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S Amin

Publications and source records attributed to S Amin.

At least 109 records · Page 6Linked to original sources
PubMed1996

Solution conformation of the (-)-cis-anti-benzo[a]pyrenyl-dG adduct opposite dC in a DNA duplex: intercalation of the covalently attached BP ring into the helix with base displacement of the modified deoxyguanosine into the major groove.

This paper reports on the combined NMR-molecular mechanics computational studies of the solution structure of the (-)-cis-anti-[BP]dG adduct positioned opposite dC in the sequence context d(C1- C2-A3-T4-C5-[BP]G6-C7-T8-A9-C10-C11).d(G12-G13-T14- A15-G16-C17-G18-A19-T20- G21-G22) duplex [designated (-)-cis-anti-[BP]dG.dC 11-mer duplex]. This adduct is derived from cis addition at C10 of (-)-anti-7(S),8(R)-dihydroxy-9(R),10(S)-epoxy-7,8,9,10- tetrahydrobenzo[a]pyrene [(-)-anti-BPDE] to the N2 position of dG6 in this duplex sequence. The exchangeable and nonexchangeable protons of the benzo[a]pyrenyl moiety and nucleic acid of the major conformation were assigned following analysis of two-dimensional NMR data sets in H2O and D2O solution. There was a general broadening of proton resonances for a three-nucleotide segment centered about the lesion site which resulted in a tentative assignment for the sugar protons of the C7 residue in the spectrum of the adduct duplex. The solution conformation of the major conformation of the (-)-cis-anti-[BP]dG.dC 11-mer duplex has been determined by incorporating DNA-DNA and intermolecular BP-DNA proton-proton distances defined by lower and upper bounds deduced from NOESY data sets as restraints in molecular mechanics computations in torsion angle space. The results establish that the covalently attached benzo[a]pyrenyl ring intercalates between intact Watson-Crick dC5.dG18 and dC7.dG16 base pairs. The modified deoxyguanosine [BP]-dG6 and its partner cytosine dC17 are looped out of the helix into the major groove. The purine ring of the [BP]dG6 residue is directed toward the 5'-end of the modified strand and stacks over the major groove edge of its 5'-side neighbor dC5 residue. The solution structure of the (-)-cis-anti-[BP]dG.dC 11-mer duplex is compared with those of the stereoisomeric (+)-trans-anti-[BP]dG [Cosman, M., et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 1914-1918], (-)-trans-anti-[BP]dG [de los Santos, C., et al. (1992) Biochemistry 31, 5245-5252], and (+)-cis-anti-[BP]dG [Cosman, M., et al. (1993a) Biochemistry 32, 4146-4155] adducts positioned opposite dC in the same duplex sequence context. A key finding is that the long axes of the intercalated benzo[a]pyrenyl rings in the solution structures of the (+)- and (-)- cis-anti-[BP]dG.dC 11-mer duplexes are oriented in opposite directions with the benzylic ring directed toward the minor groove in the (+)-cis isomer and toward the major groove in the (-)-cis isomer. In addition, a comparison is also made with the solution structure of the (+)-trans-anti-[BP]dG adduct opposite a deletion site [Cosman, M., et al. (1994a) Biochemistry 33, 11507-11517] since this adduct duplex displays several conformational features in common with the structure of the (-)-cis-anti-[BP]dG.dC 11-mer duplex. The structures of both duplex adducts exhibit intercalation of the covalently attached ligand into the helix and displacement of the modified deoxyguanosine into the major groove. Studies of the biological activities of stereochemically defined BP-DNA adducts and the comparison of the solution structure of the (-)-cis-anti-[BP]dG.dC 11-mer duplex with its stereoisomeric counterparts should lead to new insights into the relationships between defined helical distortions and mutagenic specificity and activity.

Base Composition↗
PubMed1996

Expression of a 32 kDa protein in rat mammary tumors induced by anti-benzo[c]phenanthrene-3,4-diol-1,2-epoxide.

Racemic anti-benzo[c]phenanthrene-3,4-diol-1,2-epoxide (BcPDE) is a powerful rat mammary carcinogen and a metabolite of benzo[c]phenanthrene, a polynuclear aromatic hydrocarbon found in the environment. In elucidating potential molecular mechanisms that may play a role in the development of BcPDE-induced rat mammary tumors, we have identified a 32 kDa protein in 16 of 26 tumors analyzed but in only 1 of the 15 normal mammary tissues that were examined. The 32 kDa protein was identified with antibodies to Ets, which also recognized the 55 kDa Ets-1 protein that was expressed at similar levels in normal mammary tissues. The expression of the 32 kDa protein was also observed in mammary tumor-derived cell lines of both rat and human origin and in human melanoma, but not in normal human keratinocytes or rat fibroblast cell lines. Further characterization via 2D gels revealed that the protein exhibits a PI of 5.5. Southwestern analysis using Ets-1 target sequence revealed binding of the 55 kDa Ets-1 but not of the newly identified 32 kDa protein. Overall, the preferential expression of the 32 kDa protein in mammary tumor tissues may serve as a biomarker to follow the development of this tumor type.

