Document O1dXBvk3OXOjw0JQyYZnLv6me

*C'>'.-. - H#. .1. * iH*rTtt/3^W: - r..I,'V' * '. .,* -* --r* ^1* **! Vi * * \-;`r *'. K-1711-(100) 1992 K-59G3-(100^ It., j vn> 1 yit ''f'V '- ;"W,{:' Wv'V..'-'- o rw X3*XS2&KZ&! mm. S5S 1^ '"" ' ' *vy. > ' * ."i*^'y,\y^Pv,; '?V ` ^ /- ' s * * , '>', v * * * Chem. Res. Toxicol. 1992, 5, 2-5 Communications Roles of the Vinyl Chloride Oxidation Products ^Chlorooxirane and 2-Chloroacetaldehyde in the in Vitro Formation of Etheno Adducts of Nucleic Acid Bases F. Peter Guengerich Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146 y Received August 12,1991 Vinyl chloride is of interest because of its involvement in the etiology of liver hem&ngiosarcoma and possibly other tumors in industrial workers (1). Similar tumors can be produced in animal models, and the vinyl halides are considered classic chemical carcinogens. Tumor initiation . by the vinyl halides and other vinyl monomers such as acrylonitrile and vinyl carbamate appears to involve initial oxidation to epoxides (2). In the case of the vinyl halides these halooxiranes rearrange rapidly to 2-haloacetaldehydes (Scheme 1). 'Both of these entities are reactive ` ; electrophilic species and can react with nucleic acid bases/' "Hie questiou'of which is most important has been con- ' sidered.j,Zajdela. et al- (3) found that 1-chlorooxirane ` caused-skin tumors'but-2-chloroacetaldehyde: did not,V Scheme I. Enrymttic Oxidation of Vinyl Chloride to l-Chlorooxirt&e and Rearrangement to 2-ChloroaceUldehyde DMA edduct* IM5G AA epoxide hydrolase alcohol dehydrogenase -v.-> \ :m , obtained as a SO* (w/v) aqueous solution hem Aldrich Chemical r^i ; Co. (Milwaukee, WI),` Vinyl chloride was purchased from MG, evidence'that 1-halooxiranes are the major electrophiles involved in DNA alkylation and that 2-haloacetaldehydes are the electrophiles most responsible for protein alkylation (5,< ` j photochemical ^chlorination of ethylene oxide in the preeence of tert-batyl hy-| pochlorite as deecribed elsewhere (5), and the concentration was' estimated using <-(p-ttItrobensyl)pyridine reagent (5, 18s))-----this fWl- -F/Spv 1 assay was also used in the estimation of the half-life. ATM2- OxoethyDguanin* was prepared by treatment ofguenosine with epichiorohydrin, \0,~ treatment, add hydrolysis, and preparative HPLC (19, 20); NVl-ethenoguonine was prepared jif' vdemonstrate the formation of the etheno adducts in vivo ,>, (S). However, there is now a considerable body of evidence /V indicating that these minor etheno adducts are probably (22). These latter three compounds were >98% pure asjudged by HPLC in the analytical systems and by reference of their // ufl"uoTMrescceennecee', lH NMR, and mass spectra with those hr the lit-.- i3 J1 m. I microscmeato electrophoretic hornogcueity as rteerribed elm whereJ (23).Hore'liver elcoholdehydrogenese^wes-purchssedfroma Sigmaend dialyzed before u*a\When liver microeosaeewere used! if as a source of P-dSO'ito'oiidiw'vinyl chloride? the nta'(male" i used in.tbe synthesis,of'these adducts (15) and 'as atwofisps Sprague-Dawley,' 160 g,"from Harlan Industries,'Indianapolis IN) ;'gates'forvinylhalides inDNAmodificationstudiefi(2,'iJ,'^^(ij'.weretre4ted with iaoniazid to