- Email: [email protected]
Toxicology of mustard gas
TiPS - April 1991 [Vol. 221
lb-t
Toxicologyof mustardgas Uri Wormser TIWdt~t~astatirrg effeds ofmustardgaswere Jirsl observed in World War I. The advent of the Gulf War fueled renewed fears of @the; use of toxic gases in battle, with the possible exposure of large civilian populations - while understnndi,tg of the mechanism of action of the alkykylatingsulfur mustards was sM1 quite restricted. In this arlicle Uri Wormser discusses the structureactivity studies f/rat are available, and the limited pharmacological measures that can be taken 10 protect against mustard gas attack. In addition to systemically administered sulfhydryl agents, new percutaneous preparations are being developed in the autllor’s laboratory which offer better protection tlrau is possible with simple adsorban: powders.
Mustard gas (sulfur mustard) is one of the most powerful vesicants used as a chemical weapon. Unlike the organophosphate nerve gases which instantly kill, mustard gas has delayed effects, and incapacitates far more pmple than it kills. Large amounts of the gas can rapidly be prepared for battlefield use because of the simplicity of its chemical synthesis. Mustard gas was employed for the first time in World War f in 1917 near Ypres, Belgium. This early use earned the gas two nicknames: Yperite, and Yellow Cross (due to the special sign on mustard gas-containing shells). Several exposures have since occurred. Following World War II, large stockpiles of chemical weapotls including mustard gas were dumped in the Baltic sea. Corrosion of the containers led to the liberation of shelfs which were accidently brought on board by fishing trawlers, resulting in exposure of fishermen to the vesicant’. Mustard gas was used by Iraqi forces during the Iran-Iraq conflice; the Iranian soldiers who were evacuated for treatment in hrrmpe showed classical symptoms of mustard gas poisoning? The possibility of large scale exposure of military and civilian populations to mustard gas must once again be seriously considered. This review describes the pathological effects of the vesicant, its cellular mechanism of action, and the available antidotes for prevention of skin bums caused by mustard gas.
Despite its name, mustard gas is a volatile oily liquid, slightly soluble (0.07%) in watefl, which rapidly penetrates human skin within 30 minutes of exposurd. Saturated vapor of mustard gas is absorbed at a rate of 1.4 pg cm-* min-’ at 22°C (Ref. 6). Most of the liquid form of mustard gas applied to the skin evaporates (80%) and of the 20% that penetrates, half is fixed in the skin and half is absorbed systemically’. Pathological effecta of muabrd gas lesions in humans Although there are great discrepancies in the sensitivity of individuals to mustard g&, the following description, based cn controlled human studies performed with 143 gas mask-protected volunteers in 1949 (Ref. 9), may be a typical picture of skin lesion development after exposure in times of emergency.
Unlike heat bums the typical chari&&r,ic of mustard gasinduced skin bums is its delayed effect. These start to appear at least several hours after poisoning. Exposure of humans to mustard gas vapor resulted in erythema which developed within 24 hours in the majority of subjects. In mild cases it was delayed for 3-4 days and in severely affected persons it appeared within 4 hours. In most cases the erythema gave place gradually to the pigmentation characteristic of mustard gas bums. Subcutaneous oedema was detected 3-9 days after exposure depending on the region of the body. Vesication of the skin occurred later, appearing as numerous blisters about the size of a pin’s head or smaller, and sometimes developing into large blisters. In most cases the mean time for first onset of v&cation in the various regions of the body was 7.2 daya; blistera invoked superficial layers which healed in a few days. By contraat, in severe cases (11% of the exposed individuals) deep veaicka appeared during the first four daya. Deep veaication or deaquamation produced breaches tf the epithelium surface leading to painful raw surfaces which were the main portals of infection. The mean time for the first appearance of raw surfaces was 8.8 days while maximal severity of the lesiona was obuemed after a mean of 10.9 days. The average time for healing was 19 days, while in severe cases it