Animals↗
PubMed1996

Activation of chemically diverse procarcinogens by human cytochrome P-450 1B1.

A human cytochrome P-450 (P450) 1B1 cDNA was expressed in Saccharomyces cerevisiae and the microsomes containing P450 1B1 were used to examine the selectivity of this enzyme in the activation of a variety of environmental carcinogens and mutagens in Salmonella typhimurium TA1535/pSK1002 or NM2009 tester strains, using the SOS response as an end point of DNA damage. We also determined and compared these activities of P450 1B1 with those catalyzed by recombinant human P450s 1A1 and 1A2, which were purified from membranes of Escherichia coli. The carcinogenic chemicals tested included 27 polycyclic aromatic hydrocarbons and their dihydrodiol derivatives, 17 heterocyclic and aryl amines and aminoazo dyes, three mycotoxins, two nitroaromatic hydrocarbons, N-nitrosodimethylamine, vinyl carbamate, and acrylonitrile. Among the three P450 enzymes examined here, P450 lB1 was found to have the highest catalytic activities for the activation of 11,12-dihydroxy-11,12-dihydrodibenzo[a,l]pyrene, 1,2-dihydroxy-1,2-dihydro-5-methylchrysene, (+)-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene, 11,12-dihydroxy-11,12-dihydrobenzo[g]chrysene, 3,4-dihydroxy-3,4-dihydrobenzo[c]phenanthrene, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole, 2-aminoanthracene, 3-methoxy-4-aminoazobenzene, and 2-nitropyrene. P450 1B1 also catalyzed the activation of 2-amino-3,5-dimethylimidazo[4,5-f]quinoline, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, 2-amino-3-methylimidazo[4,5-f]quinoline, 2-aminofluorene, 6-aminochrysene and its 1,2-dihydrodiol, (-)-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene, 1,2-dihydroxy-1,2-dihydrochrysene, 1,2-dihydroxy-1,2-dihydro-5,6-dimethylchrysene, 2,3-dihydroxy-2,3-dihydrofluoranthene, 3,4-dihydroxy-3,4-dihydro-7,12-dimethylbenz[a]anthracene, and 6-nitrochrysene to appreciable extents. However, P450 1B1 did not produce genotoxic products from benzo[a]pyrene, trans- 3,4-dihydroxy-3,4-dihydrobenzo[a]anthracene, trans-8,9-dihydroxy-8,9-dihydrobenzo[a]anthracene, 7,12-dimethylbenz[a]anthracene and its cis-5,6-dihydrodiol, 5-methylchrysene, 11,12-dihydroxy-11,12-dihydro-3-methylcholanthrene, 1,2-dihydroxy-1,2-dihydro-6-methylchrysene, benzo[c]phenanthrene, 2-amino-6-methyldipyridol[1,2-a:3',2'-d]imidazole, 2-acetylaminofluorene, benzidine, 2-naphthylamine, aflatoxin B1, aflatoxin G1, sterigmatocystin, N-nitrosodimethylamine, vinyl carbamate, or acrylonitrile in this assay system. P450 1B1 is expressed constitutively in extrahepatic organs, including fetal tissue samples, and is highly inducible in various organs by 2,3,7,8-tetrachlorodibenzo-p-dioxin and related compounds in experimental animal models. Thus, activation of procarcinogens by P450 lB1 may contribute to human tumors of extrahepatic origin.

Adult↗
PubMed1996

Lack of tumorigenicity of cholesterol epoxides and estrone-3,4-quinone in the rat mammary gland.

