the colorimetric assay using 2-naphthol as substrate. There was a good correlation between the phenotyping assay and radiochemical assays for both 2-naphthol sulfotransferase and p-nitrophenol sulfotransferase activity (r = 0.85 and 0.69, respectively). However, SULT1A1 activity was approximately 10 to 20 times higher with the colorimetric determination. As anticipated, there was no correlation between SULT1A1 activity and dopamine sulfotransferase activity (r = 0.07) in these human platelet preparations. This inexpensive and rapid method for phenotyping SULT1A1 activity may help investigators assess a role for this enzyme in disease susceptibility.

Adult↗

PubMed2000

Pharmacogenetics of the arylamine N-acetyltransferases: a symposium in honor of Wendell W. Weber.

This article is a report on a symposium sponsored by the American Society for Pharmacology and Experimental Therapeutics presented at the joint meeting of the American Society for Biochemistry and Molecular Biology and the American Society for Pharmacology and Experimental Therapeutics, June 4-8, Boston, Massachusetts. The presentations focused on the pharmacogenetics of the NAT1 and NAT2 arylamine N-acetyltransferases, including developmental regulation, structure-function relationships, and their possible role in susceptibility to breast, colon, and pancreatic cancers. The symposium honored Wendell W. Weber for over 35 years of leadership and scientific advancement in pharmacogenetics and was highlighted by his overview of the historical development of the field.

Animals↗

PubMed1999

Detoxification of carcinogenic aromatic and heterocyclic amines by enzymatic reduction of the N-hydroxy derivative.

The metabolic activation pathways associated with carcinogenic aromatic and heterocyclic amines have long been known to involve N-oxidation, catalyzed primarily by cytochrome P4501A2, and subsequent O-esterification, often catalyzed by acetyltransferases (NATs) and sulfotransferases (SULTs). We have found a new enzymatic mechanism of carcinogen detoxification: a microsomal NADH-dependent reductase that rapidly converts the N-hydroxy arylamine back to the parent amine. The following N-OH-arylamines and N-OH-heterocyclic amines were rapidly reduced by both human and rat liver microsomes: NOH-4-aminoazobenzene, N-OH-4-aminobiphenyl (N-OH-ABP), N-OH-aniline, N-OH-2-naphthylamine, N-OH-2-aminofluorene, N-OH-4,4'-methylenebis(2-chloroaniline) (N-OH-MOCA), N-OH-1-naphthyamine, N-OH-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP), N-OH-2-amino-alpha-carboline (N-OH-AalphaC), N-OH-2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (N-OH-MeIQx), and N-OH-2-amino-3-methylimidazo[4,5-f]quinoline (N-OH-IQ). In addition, primary rat hepatocytes and human HepG2 cells efficiently reduced N-OH-PhIP to PhIP. This previously unrecognized detoxification pathway may limit the bioavailability of carcinogenic N-OH heterocyclic and aromatic amines for further activation, DNA adduct formation, and carcinogenesis.

Animals↗

PubMed1999

The effects of phytoestrogens on human pancreatic tumor cells in vitro.

Diet has been implicated as a possible link to the etiology, promotion and/or progression of many diseases, including cancer. Recently, interest has been focused on the cancer-protective role of several of the hormone-like diphenolic phytoestrogens, lignans, and isoflavonoids. This study examined the chemoprotective effects of genistein, biochanin A, equol, and coumestrol on human pancreatic adenocarcinoma cells in vitro. Two human adenocarcinoma cell lines, HPAF-11 from a male and Su 86.86 from a female, were used. HPAF-11 cells were exposed for 24 h to these agents at concentrations of 1 and 10 microM. Su 86.86 cells were exposed for 24 h at a concentration of 1 microM. Coumestrol and equol at higher concentrations were toxic to the Su 86.86 cells. These agents displayed marked differences between cell lines in inhibition of growth. Equol and coumestrol inhibited the growth of the female pancreatic tumor cells by 95%; however, these agents stimulated the growth of pancreatic tumor cells from the male. Genistein also stimulated growth in the male pancreatic tumor cells, but had little effect on pancreatic tumor cells from the female. Biochanin A inhibited growth of both male and female tumor cells, but to a lesser extent than other agents. This study also indicated a difference in K-ras expression in pancreatic tumors cells treated with these agents. Equol and coumestrol decreased K-ras expression in the female tumor cell line. Genistein increased expression of K-ras in both male and female pancreatic tumor cells. Genistein also increased expressions of the multidrug resistant (mdr-1) gene in the male tumor-cell line, while coumestrol and biochanin A decreased expression. Equol had no effect on mdr-1 expression. Whether the chemoprotective potential of equol and coumestrol against pancreatic cancer is greater in females than males is being further studied.

Analysis of Variance↗