EPS - Apriil1991 /Vd. 121 could lengthen to months, particularly on the penis. Mustard gas vapor differentially affected various regions of the body. The most severely injured organs were the wet and warm areas, i.e. scrotum, axillae, penis and buttocks. The trunk, dorsum of hands (palms and soles were rarely affected) and forearms were also frequently injured to different degrees of severity9. Additional delayed symptoms such as epigashic distress and vomiting, appearing hours after exposure were ohsemd in soldiers attacked by mustard gas during World War I (Ref. 10). Occnlar damage such as conjunctivitis, lacrlmation, photophobia and eyelid oedema occurred in most of the affected individuals, while in severe cases cornea1 involvement, iritis and chemosis also appearedr”. The respiratory system was also highly vulnerable to mustard gas vapor. LaryngItls and bronchitis were primarily irritative and in Severe cases were complicated, 3-4 deys after exposure, by bronchopneumonia or pulmonary o&ma. The fatal cases (2-5s of exposed individuals) following ~~m~;;q~;~$;$?{ gone marrow toxicity was recorded in the most severely affected soldiers2 as well as in accidental exposure’*. The main long-term effect of mustard gas is its tumorigenic activity. The mutagenic and carcinogenic potential of the vesicant was proven by in vitro and in viuo experiments, as well as by epidemiologlcal studies’“. A 4%year follow-up of workers in British mustard gas manufacture during World War I1 has shown that the death rate from respiratory tract malignancies was higher (up to 5.5 times) than that of the onexposed populationr4. An even higher incidence of lung cancer (up to 35-fold) was observed in empIoyees in Japanese and German mustard gas factories”. Duration of employment significantly affected risk for certain cancers”. High mortality from chronic nonmalignant respiratory diseases was also observed”. Long-term keratopathy, including deposition of hyalin, Ca2* and crystals, was also detected 35-50 years after exposure to mustard gas during World War 1t5.
Cl-CH,-CH,-S-C&-CH,-Clmustard gas OH-C~J~~~-S-CH~-CH,-OH
h,,,lysis/ drtoxif&i&l
ion r-p
1 conjugation
excretion Cl-CH,-CH,-S-Cl-$-CH,-R, mono-alkylaticn R,-CH,-CH,-S-Ct$-CH,-R, di-alkylation
DNAALKYlAltoN
DNA,R?4AandPROlEtNALKYUVON
CELLUIAR PAwoLoQY
GROSS PAlMoLoGv
carcinogenicity
inhibition of DNA RNAand
pmtein synthesis, dwomatid aberrations, mduoed glycdysii, NADdepletion
erytherna
t basacelldmage, localewtraveaatkn, leukocyte
krfiltration,relsased inflammatorymediators
o&ma
J Miter formationand desquamation
Mechanismof action of mustard
gas It is established that the key reaction of mustard gas is the intramolecular cyclization to form the electrophilic ethylene episulfonium intermediate and the liberation of free chloride anion $I=PFig. 1). The conversion to this reactive deri-rative is temperature dependent”’ and is facilitated in the presence of aqueous solution’. This fact may explain the high vulnerability of warm and wet regions and the mucosal tissues of the eye and respiratory tract to mustard gas. Similar chemical cyclization occurs with the antineoplastic drug nitrogen mustard
(mechiorethamine) to form the ethylene immonium derivative”. The cyclic -onium intermediate alkylates the nucleophilic residues of macromolec4es. The major alkylation site of nucleic acids of mammalian origin is the N-7 of the guanine residue“‘. The second chloroethyl moiety of the mustard can attack additional I%7-guaninr to create inter&and” and intrastrandm crosdinklng. Additional targets for alkylation are N-3and b-g~anfne~‘-~. adeninc aberrations TllUS, chromatid occt~~‘*~,synthesis of DNA, RNA and proteins is inhibited and cells are blocked at the interfaceof the C&/Mphases of the cell cyclers.