The purpose of this study is to test the long-standing hypothesis that endogenous agents found in human breast fluid and in plasma are potential initiators of breast cancer. Therefore, we evaluated the tumorigenicity in the mammary glands of female CD rats of cholestan-5 alpha,6 alpha-epoxy-3 beta-ol (cholesterol-alpha-epoxide), cholestan-5 beta,6 beta-epoxy-3 beta-ol (cholesterol-beta-epoxide), and 1,5(10)estradiene-3,14,17-trione (estrone-3,4-quinone). As a positive control, trans-3,4-dihydroxy-anti-1,2-epoxy-1,2,3,4-tetrahydrobenzo[c]phenanthren e (BcPDE) was used. Rats were fed a high-fat AIN-76A diet (23.5% corn oil) to mimic the Western dietary composition. Because literature data suggest that the endogenous agents tested in this study are weak electrophiles, the total doses of cholesterol epoxides (12.3 mumol/rat) and of estrone-3,4-quinone (30 mumol/rat)were 10- and 25- fold higher, respectively, than that of BcPDE (1.2 mumol/rat). Each agent was dissolved in DMSO, and one-sixth of the total dose was injected under each of six nipples on the right side. The thoracic glands of the rat were treated at 30 days of age, and those located in the inguinal area were treated on the following day. The experiment was terminated at 44 weeks after treatment. Consistent with our previous study, BcPDE was a strong mammary carcinogen. However, there were no differences between rats treated with DMSO alone or those receiving DMSO containing cholesterol-alpha-epoxide, cholesterol-beta-epoxide, or estrone-3,4-quinone. The results of this study clearly indicate, for the first time, that metabolites derived from cholesterol and estrone lack tumorigenic activity in the rat mammary gland, at least under the conditions of the present protocol.

Animals↗
PubMed1996

Tnk1: a novel intracellular tyrosine kinase gene isolated from human umbilical cord blood CD34+/Lin-/CD38- stem/progenitor cells.

Degenerate PCR was employed to identify novel tyrosine kinase genes from an enriched population of human umbilical cord blood hematopoietic stem/progenitor cells. One novel tyrosine kinase gene, designated Tnk1, was cloned. The sequence of the complete Tnk1 coding region predicts a 72 kD protein. Comparison of Tnk1 to available sequences in protein databases reveals that it is most homologous to Ack, an intracellular tyrosine kinase which associates with the GTP-bound form of p21cdc42Hs. Like Ack, Tnk1 consists of an N-terminal kinase domain, a putative SH3 domain immediately C-terminal to the kinase domain, and a proline-rich C-terminal region. Analysis of Tnk1 mRNA expression demonstrates that Tnk1 is expressed in all cord blood, bone marrow and adult blood sub-populations, as well as in most of the leukemia cell lines examined (16 of 20). Hybridization to fetal multi-tissue Northern blots detected several different Tnk1 transcripts in all fetal tissues examined. In contrast, a single Tnk1 transcript was detected in only five of 16 adult tissues examined (prostate, testis, ovary, small intestine and colon). Fluorescence in situ hybridization (FISH) analysis of metaphase chromosomes localized the Tnk1 gene to the short arm of chromosome 17 (17p13.1), near the p53 locus. Thus, Tnk1 is a novel tyrosine kinase that may be involved in signalling pathways utilized broadly during fetal development, more selectively in adult tissues and in cell of the lymphohematopoietic system.

ADP-ribosyl Cyclase↗
PubMed1996

Expression of the hematopoietic growth factor receptor FLT3 (STK-1/Flk2) in human leukemias.

Normal expression of the hematopoietic growth factor receptor FLT3 (STK-1@Flk2) is limited to CD34+ stem/progenitor cells. We have evaluated the expression of FLT3 by RNase protection assay and Western blotting in 161 primary bone marrow (BM) samples from patients with leukemia. FLT3 RNA was found to be expressed at a higher level than in normal BM controls in 33 of 33 B-lineage acute leukemias, 11 of 12 acute myeloid leukemias (AMLs), and 3 of 11 T-cell acute leukemias (T-ALLs). Expression of FLT3 RNA was also observed in some cases of blast crisis CML. The FLT3 signal resulted from expression on the leukemic blasts, and was not caused by increased FLT3 expression on normal CD34+ stem/progenitor cells in the leukemic samples. To determine if FLT3 protein was also overexpressed, proteins were extracted from leukemic BM samples and screened by Western blotting with anti-FLT3 antisera. FLT3 protein was not detected in normal BM controls, but was found in 14 of 14 B-lineage ALLs, 36 of 41 AMLs, and 1 of 4 T-ALLs. Stimulation of patient samples with FLT3 ligand resulted in autophosphorylation of the FLT3 receptor, suggesting the receptor is functional in these cells. These data show that FLT3 RNA and protein are aberrantly expressed by AML and ALL cells in that CD34 expression and FLT3 expression are no longer synchronous, and suggest the possibility that overexpression of FLT3 could play a role in the survival and/or proliferation of malignant clones in acute myeloid and lymphoid leukemias.