induce'P;460,2El 5iI*2?,?,I^6I,6^,^^^^^^y^kein`lnehUe^ti^?daihndhiinbhitibioitnio'na'apmpr>6rdaacchh`eM8'uuseaded^^^"?A***y*vV^? conteining chloride ion ewe. '^Her/^^appU^spe^^^to'theque8tionJbf how| avoided to prevent the'poeaibility of nudeophilic-'attack,on 1^ ethenpodducUare'generated undMcbnditions''reemblingM thbee'encountored ihlcell^'^*l*!!sWi!esJ^^'^*^w'';''"'TM ^ehloroorirane (IS).tCalf thymus DNA'(2^ mg mL'1) was'incu/ 'Abated;with.l<hlorooxirane or 2-chloroecetsldehyde'in 0J.0 Mw 'potassium phosphate buffer'(pH7A) for IB.min at23.*CADNA`S ^^^^^%Eiq>erlmntafProcduresl^ .was'precipitated bytheaddition'of Bvolumes'of coldC^OH,? ^rocbveredbycentrifugation'at300Qf for.10 mitCdried under anl K<.f&C*ution!j-TftefoUowuig chemicals are fuuardous and should f; N,*'itresm,;diasolyed In HjQ.'and digested .with'0.1 NHC1 (70j ' behahdled^nefuUy\il<hlorb6xupnefuul2<hloroacetaldehydef^ i^C,;45 min) or emymaticelly [in GO mMTris-HCl bufler (pH &8)f .These,should'.be'handUd inth''`pntectwe^ehthingdrr a'weU-^S w containing 10 nM MgClJ with bovine pancrees DNase I [Sigma,! ventilated tm, h at 37 *C] end then a mlxture'm snakes Cheinirwls f# Cell thymus: DNAjyadenoeine'i^ltN^retheno^ |edah6sine.nd LAAethenodeoiyadendsine were' purchased fromi S Sigma Chemical Co. (St. Louh, MO).ti2-Chloroaceteldehyde waa'| !/.. ------------------------------ ` -------------------------- ---------------- - -1'- 4 V1 Abbroriaham GSK r-^utethioo--s--;-P'S~-t---0--,-n---daoem---a-a--l <7--t-o-d--u--o-m---e-5" 3 i3>S,JWMr! '..fi i' r.V ' -t Communications 50 40 Chem. Res. Toxicol., Vol. 5, No. I, 1992 3 1,N`.eihenedeozyedenoilne, nmol 20 40 60 80 100 120 1-Chlorooxirane, pmol ff*>Elhenoguanlne, nmol si Figure 1, Analysis of adducts formed by treatment of DNA with 1-chiorooxirane. Calf thymus DNA (10 mg) was treated with the indicated amount of l-chlorooxirane for 16 min in a total volume of 2.0 mL of 0.10 M potassium phosphate buffer (pH 7.4) at 23 *C. He DNA was precipitated by the addition of 10 volumes of cold C]H(OH, and adducts were measured as described under Experimental Procedures. The inset shows the same information with an expanded scale. All points presented are means from duplicate experiments: NM2-oxoethyl)guanine, ; 1,N*ethenodeoxyadenosine. A; A^^-ethenoguanine, . venom phosphodiesterase [Boehringer-Mannheim, Indianapolis, IN; 5 pg (mg of DNA)'1] and Escherichia coli alkaline phosphatase (Sigma, 3 units (mg of DNA)'1] for 12 h at 37 *C (25). In some of the incubations vinyl chloride (5000 ppm in head . apace) was oxidized by rat liver P-450 m the presence of adenosine (30 mM) or calfthymus DNA (2J> mg mL*1). DNA was separated by extraction with butanol and phenol solutions and recovered by C,H*0H precipitation prior to digestion (26). Recovery and enzymatic digestion were estimated by quantitation of deoxy- adenosine by HPIX) (AjjJ.'vide infra), r jarSS--' 1 A/LCZ-OxoethyOguanioe and A^^ethenoguanine were separated and quantified by HPLC and fluorescence measurements as described elsewhere (27). The procedure for the separation and estimation of ltAf*-ethenoadenosine used in this laboratory has been described previously (28,29). The same HPLC procedure was utilized for tbs analysis