TiPS-April 1991[Vol. 121
166
The sulfoxide form of mustard
TABLE I. V~!dMng activityd IWSWd geSdSrivatiVSS coAIpound-----
Rob
VW ;+
VI VII
s(WCICHC!,)(CH,CH,CI) 6(CHClCHCM* 6(CHClCH,)P
VM
S(CH2CHZCHzCl)z s(CHBcHd(W,CHzCI) .%CaHJ(cH2CH2cI)
+.++’ ++ +
XI XII XIII
s(CycsHs)(CH&HzCI)
XIV XV
02s(CH2CH2cl)z OS(CH=CH&
+
XVI
og5(CH=CH&
+
os(cH?cHzC1)2
6.36,*1 41 17.36 41 41 41 17 ii 6 I7 6 17.37 37,41 17 17
++,slmng;+,moderate:-,t~vesicanl.’extentdvesicstingectMtynotde~
Nucleic acids are not the only target macromolecules attacked by sulfur mustard. Inhibition of various glycolytic and respiratory enzyme@ and impaired glucose uotake2b have also been observed f&owing exposure t0 sulfur mustard. Gross et nl. have shown reduced NAD+ levels after sulfur mustard exposure in human skin grafted to athymk nude mi@. This phenomenon has also been observed in other biological systemP2. In human keratinocyte cultures, prevention of NAD+ depletion did not reverse glucose uptake inhibition, which !Y a measure for cellular viabilip, mdicating that reduced NAD+ levels could be one of several mechanisms of sulfur mustardinduced cell injury in human skin. Dannenberg and coworkers developed an organ culture from rabbit skin exposed in vitro to mustard garP. This ex oiuo system was used to follow the inflammatory processes accompanying the various stages of sulfur mustardinduced skin lesions, including migration of polymorphonuclear, basophil and mononuclear teukocytesm, release of proteinases3’ and proteinase inhibitor!?*, and entry and turnover of serum proteins in developing and healing of skin lesionsm. Despite the extent of the nonspecific cellular processes impaired by its potent alkylating activity it appears that sulfur mustard primarily disrupts the normal homeostasis of cell proliferation particularly of the germinative (basal) cells of the ski#, resulting in separation of the epidermis from the dermis and blister formation.
Structu~vesicatlng activity relationships The key reaction of sulfur (~,fl’-dichlorodiethyl mustard sulfide) involves intramolecular cyclization to form the electrophilic ethylene episulfonium derivative. Compounds that do not produce this intermediate (see Table I) either because of the substitution of chloromethyl moieties (compound II) for chloroethyl groups (I), or replacement of Cl- by hydroxyl groups to form the urinary metabolite dihydroxydiethyl sulfides’ (Ill), have no vesicating activity. Similarly the introduction of a Cl- into the aposition of either one arm of the difunctional sulfur mustard (IV, V) or both arms (VI, VII) produced non-vesicating compounds. This might be a result of reduced electron density of the sulfur atom and decreased intramolecular cycliition. The presence of shlorlde in the y positions of the dichlorodipropyl sulfide derivative (VW) eliminated its blisterogenic activity whereas the S,@‘-dlchlorodipropyl sulfide (lx) was bund to have comparable activity to that of mustard gasas. In spite of its single alkylating group, chloroethykthyl sulfide (X) was shown to have strong veelcating actlvlty because of its rapid penetration of the skin, probably due to the presence of lipophilic moiety. Its penetration is more rapid than compound I (Ref. 6). Another monofunctional sulfur mustard in which the ethyl was replaced by a phenyl group (Xl) had blis nit activity, but introduction“gp0 benzyl moiety (XII) caused the molecule to be inactive.