Antigens, CD34↗
PubMed1996

Metabolism of 5-methylchrysene and 6-methylchrysene by human hepatic and pulmonary cytochrome P450 enzymes.

The metabolism of environmentally occurring methylated polynuclear aromatic hydrocarbons by human cytochrome P450 (P450) enzymes has not been examined previously. We compared the metabolism of the tobacco smoke constituents 5-methylchrysene (5-MeC), a strong carcinogen, and 6-MeC, a weak carcinogen, in 18 hepatic and 11 pulmonary human microsomes. Major metabolites of 5-MeC were its proximate carcinogen trans-1,2-dihydroxy-1,2-dihydro-5-methylchrysene (5-MeC-1,2-diol), as well as 5-MeC-7,8-diol, bay region dihydrodiols, and phenols. 5-MeC-1,2-diol and 5-MeC-7,8-diol were formed stereoselectively, with the R,R enantiomers predominating. Major metabolites of 6-MeC were 6-MeC-1,2-diol, bay region dihydrodiols, phenols, and 6-(hydroxymethyl)chrysene. 6-MeC-1,2-diol was also formed stereoselectively in the 1R,2R configuration. All human liver samples formed the proximate carcinogenic 1,2-diols (0.2-2.3 pmol/mg protein/min for 5-MeC and 0.3-3.1 pmol/mg protein/min for 6-MeC). Comparable results were obtained in pulmonary microsomes, but the extent of metabolism was less than in the hepatic samples, and only 4 of 11 samples showed activity. Catalytic activities known to be associated with specific P450s were analyzed in each hepatic sample and correlated with levels of 5-MeC and 6-MeC metabolites in the same samples. The results of the correlation analysis indicated that P450s 1A1 and 1A2 were active in the formation of 5-MeC-1,2-diol and 6-MeC-1,2-diol, as well as several other metabolites resulting from ring oxidation. The formation of the hydroxymethyl metabolites was catalyzed by P450 3A4 (for 5-MeC) or P450s 3A4 and 1A2 (for 6-MeC). Experiments with chemical inhibitors and antibodies supported these results. The metabolism of 5-MeC and 6-MeC was also investigated using purified recombinant human P450s 1A1, 1A2, 2C10, 2D6, 2E1, 3A4, and 3A5. P450s 1A1, 1A2, and 2C10 had higher activities than the other enzymes for ring oxidation of 5-MeC and 6-MeC, whereas P450s 1A2 and 3A4 were more active than the other enzymes for methyl hydroxylation of 6-MeC. Only P450 3A4 showed substantial catalytic activity for methyl hydroxylation of 5-MeC. Collectively, the results of these studies demonstrate that P450s 1A2 and 2C10 are important catalysts of the metabolic activation of 5-MeC and 6-MeC in human liver, whereas P450 1A1 plays a major role in the metabolic activation of these compounds in human lung.

Antibodies↗
PubMed1996

Detection of DNA and globin adducts of polynuclear aromatic hydrocarbon diol epoxides by gas chromatography-mass spectrometry and -3H-CH3I postlabeling of released tetraols.

Gas chromatography-negative ion chemical ionization mass spectrometry--selected ion monitoring (GC-NICI-MS-SIM) was employed to detect tetramethyl ether derivatives of tetraols formed upon hydrolysis of DNA and globin adducts derived from diol epoxides of benzo[a]-pyrene (BP) and other polynuclear aromatic hydrocarbons (PAH). The tetramethyl ether derivatives could also be detected by [3H]CH3I postlabeling. The methodology involves the following steps: (1) isolation of DNA or globin; (2) mild acid hydrolysis under vacuum; (3) isolation of the resulting tetraols and derivatization to the corresponding tetramethyl ethers using methyl sulfinyl carbanion and unlabeled or 3H-labeled CH3I; (4) analysis by GC-NICI-MS-SIM or HPLC with radioflow detection. The optimum conditions for hydrolysis of adducts and derivatization of the resulting tetraols as well as the feasibility of this approach for detecting PAH adducts in mice and humans were explored. Using the set of four BP-tetraols that can be formed upon hydrolysis of adducts formed from r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (anti-BPDE) or r-7,t-8-dihydroxy-c-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (syn-BPDE) as models, the stability of the tetraols under the hydrolysis conditions was investigated. Adducts derived from anti-BPDE yield predominantly the stable r-7,t-8,9-c-10-tetrahydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (trans-anti-BP-tetraol), while adducts derived from syn-BPDE released cis-syn-BP-tetraol as a major hydrolysis product. Hydrolysis under vacuum significantly increased the recovery of tetraols. Conditions for derivatization of the BP-tetraols as well as tetraols derived from several other PAH anti-diol epoxides were investigated. Tetramethyl ethers proved to be superior derivatives that were stable, easy to prepare in high yields, and detectable with high sensitivity by GC-NICI-MS-SIM (1-50 fmol per injection). Alternatively, these derivatives could be detected by HPLC with radioflow detection if [3H]CH3I were employed for derivatization. The methodology was tested by comparing levels of DNA and globin adducts in mice treated with either unlabeled or 3H-labeled BP. Good agreement was obtained among the GC-NICI-MS-SIM, [3H]CH3I postlabeling, and conventional radiometric methods. Moreover, analysis of human hemoglobin by GC-NICI-MS-SIM resulted in detection of adducts derived from anti-BPDE and r-1,t-2-dihydroxy-t-3,4-epoxy-1,2,3,4-tetrahydrochrysene. The results of this study demonstrate that GC-NICI-MS-SIM of tetramethyl ethers of tetraols formed by hydrolysis of PAH diol epoxide DNA and globin adducts is a promising approach for detection and quantitation of adducts derived from a broad range of PAH.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1996