of l^Vfathenodeoxyadenosine, with reduction of the CH,CN concentration to 6% (v/v), and this separation system was also used to estimate deoxyadenosine recovery (the gradient system of Cirroussel et al. (25) was used in preliminary studies but did not appear to offer any advantage]. Reactant, junol Figure 2. Comparison of levels of formation of etheno adducts formed in DNA from 1-chlorooxirane and 2-chloroacetaldehyde. Calf thymus DNA (10 mg) was incubated with the indicated amount of reactant [1-chlorooxirane () or 2-chloroacetaldehyde ()] for 15 min at 23 *C in a total volume of 2.0 mL of 0.10 M potassium phosphate buffer (pH 7,4). He DNA was precipitated by the addition of 10 mL of cold CiHjOH, and adducts ware measured as described under Experimental Procedures. (A) lAi'-Ethenodeozyadenoaiae. (B) AP^-Ethenoguanine. All points presented are means from duplicate experiments. t'ij.H ' -1 ... 1 (epoxide hydrolase) or 2-chloroacetaldehyde (alcohol de-7 hydrogenase, plus NADH) were examined (5,6). Epoxide '" hydrolase almost completely blocked 1JV^-ethenoadenosme formation in both cases, but alcohol dehydrogenase had no effect. It was also found that tha addition of 10 mM GSH to the incubations containing adenosine and DNA (without added epoxide hydrolase or alcohol de hydrogenase) reduced l^-ethenoadenosine formation by ' 1. Sf.t,-*/ 76 and 61%, respectively.1 I 1 Discussion ^.^1<3UoWM^^Mai^:md'Ktedr^ti|,'calf ; The oxidation of vinyl chloride to 1-chlorooxirane and to 2^hloro^talddiyde DNA_vAnalvsis ofthe moducts indicated forma- - -Vvebeen known for some time (for review see ref j l-Chloroaxhane and 2-<&kroeoetzd<iehyde can showsimilar., ijSviadenoisme>, . gmutagenidtiea in some bacterial and mammalian'cellhi ^' eeththeennoo prro^duucctsts.,^, S3^eeiintheeaioK<x3yrMto8sminee;'aannad^ i^mexKr-:0~syeti^ ^ the question of which of theae'two^l tewwfyjri&nmreawt properhesandwere electrophilic halogen apedes is moat important in forma-fl Ofbiologically relevant DNA adducts has arisen.,WagMZd sWh?St?`r^rt>^^A^'fXDNAr.to^f<^^^`l^-|'0rieiliaIJy utilized two in vitro approaches to wmtaa ^..^^ethenoadenoame mdW,3;ethenoguamne!Mnsiderably>^''qUegtion of whether l-halooxinmes or 2-haloacetaldahydeai mote effectivelv than did 2-chloroacetaldehyde in exper-l>.&` ,,e m08t important (5,6): reaction Idnetica'can he mea-^ . sured directly with the chemicals, and epoxide^hydrolaseI`^^`^l *"AV1-(2-ox'oethyl)guar^,eJsfrom'. *rid ftlf'fiVinl dfthvdroffanAftA Tor aldehvdA dehvdroffimAAA^^W .consistent .with the;reported(A fo.L adduct8 <Rur" - Owprevioi work indicates#^^ m|MmpM:u3Ad) oNrA^oDsPlf.Hthyinm,tuhsq^pDreNsAen(cFeigoufreeitShBerl.^asdhadnol^eintehxEeignuwo-^ 'r-- - ^-J3le--ae& ^I1 1s^SS>S'f;iaTInrd, ebnoothdnceas'oer*l,tNhe*-etfhfeecnts6dOefoinxcyraedaeshinoegircieonwcaesnmtraMtisounrseOd.I^*V^R#c-h'*lo/r.o1a1 ix*i,ruaifnncel0o*tuo1rwbulhoryciikcerthsetundoieasdd(6u)ci;nt hdfooibrumitrialoytn.ioGonf,SDeHNltheAosubaifnhrdenainocgtt uwwsietshffoo1oc--tJ1j;j R&S150737 -r , R;"-, v 'l - . i'- ` jV;: f* , ^ * 4 Chem. Res. Toxicol., Vol. 5, No. 1, 1S92 Communications Figure 3. Effects of the addition of purified epoxide hydrolase and alcohol dehydrogenase on the formation of l^-ethanoadenosine