gas (XIII) had no toxic activity. However, the sulfone mustard (XIV), whose conjugated form was
found as a urinary metabolitr, did act as a vesicant although less potently than sulfur mustard itself. Although neither compound is able to cyclize to form the isulfonium ion it electrophilic was assumed 2 that they are converted in vioo to the corresponding non-active divinyl &oxide (XV) and to the toxic vesicant divinyl sulfone (XVI), respectively. The possibility of divinyl sulfone activation via epoxide formation cannot be excluded. These assumptions need to be further investigated. Neutralizing agenln for mustard grs The ele&ophilic nature of the ethyleneepi&onium intermediate can be neutralilntd by nucleophilic compounds. Sulfhychyl agents such aa dlthiothwitd
and
mercaptoethylamlne reduce sulfur mustard toxicity in in titro systemP. In in ufuo studies in fmimalt3 sodium thlosulfate’ combined with various drugs such as cortlcosteroids, antlmddanb, anticoagulants and antlbistunines was able to prevent, to smno extent, mustard gas t&cl@‘. HowS ever, neither these amt nor numerous others7 JFZ? completely protective against systemic or topical toxic&y of mustard gas. Skin lesions caused by percutaneous appliattion of sulfur mustard can be prevented by adsorbent powders such as Fuller% earth, charcoal and t&urn* provided they are used within 10 minutes of exposure. Recently we have patented a non-toxic and non-irritant iodlne/povidoneiodine-containing preporation which efficiently protects guineapig skin from must& gas even when applied 20 minutes after exposu#. Preliminary results of similar newly developedprepar. ations obtained by us indicate that even longer intervals such as 40 and 60 minutes between exposure and treatment achieve high degrees of protection against mustard gas. cl
cl
cl
Although mustard gas has been known for decades as a powerful
TiPS - April 1992 [Vol. 121 chemical weapon to which mass populations might be exposed, understanding of its mechanism of action is quite limited. A combination of detailed structureactivity zrudies with inves’igation of the molecular and cellular pathology of mustard gas may lead to practical solutions for the current military medical problems.
References 1 Aasted, A., Dane, E. and Wulf, H. C. (19S7)Ann. Plasf. surg. 19, X30-333 2 Suhrabpuur, H. (1987) Med. J. Islamic Rep. irau 1,32-37 3 Requena, L. et al. (1988) J. Am. Acad. Dcmutol. 19,5=6 4 Hopkins, E. F. (1919)J. PJ~anacoI. Exp. 77Icr.lz35?uu3 5 Culltunbine, H. (1946) Br. 1. Dmalol. 5B,291-294
6 Nagy, S. hi., Golumbic, C., Stein, W. H., Frutun, J. S. and Bergmann, M. (1946) /. Cen. Physfol. 29,441&9 7 Sumani, S. M. and Babu, S. R. (1989) 1. C&I. Pharatacal. Thcr. Toxicof. 27, 419-435 8 Mamhali, E. K., Lynch, V. and Smith, H. W. (1918)J. Pharmrcul.Exp. Thcr. 12, 291301 9 Sfndair, D. C. (1949) Br. I. Derwtol.
Six volumes on medicinal chemistry Comprehensive Medicinal Chemistry edited by Cotwin Hansch, Pergamon Press, 1990. $1995.00/E1145.00 kii + 5494 pages in six volumes) ISBN 0 08 032530 0
It is not insignificant that the first chapter of Comprehensive Mcdicinal Chemistry, written by a team of over 250 authors with Corwin Hans& chairing the board of editors, comes from Alfred Burger. Indeed, this book may be regarded as a successor to Burce; Medicinal Chemistry, ’ appeared in l%O. In 1980 three volumes sufficed, but now an impredvese&es of six volumes is needed in order to be comprehensive. The new book contains a wealth of information but also has some drawduplications occur; some subjects are treated in dif-
167 Syphilis 61,113-125 10 Derby, G. S. (1918)Am. I. Med. Sri. 156, 733-736