Effects of dietary fat content on the metabolism of NNK and on DNA methylation induced by NNK.

The available data support the concept that high-fat diets increase cytochrome P-450 activities in the liver, leading to increased rates of carcinogen metabolism and, in some instances, DNA adduct formation. Therefore we investigated whether a high-fat diet can also influence DNA methylation by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in the lungs of rats. Male F344 rats were fed a regular AIN-76A low-fat (5% corn oil) or AIN-76A high-fat (23.5% corn oil) diet. After three weeks on this dietary regimen, the animals were injected subcutaneously once daily for four days with NNK at 0.39 mmol/kg body wt. Groups of rats were sacrificed 4 and 24 hours after the last NNK administration; livers and lungs were excised for DNA isolation. We found that the high-fat diet significantly enhanced the formation of O6-methylguanine (O6-mGua) in the rat lung four hours (p < 0.01) after the last carcinogen administration. This may, in part, account for our previous finding in regard to the enhancing effect of the high-fat diet on NNK-induced lung carcinogenesis. There was no effect on O6-mGua or 7-mGua in the rat liver at either time point. To further elucidate the enhancing effect of the high-fat diet on DNA methylation by NNK in the lung, we determined its effect on the in vitro and in vivo metabolism of NNK. The in vitro data indicated that dietary fat has no measurable effect on liver and lung microsomal mixed-function oxidase in catalyzing the metabolic activation of NNK. The results of the metabolism study of NNK in vivo appear to be consistent with the in vitro finding, in that fat had no effect on the excretion pattern of NNK or on the distribution pattern of its urinary metabolites. It is apparent that the enhancing effect of the high-fat diet on O6-mGua in the lung of rats that was measured four hours after NNK injection requires future investigations.

Animals↗
PubMed1996

Benzo[c]phenanthrene is activated to DNA-binding diol epoxides in the human mammary carcinoma cell line MCF-7 but only limited activation occurs in mouse skin.

Benzo[c]phenanthrene (B[c]Ph) is an environmental contaminant with low carcinogenic activity in rodent bioassays. B[c]Ph-3,4-diol-1,2-epoxides (B[c]PhDE), however, are among the most tumorigenic diol epoxides known. To determine whether human cells are capable of activating B[c]Ph to DNA-binding metabolites, cultures of the human mammary cell line, MCF-7, were exposed to 10 microM B[c]Ph for 48, 72 and 96 h or to 1 microM (+/-)-B[c]Ph-3,4-dihydrodiol for 48 h. The B[c]Ph-DNA adducts were analyzed by 33P-postlabeling and reverse-phase HPLC. The major B[c]Ph-DNA adducts were formed by the trans-addition of (4R,3S)-dihydroxy-(2S,1R)-epoxy-1,2,3,4-tetrahydro-B[c]Ph to deoxyadenosine [(-)-B[c]PhDE-2dAt] and by the cis- and trans-addition of (4S,3R)-dihydroxy-(2S,1R)-epoxy-1,2,3,4-tetrahydro-B[c]Ph to deoxyadenosine [(+)-B[c]PhDE-1dAc and (+)-B[c]PhDE-1dAt]. Smaller amounts of the trans-addition of (-)-B[c]PhDE-2 were bound to deoxyguanosine. To determine whether B[c]Ph can be metabolically activated to diol epoxides in mouse epidermis, female SENCAR mice were treated topically with 2 micromol B[c]Ph for 24, 48 or 72 h or with 0.4 micromol (+/-)-B[c]Ph-3,4-dihydrodiol for 24 or 48 h. In B[c]Ph-treated mice, only small amounts of three B[c]PhDE-DNA adducts were detected [(-)-B[c]PhDE-2dAt, (+)-B[c]PhDE-1dAt and (+)-B[c]PhDE-1dAc] at 24, 48 and 72 h. In contrast, mice treated topically with 0.4 micromol (+/-)-B[c]Ph-3,4-dihydrodiol formed B[c]PhDE-DNA adducts at levels 6-fold greater than those observed with B[c]Ph at 48 h. The higher formation of B[c]PhDE-DNA adducts by (+/-)-B[c]Ph-3,4-dihydrodiol correlates with the greater potency of (+/-)-B[c]Ph-3,4-dihydrodiol than of B[c]Ph as a tumor initiator in mouse skin. The low extent of formation of B[c]PhDE from B[c]Ph in mouse epidermis may explain the low tumorigenicity of B[c]Ph in this tissue. These results indicate activation of B[c]Ph in mouse skin and tumorigenesis results in that tissue may not adequately assess the potential capability of cells from humans to activate B[c]Ph to ultimate carcinogenic metabolites.