adducts in systems containing rat liver microeomes and ' vinyl chloride. The indicated concentrations of purified rat liver , epoxide hydrolase () or horse liver alcohol dehydrogenase () ' - (plus 1 mM NADtfi were added to systems containing 0% (v/v) vinyl chloride gas in the head apace, 0.1 M potassium phosphate - buffer (pH 7.4), an NADPH-generating system, and either (A) 0.10 mg of liver microsomal protein prepared from isoniazidtreated rats and 30 mM adenosine (total liquid volume 0.12 mL and head space 3.9 mL) or (B) 1.0 mg of liver microsomal protein prepared from isoniazid-treated rats and 2.5 mg of calf thymus DNA mL"1 (total liquid volume 2.0 mL and head space 5.0 mL). In both cases reactions proceeded for 45 min at 37 "C and either (A) l,ft*-cthenoadenosine or (B) l/^-ethenodeaxyadenosine was measured os described under Experimental Procedures. All points are presented as means of duplicate experiments. pounds. It is known that the enzyme catalyzes the 0- deethylation of diethyl ether (35), and vinyl acetate might be readily hydrolyzed by esterases. It is unclear which of the etheno bases is most likely to be responsible for the tumorigenic effects of vinyl halides and other vinyl monomers. l,N*-Ethenoadenine, N3,4- ethenocytosine, and A^,3-ethenoguanine all appear to be formed at roughly similar levels in vivo (25,36). Differing results have been obtained with regard to miscoding in various in vitro polymerase fidelity experiments (10,11, 13, 14). While l^V^-ethenoadenosine has not been as dramatic as other etheno adducts in its miscoding prop erties in some of these assays, six of seven hepatomas raised in B6C3 FI mice treated with vinyl carbamate (which yields the same etheno adducts as vinyl chloride) showed an AT to TA transversion at the second position in the 61st codon of the rosH gene, which is probably consistent only with lr!V6-ethenoadenme as the mutagenic lesion (37). Most of the work done here focused on the use of IjA^-ethenoadenine as a prototypic adduct. How ever, the results also appear to apply to N^l-ethenoguanine (Figure 1), and selective 13C-labeling experiments indicate that the mechanisms of formation of 1/^-ethenoadenosine and 3,N4-ethenocytosine both involve attack of the basic endocyclic nitrogen of the pyrimidine ring (Nl of adenine or N4 of cytosine) on the unsubstituted methylene of the 1-halooxirane (32). Thus, the conclusions regarding the roles of 1-halooxiranes probably apply to all of the etheno adducts in DNA. , v Acknowledgment.- This work was supported in part `f by NIH Grants CA 44353 and ES 00267., I thank K. A. Atkins for synthesis of N7-(2-oxoethyl)guanine and N*,3- , ethenoguanine and M. V. Martin for technical assistance; in the purification of epoxide hydrolase. Registry No. 1-Chlorooxirane, 7763-77-1; 2-chloroacetaldehyde, 107-204); JV,-(2-oxo*thyl)guanine, 73100-87-5; IJN*ethenodeoxyadenosine, 68498-25-9; N*,3-ethenoguanme, 5628713-9; vinyl chloride, 75-01-4; epoxide hydrolase, 9048-63-9; alcohol dehydrogenase, 9031-72-5; 1^-ethenoadenosine, 39007-51-7. References that formation of A'7-(2-oxoethyl)guamne is probably also due to 1-chlorooxirane (Figure 1), and we previously con- (1) Forman, D., Bennett, B,, Stafford, J., and Doll, R, (1985) Ex posure to vinyl chloride and angiosarcoma of the liven a report of the register of cases. Sr. J. Ind, Med. 42, 750-763. .. . eluded that protein modification is the result of modifi- (2) Bolt, H. M. (1988) Roles of etheno-DNA adducU in turnon- . cation by 2-haloacetaldehydea (5,6). 