Animals↗
PubMed1996

Bay-region distortions in a methanol adduct of a bay-region diol epoxide of the carcinogen 5-methylchrysene.

The three-dimensional structure of the product of the reaction of a diol epoxide of the carcinogen 5-methylchrysene with methanol has been determined by an X-ray diffraction analysis. The diol epoxide used to obtain this compound contains a stereochemically hindered bay region because of the location of the 5-methyl group, and this might be expected to affect the type of chemical reaction that occurs. The crystal structure analysis of this adduct of a polycyclic aromatic hydrocarbon (PAH) showed that a methoxy group has been added at the carbon atom of the epoxy group that is nearest to the aromatic system. The bond that is formed is axial to the ring system so that the carbon and hydrogen atoms of the methoxy group are considerably displaced from the PAH ring plane. The bay-region methyl group at position 5 is displaced out of the ring plane in the opposite direction. The major steric distortion in this methanol adduct is shown, by a comparison with crystal structures of related non-methylated compounds, to be in the area of the 5-methyl group and not in the tetrol-bearing ring. The steric effects that caused the axial conformation of the newly formed bond would also be expected to pertain in the DNA adduct of a PAH with a bay-region methyl group. Since the presence of the bay-region methyl group in 5-methylchrysene has been shown to enhance the carcinogenicity of this PAH over the parent compound or compounds with methyl groups in other positions of the molecule, it might be anticipated that this axial bond is found in carcinogenic lesions in DNA, and that any factor that ensures this axial conformation may accentuate the carcinogenic potential of a PAH of the appropriate size.

Carcinogens↗
PubMed1996

Effects of dietary 1,4-phenylenebis(methylene)selenocyanate on 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone-induced DNA adduct formation in lung and liver of A/J mice and F344 rats.

1,4-Phenylenebis(methylene)selenocyanate (p-XSC) was tested for its ability to inhibit DNA adduct formation induced by the tobacco-specific N-nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in the liver and lung of A/J mice and F344 rats. Dietary p-XSC, providing a dose of 5 p.p.m. selenium, significantly inhibited the formation of 7-methylguanine (7-mGua) induced by a single i.p. injection of 10 mumol of NNK(12.8% inhibition at 4 h and 19.9% at 96 h) and O6-methylguanine (O6-mGua) (16.5% at 4 h and 34.8% at 96 h) in the liver of A/J mice. Dietary supplements of p-XSC providing 15 p.p.m. of selenium reduced the levels of 7-mGua by 17.3% (4 h) and 33.6% (96 h). The formation of O6-mGua was inhibited by 69.5% (4th) and 73.8 (96h). In A/J mouse lung DNA the most significant reduction was observed in levels of O6-mGua. Dietary p-XSC at 5 p.p.m. as selenium inhibited the formation of this adduct by 73.1% (4 h). Ninety-six hours after NNK injection, and at both time points with p-XSC providing 15 p.p.m. selenium, O6-mGua was not detected. Although levels of 7- mGua in mouse lung DNA were also reduced, this was significant only 4 h after carcinogen administration. In general, selenite at a5 p.p.m. as selenium had no significant effect on the levels of these lesions; however, it inhibited O6-mGua in the liver only 4 h after NNK administration. These effects may explain why there is chemopreventive activity for p-XSC, but not for selenite, in NNK-induced lung carcinogenesis in A/J mice. Moreover, these findings raised our interest in determining the potential chemopreventive activity of p-XSC against NNK-induced lung adenocarcinomas in male F344 rats by first determining its effects on NNK-induced DNA methylation in the lungs of rats. Diet supplemented with 10 p.p.m. selenium as p-XSC did indeed inhibit the formation of adducts in pulmonary DNA of F344 rats treated with four consecutive injections of 81 mg/kg of NNK. Statistically significant inhibition of O6-mGua formation was observed 4 h after carcinogen treatment in both pulmonary (49.1% inhibition) and hepatic (39.8%) DNA. Statistically significant inhibition of 7-mGua formation was also measured in lung DNA isolated 24 h after the last NNK injection (45.0%) and in liver DNA 4 h after carcinogen treatment (31.8%). These results suggest that p-XSC would also inhibit induction of lung adenocarcinoma in male F344 rats by NNK.