'The reaction of 2- genicity of olefine. CRC Crit. Beo. Toxicol. 18,299-309. 1 '-0 : . . haloacetaldehydes with nucleic add bases is known to be .< : relativelyslow (6,' 22);'and the preferential reaction of the. ^*|^|baseswith l-halooxiranea can probably be attributed sim- ' ply to`their greater electrophilicity.- For further consid- . (3) Zajdela, F, Croity, A, Btrbin, A, Mslavsllle, C., Tomatis, L, . and Bartsch, H. (1980) Carcinogenicity of chloroethylene oxide,' .**, an ultimate reactive metabolite of vinyl chloride, sndbis(chloro- .. ' - methyl)ether after subcutaneous administration and in initia tion-promotion experiments in mice. Cancer Ree. 40,352-356. . -. 1 erationof the mechanism see Guengerich and Raney (32). (4) Gwinner, L M., Laib, R, J., Filser, J. G, and Bolt, H. M. (1983) ,V: 'l . -J^TheseJsbhduaiohs regarding the origin of the etheno de; rivatives are consistent'with the'lack of tumorigenicity of ` 2-chloroacetaldehyde'(3) and compounds such'as bis(2chloroethyl) ether which generate 2-chIoroacetaldehyde (4), . . v if indeed the etheno adducts are responsible for the bio- y.-. Evidence of chlorosthylsne oxide being the reactive metabolite of ' v-vinyl chloride towards DNA:' comparative studies with 2,2^-di. ' cblorodietbyletber. Carcinogenesis 4,1483-1486. (6) Guengerich, F. P., Crawford, W. M., Jr., and Watanabe, P. G. ` . (1979) Activation of vinyl chloride to covalently bound matabo-'. 1 lites: roles of 2-chloroethylene oxide and 2-chloroacetaldehyde. , , , '\-d'C logicaTeffectAl,2-Dibaloalkanes are tumorigenic.'but the ' Biochemistry 18, 5177-6182. ; ' mechanUm' involves GSH conjugation (30);: indeed, in- (6) Guengerich,'F. P,, Mason, Pi S., Stott, W. T, Kox, T. R.. and ,lihibitorsof P-450oxidation actually are potent cocaicino- . gensin'ratliver'^)^;^..-5-^ ;r ^ ^7v'^t.WeTcoh8idered'fthe',poesibility thatother4vinyl'com- .Watanabe, P. G. (1981) Roles of 2-halosthylens oxides and 2- '. haloacetaldehydes derived from vinyl bromide and vinyl chloride ' -i V in irreversible binding to protein and DNA Cancer Rts. 41,'- > : 4391-4398. 'TSS"1' ^^'pounds'bearing good.leaving groups might also be sub-. ' ``A'. (7) Laib, R. J.; Gwinhernl M.;'and Bolt, H. M. (1981) DNA al- vfel ^*tw*trat^fpr'P-450J2E$and''giveirise;toi etheno'adducts.' ' - kylatlon by vinyl chloride metabolites:' etheno derivatives or 7 ^si^Hiiweyer^heither viriyrsiwtate nor.'vinyl ethyl,ether did Jat;leyelsl>0.2 thatfaeeritwith^vinyl chloride* [nor did JltUethyl ether;which could conceivably be dehydrogenated fW.to vinyrethyl etherv(34)].4'Thua;:rat P-450 2E1 appears : alkylation of guaiuneT: Chem.-Biol. Interact. 37,219-231,s&fAi ( Singer. B,, and Bartach, H. (1986) The Role of Cyclic Nucleic iSizSK Acid Adducts in Carcinogenesis and Mutagenesis, 1ARC Srien-'';,* - -Qv - tific Publications, Lyon. ` 1'. ,- . - (9) Fedtks, N,, Boucheron, J. A, Walker, V. E., and Swsnberg, J. V-f ' A (1990) Vinyl chloride-induced DNA addueta. II. Formation - '% Chcm. Res. Toxicol. 