Adenocarcinoma↗
PubMed1996

The essential role of the functional group in alkyl isothiocyanates for inhibition of tobacco nitrosamine-induced lung tumorigenesis.

The importance of the isothiocyanate group in alkyl isothiocyanate for inhibition of tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3- pyridyl)-1-butanone (NNK)-induce lung tumorigenesis was examined in A/J mice. Our previous structure-activity relationship study of isothiocyanates showed that 1-dodecyl isothiocyanate [CH3(CH2)11NCS], a simple alkyl isothiocyanate, is a potent inhibitor of NNK-induced lung tumorigenesis. It was chosen for this study due to its structural features and potency. A single dose of 1-dodecyl isothiocyanate given by gavage at 1 micromol/mouse 2 h prior to NNK administration completely inhibited lung tumorigenesis, while removal of the isothiocyanate group or replacing it with a hydroxyl group abolished the inhibitory activity. These results demonstrate that the isothiocyanate functional group is critical for the inhibitory activity of isothiocyanates in NNK-induced lung tumorigenesis. To gain more insights into the relationship of in vivo inhibition of tumorigenesis with the cytochrome P-450 enzyme inhibitory activity, the effects of these compounds on metabolism of NNK in mouse lung microsomes were studied. 1-Dodecyl isothiocyanate inhibited all three known oxidative pathways of NNK metabolism, with a stronger inhibitory activity toward NNK N-oxidation (IC50 430 nM) and keto alcohol formation (IC50 500 nM) than keto aldehyde formation (IC50 13,000 nM). 1-Dodecanol had a similar selectivity in inhibition of these metabolic pathways, but was less potent than 1-dodecyl isothiocyanate. Dodecane showed little or no inhibitory activity in the same concentration range. These results indicate that the isothiocyanate group of 1-dodecyl isothiocyanate is important for inhibition of NNK-induced lung tumorigenesis and also for effective inhibition of cytochrome P-450 enzymes involved in NNK oxidation.

Adenoma↗
PubMed1996

Site-specific adducts derived from the binding of anti-5-methylchrysene diol epoxide enantiomers to DNA: synthesis and characteristics.

The direct synthesis and characterization of site-specific adducts derived from the binding of (+)-1R,2S-dihydroxy-3S,4R-epoxide-1,2,3,4-tetrahydro-5-methylchrysene and the (-)-1S,2R,3R,4S-enantiomer [(+)- and (-)-5-MeCDE, respectively], to the N2-guanine residues in the oligonucleotide d(CCATCGCTACC) are described. The spectroscopic characteristics of the 5-MeCDE-modified oligonucleotides are discussed, and it is shown that their CD characteristics can be used to distinguish between the trans-addition products of the binding of the (+)- and (-)-enantiomers of 5-MeCDE (C4 position). The 11-mer duplexes with the normal complementary strands are destabilized by the site-specific, covalently bound 5-MeCDE residues: the melting points, Tm, are 5-10 degrees lower than in the case of the unmodified duplex. Stereoselective exonuclease enzyme digestion patterns of the single-stranded (+)- and (-)-trans-5-MeCDE-modified oligonucleotides (Mao et al, 1993, Biochemistry, 32, 11785-11793) were used to probe the orientations of the covalently bound 5-MeCDE residues relative to the modified guanine and the 5'-3' strand polarity; the aromatic residues are positioned either on the 5'-side [(+)-5-MeCDE], or the 3'-side [(-)-5-MeCDE adduct] of the modified guanine residues. The electrophoretic mobilities of the (+)-5-MeCDE-modified 11-mer duplexes in native polyacrylamide gels are slower than those of unmodified and modified duplexes containing the stereoisomeric (-)-5-MeCDE-N2-dG lesions. This indicates that the lesions derived from the tumorigenic (+)-5-MeCDE induce greater degrees of bending or local flexibility than the non-tumorigenic (-)-5- MeCDE enantiomer. These differences in the orientational and structural characteristics are similar to those observed with analogous DNA adducts derived from the tumorigenic (+)-7R,8S-dihydroxy-9S,10R-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene and the non-tumorigenic 7S,8R,9R,10S-enantiomer, respectively. The adducts derived from BPDE and 5-MeCDE enantiomers thus display similar characteristics that depend primarily on the PAH diol epoxide enantiomer stereochemistry. This direct synthesis approach can be used to generate milligram quantities of site-specific 5-MeCDE-modified oligonucleotides that are suitable for NMR studies (Cosman, et al., 1995, Biochemistry, 34, 6247-6260).