1992, 5, 5-7 and persistence of 7-(2'-oxocthyl)guanine end N^S-eihenoguanine in rat tissue DNA. Carcinogenesis 11, 1287-1292. (10) Bsrbin, A., Leib, ft J., and Bartsch, H. (1985) Lack of mis coding properties of 7-(2-oxoethyl)guanine, the major vinyl chloride-DNA adduct. Cancer Res. 45, 2440-2444. (11) Singer, B., Abbott, L. G., and Spongier, S. J. (1984) Assessment of mutagenic efficiency of two carcinogen-modified nucleosides, lN**ethenodeoxyadenoeino and O4*methyldeoxythymidine, using polymerases of varying fidelity. Carcinogenesis 5, 1165-1171. (12) Kusmierek. J. T,, and Singer, B. (1982) Chloroacetaldehyde- treated ribo- and deoxyribopolynucleotides. 2. Errors in tran scription by different polymerases resulting from elhcnocytoaine and its hydrated intermediate. Biochemistry 21, 5723-5728. (13) Singer, B., Spengler, S. J., Chavez, F., and Kusmierek, J. T. (1987) The vinyl chloride-derived nucleoside, N*,3-etheno* guanoaine, is a highly eficient mutagen in transcription. Carci nogenesis 8, 745*747. (14) Singer, B., Kusmierek, J. T., Folkman, W., Chavez, F,, and Doaanjh, M. K. (1991) Evidence for the mutagenic potential of the vinyl chloride induced adduct, N\3-ctheno-deoxyguanosine, using a site-directed kinetic assay. Carcinogenesis 12, 745-747. (15) Leonard. N. J. (1984) Etheno-substituted nucleotides and co enzymes: fluorescence and biological activity. CRC Crit. Rev. Biochem 15,125-199. (16) Barbin* A., Friesen, M., O'Neill, I. K., Croisy, A., and Bartsch, H. (1986) New adducts of chloroethylene oxide and chloroacet- aldehyde with pyrimidine nucleosides. Chem.-Biot. Interact. 59, 43-54. (17) Bedell, M. A., Dyroff, M. C, Doerjer, G., and Swenberg, J. A. (1986) Quantitation of etheno adducta by fluorescence detection. In The Rob of Cyclic Nuclcie Acid Adducts in Carcinogenesis and Mutagenesis (Singer, B., and Bartsch, H., Eds.) pp 425-436, IARC Scientific Publications, Lyon. (18) Barbin, A^ Br6ziat, J. C., Croisy, A-, O'Neill, L K,, and Barlach, H. (1990) Nucleophilic selectivity and reaction kinetics of chip* roethylene oxide assessed by the 4-(p-nitrobeozyl)pyridine assay and proton nuclear magnetic resonance spectroscopy. Chenu-Biol. Interact. 73,261-277. .. (19) Roe, Jr., Paul, J. S., and Montgomery, P. 0., Jr. (1973) Synthesis and PMR spectra of 7-hydmxyalkylguanoiinium ace tates. J. HeterocycL Chem. 10,849-657. (20} Piper, J. R., Laseter, A. Gn and Montgomery, J. A. (1980) Synthesis of potential inhibitor! of hypoxanthine-guanine phos- phoribosyltransferase for testing as antiprotozoal agents. 1. 7- Substituted 6-oxopurinea. J. Med. Chem. 23, 357-364. (21) Sattsangi, P. D,, Leonard, N. J., and Frihart, C. R, (1977) IJVZ'Etheoeguanine and A^^-etbenoguainine. Synthesis and comparison of the electronic spectral properties of these linear and Angular triheterocycles related to the Y bases. J. Org. Chem. 42, 3292-3296. (22) Barrio, J. R., Secrist, J. A-, III, and Leonard, N. J. (1972) Fluorescent adenosine and cytidine derivatives. Biochem. Bio- phys. Res. Commmun* 46,597-604. (23) Guengerich, F- P-, Wang, P., Mitchell, M. B., and Mason, P. S. (1979) Rat and human liver microsomal epoxide hydratase. Pu rification and evidence for the existence of multiple forms. J. BioL Chem. 254,12248-12254. fcy:; , i- ;< (24) Thomas, P. E., Bandiera, S., Maincs, S. L., Ryan, D. E., and Levin, W. (1987) Regulation of cytochrome P-450j. a high-affinity N-nitrosodimcthylaminc demethylase, in rat hepatic microsomea. Biochemistry 26, 2280-2289. (25) Ciroussel, F., Barbin, A., Ebcrle, G-. and Bartsch, H. (1990) Investigations on the relationship between DNA ethenobase ad duct levels in several organs of vinyl chloride-exposed rats and cancer susceptibility. Biochem. Pharmacol. 39, 1109-1113. (26) Kadlubar, F. F., Miller, J. A., and Miller, E. C. (1976) Micro somal N-oxidation of the hepatocarcinogen N-methyl-4-aminoazobenzene and the reactivity of N*hydroxy-N-methyl-4-aminoazobenzcnc. Cancer Res. 36, 1196-1206. (27) Fedtkc, N,, Walker, V. E., and Swenberg. J. A. (1989) Deter mination of 7-(2'*oxoethyl)guanine and N*.3-ethenoguanine in DNA hydrolysates by HPLC. Arch. Toxicol. Suppl. 13, 214-218. (28) Leithauser, M. T., Liem, A., Stewart, B. C.. Miller, E. C.. and Miller, J. A. (1990) l,N*-Ethenoedeno6ine formation, mutagenicity and murine tumor induction as indicators of the generation of an electrophilic epoxide metabolite of the closely related carcinogens ethyl carbamate (urethane) and vinyl carbamate. Carcinogenesis U, 463-473. (29) Guengerich, F. P., Kim, p.*H., and Iwasaki, M. (1991) Role of human cytochrome P-450 1I&1 in the oxidation of several low molecular weight cancer suspects. Chem. Res. Toxicol. 4.168-179. (30) Kim, D.-H., and Guengerich, F. P. (1990) Formation of the DNA adduct 5*(2*(N7*guanyl)ethyl]glutathione from ethylene dibromide; effects of modulation of glutathione and glutathione S-transferaae levels and the lack of a role for sulfation. Carcino genesis 11, 419-424. (31) Malaveille, C., Bartsch, H., Barbin, A., Camus, A. M., and Montesano, R. (1975) Mutagenicity of vinyl chloride, chloroethyleneoxide, chloroacetaldehyde and chloroethanoL Biochem. Biophys. Res. Commun. 63,363*370, (32) Guengerich, F. P,, and Raney, V. M. (1991) Formation of etheno adducts of adenosine and cytidine from 1-halooxiranea. Evidence for a mechanism involving initial reaction with the endocydic nitrogen atoms. J. Am. Chem. Sac. (in press). (33) Wong, U C. 1C, Winston, J, M.,'Hong, C. B., and Plotnick, H. (1982) Carcinogenicity and toxicity of l^-dibromomethane in the rat. Toxicol. Appl. Pharmacol, $3,155*165. (34) Guengerich, F. P., and Kim, D.*H. (1991) Enzymatic oxidation of ethyl carbamate to vinyl carbamate and its role as an inter* mediate in the formation of l,N*-ethenoadenoeme. Chem. Res. Toxicol. 4.413-421. (35) Brady, J. F., Lee, M. J., Li, M., Ishizaki, H., and Yang, C. S. (1988) Diethyl ether as a substrate for acetone/ethanol-inducible cytoclirome P-450 and as sn inducer for cytochrome(s) P-450. Mot. Pharmacol, 33,148-154. (36) Eberie, <i., Barbin, A., Laib, R. J., Ciroussel, F., Tbomale, J., Bartsch, H., and Rajcwsky, M, F. (1969) lJVt'etheno-2/-deoxy- adenosine and SpN^etheno^-deoxycytidine detected by mono clonal antibodies in lung and liver DNA of rata exposed to vinyl chloride. Carcinogenesis 10, 209-212. (37) Wiseman, R, W., Stowers, S. J,, Miller, E. C., Anderson* M. W., and