Base Sequence↗
PubMed1996

Inhibitory effects of 6-phenylhexyl isothiocyanate on 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone metabolic activation and lung tumorigenesis in rats.

This study examined the effects of 6-phenylhexyl isothiocyanate (PHITC) on lung tumorigenesis in F344 rats induced by the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). Two biomarkers of NNK metabolism, 4-hydroxy-1-(3-pyridyl)-1-butanone (HPB)-releasing hemoglobin adducts and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and its glucuronide (NNAL-Gluc) in urine, were also quantified during the course of the tumor induction experiment. Rats were divided into groups as follows: (1) NNK, 2 p.p.m. in drinking water, 60 rats; (2) NNK, 2 p.p.m. in drinking water and PHITC, 1 micromol/g NIH-07 diet, 60 rats; (3) PHITC, 1 micromol/g NIH-07 diet, 20 rats; (4) control, 20 rats. PHITC was added to the diet for 1 week prior to and during 111 weeks of NNK treatment. There were no effects of PHITC on body weight, mortality, blood chemistry or hematology. Seventy percent of the rats treated with NNK had adenoma or adenocarcinoma of the lung. In the rats treated with NNK plus PHITC, the total percent incidence of lung tumors was 26% (P < 0.01 compared with NNK). PHITC had no effect on the total incidence of exocrine pancreatic tumors induced by NNK. The rats treated with PHITC and NNK had significantly lower levels of HPB-releasing hemoglobin adducts throughout the course of the bioassay than did those treated with NNK alone and significantly higher levels of NNAL plus NNAL-Gluc excreted in urine at two time points during the bioassay. These results demonstrate that near lifetime administration of PHITC to rats strongly inhibits the metabolic activation and lung tumorigenicity of NNK.

Animals↗
PubMed1996

Synthesis of tobacco-specific N-nitrosamines and their metabolites and results of related bioassays.

Tobacco-specific N-nitrosamines (TSNA) are the most abundant, strong carcinogens in tobacco smoke. Seven TSNA have been identified in tobacco products: N'-nitrosonornicotine (NNN), N'-nitrosoanabasine (NAB), N'-nitrosoanatabine (NAT), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), 4-(methylnitrosamino)-4-(3-pyridyl)-1-butanol (iso-NNAL), and 4-(methylnitrosamino)-4-3-pyridyl)butyric acid (iso-NNAC). The syntheses of these compounds are reviewed. The syntheses of 14C- and 3H-labeled NNK as well as metabolites of NNK and NNN are also discussed. Comparative assays for lung tumorigenesis in female A/J mice were carried out for six of the TSNA and for two related compounds, N-nitrosodimethylamine (NDMA) and N-nitrosopyrrolidine (NPYR). They yielded the following ranking of potency: NDMA > NNK > NNAL > NPYR > NNN > NAB. Iso-NNAL and iso-NNAC were inactive. These results are also compared with previous assays of TSNA carcinogenicity in rats and hamsters.

Adenoma↗
PubMed1995

Ipomeanol analogs as chemopreventive agents: effect on the in vitro metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK).

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a tobacco-specific, powerful, organospecific lung carcinogen. 4-Ipomeanol (IPO) is an investigational chemotherapeutic drug with specific toxicity towards the lung. We hypothesized that non-toxic analogs of IPO could be competitive inhibitors of the metabolic activation of NNK. We had shown previously that 4-hydroxy-1-phenyl-1-pentanone (HPP) and 7-hydroxy-1-phenyl-1-octanone (4-HPO) are effectively inhibiting the lung tumor activity of NNK in A/J mice. In these extended studies we have synthesized 11 new analogs of HPP and tested them for their in vitro activities as inhibitors of the metabolism of NNK. The present study demonstrated that the lipophilicity in the molecule is playing an important role for the inhibition of NNK metabolism with pulmonary and hepatic microsomes of A/J mice.

Animals↗