Miller, J. A. (1986) Activating mutations of the c-Ha-roa protooncogene in chemically induced hepatomas of the male B6C3 Fl mouse. Proc. Natl. Acad. Sei. UJSjL 83, 5825*5829. R & S 150739 Semiemplrical Self-Consistent Field (CNDO) Calculations of 7 Arsenical-Antidote Adducts Dennis W. Bennett,1 Lihua Huang,* and Kilian Dill*-* Department of Chemistry, University of Wisconsin, Milwaukee, Wisconsin 53201, and Department of Chemistry, Clemson University, Clemson, South Carolina 29634 Received August 8,1991 1 - .OX'-. '-..T ' Introduction . !f A'requisite feature of antidotes for heavy metals such m-t, i arsenic is the ability of the antidote to sequester the - . <* --A`* , /. .v i" qti ^ v. 9 Author to whom correspondence should be addressed. * University of Wisconsin. ^ * Clemson University. ; (Vf%' *"'* ," rV.-Nk " ' ' 0893-228i/92/2705-0005103.00/0 metal and eventually excrete the adduct. Thus, the pharmacokinetics of the antidote and the stability of the adduct formed are of utmost importance as the first step in the detoxification process. In recent years, we have published extensively on the reactions of organic arsenicals with simple thiol-containing compounds in order to develop a strategy in the search for 1992 American Chemical Society < < V' Identification and Quantitation of BP-DNA Adducts (12) Jankowiak, R, and Small, G. J. (199}} Fluorescence line nar rowing: A high-resolution window on DNA and protein damage from chemical carcinogens. Chctn. Res, Toxicol, 4, 266-269. (13) NIH Guidelines for the Laboratory Use of Chemical Cardnogens (1961) NIH Publication No. 61-2385, U.S. Government Printing Office, Washington, DC. (14) Bodell, W. J,, Devanesan, P. D>, Ro"^n, E. G., and Cavclieri, . L. (1989) S2P-Postlabeling analysis of benzo[a]pyrene-DNA adducts formed in vitro and in vivo. Chem. Res. Toxicol. 2, 312-316. (15) Zamxow, D., Jankowiak, R, Cooper, R S., Small, G. J.t Tibbcla, S. R, Cremonesi, P., Devuneaon, P., Rogon, E. G., and Cavolicri, Chcm. Bes. Toxicol, Vol 5, No. 2, 1992 309 E. L. (1989) Fluorescence line narrowing spcctrometric analysis of benxofaJpyrewHDNA adducts formed by one-electron oxida tion. Chem. Res. Toxicol. 2, 29-34. (16) Lu, P,, Jeong, H., Jankowiak, R., Small, G. J., Kim, S. K., Gasman, M., and Geacintov, N. E. (1991) Comparative laser spectroscopic study of DNA and polynucleotide adducts from the (+)-4ffli-diol epoxide of bcno(ajpyrenu. Chem. Res. Toxicol 4, 58-69. (17) Jeong, H. (1991) Ph.D. Dissertation, Iowa State University. (18) Sage, E., and Hoseltine, W, A. (1984) High ratio of alkali-sen sitive lesions to total DNA modification induced by benzo(o)pyrene diol epoxide. J, Biol Chem. 259,11098-11102. Additions and Corrections Roles of the Vinyl Chloride Oxidation Products 2Chlorooxirane and 2-Chloroacotaldebydc in the in Vitro Formation of Etheno Adducts of Nucleic Acid Bases [Volume 5, Number 1, January/February 1992, pp 2-5). F. Peter Guengerich Page 2. The title should read Roles of the Vinyl Chloride Oxidation Products 1-Chlorooxirane and 2-Chloroacetaldehyde in the in Vitro Formation of Etheno Adducts of Nucleic Acid Bases. ft- <I*'*' <i.V nr v ^' 1 V'.v- & , Ji. '. ,-xfSW' h ..yv (h %