11 Mandel. M. and Gibson, W. S. (1917) J. Am. Med. Assoc. 69,1978-1971 12 Hobbs, E. 8. (1944)Br.Med. j. 2,306-y17 13 IARC Monographs (1975) Vol. 9, pp. i81-i92, WHG, Geneva 14 Easton, D. F., Pete, J. and Doll, R. (1988) Br. 1. Ind. Med. 45.652-659 15 Blodl, F. C. (197l) InL Ophthalmol. Clin. ll, l-13 16 Ward, J. R. and Seiders, R. P. (1984) Thcrmochim. Acfa 81.343-348 17 Dixon, hi. and Needham, D. M. (1946) Nature 158,432-438 18 Wheeler, C. P. (1962) Cancer Rts. 22, 651-688 19 Ball, C. R. and Roberts, J. J. (1972) Chrm-tliol. Interact.4.297303 20 Walker, 1. C. (197l) Con. 1. Biochcm.49, 332-336 21 Ludhun, D. B., Kent, S. and Mehta, J. R. (1986)Carcinogenesis7,1203-12% 22 Habraken, Y. and Ludhun, D. B. (1989) carcfnogcnesii 10,489-492 23 Kirchner, M. and Brendei, M. (1983) Chem-Biol. Interacf. 44, n-39 24 Savage, J. R. K. and Breckon, C. (l%l) Mutat. Rn. 84,37u87 25 Rub&s, J. J., Friedlos, F., Scott, D., Cbmerud, M. C. and Rawlings, C. 1. (1986)Mulot. Res. 166,169-181 26 Md, M. A. E., Van De Ruit, A. M. 5. C. and Kluivers, A. W. (1989)Toxicol.Appl. Pharmacol.98,159-165 27 Gross, C. L,, Meier, H. L., Papirmeis~er,
ferent volumes, which makes it difficult to get complete information quickly; the style differs substantially tinm chapter to chapter but, more importantly, the difkrent subjects are not all pitched at the same level. The presentation of the book is excellent, but this is to be expected as the price of the book is extraordinary. Very few printing errors appear throughout the book. In the prefaace of the book the aim is presented as ‘. . . to present the subject, the modem role of which is the understanding of relationships structure-activity and drug design from the mechanistic view-point, as a field of its own right, integrating with its central chemistry all the necessary ancillary disciplines’. The first volume covers general principles. Several chapters are very general and offer familiar information, and the presentation makes the volume very useful for educational purposes (particularly the chapter by Sneaders on the
B., Brinkley, F. B. and Johnson, J B. (1985) Toxicof. Appt. fharmoml. 81, 85-90 28 Petrali, I. P., ogksby, S. 8. and Meier, H. L. (1990) Ultrostruct. Parhol. 14. 253-262 29 Ku, W. W. and Bernstein, I. A. (1988) Toxicol.Appl. Phermacol. 95,397+1 30 Darwten& A M. Jr et al. (1985)AM. 1. Pufhol. 121.15-27 31 Higuchi, K. et al. (1988)lnflummrtion12, 311-334 32 Harada,5. et al. (1987)Am. 1. Pathol.l26, 148-163 33 Handa, S. et al. (1985)Am. 1. Pathol. 121, 28-38
34 Davison, C., Rozman. R. S. and Smith, P. K. (l%l) Biochem. I%wmucol. 7,65-14 35 Lynch. V., Smith, H. W. and Marshall, E. K.Jr(1918)1. Pharmacol.Exp. ntr. 12, 265-290 # Renshaw, B. (1947)1. Invest. Dennatol. 9, 7S-85 37 Banks, T. E., BoursnelL J. C., Francis, C. 8.. Hupwuud. F. L. and Wonnall, A. (1946)Biochrm.J. 40,734-736 38 Walker, I. G. and Smith, J. F. (1969)Can. J. Physiol. Pharmaco~.47,143-151 39 Vojvudic, V., Milusavljwic, Z., Buskovic, B. and Bojanic, N. (1985) Fundurn. A@. ToticoI. 5, S165S168 40 Sohnan, T. (1919) J. Pharmrcol. Exp. Ther. 12,303-318 41 Peters, R. A. and Walker, E. (1923) Biochm. I. 17,~W6 42 Wonnser, IJ. (1990) Israeli Patent No. 95087
chronology of drug introductions and the chapter on sources of information). The chapters on physiological aspects are very condensed, but informative. Some of the chapters are too limited (even superficial) to be of interest (e.g. the chapter on selectivity by Lipnick, which seems to be an abstract of Albert’s famous book). Too much attention is paid to aspects of recombinant DNA technology procedures. Information on organization and funding of medical research and on healthcan? systems is included, but only for the USA and UK. The next four volumes treat medicinal chemistry subjects impressively and concord with the promises made in the preface. Volume 2 covers enzymes and other molecular targets; a lively, factua! and fascinatig account but again there is variation in presentation, e.g. treatment with fi-lactams begins with chemical aspects, while in another case biological mechanisms open the chapter. Volume 3. Membranes and Receptors, also shows remarkable differences between chapters; the information on p-adrenoceptors
lb-t
Toxicologyof mustardgas Uri Wormser TIWdt~t~astatirrg effeds ofmustardgaswere Jirsl observed in World War I. The advent of the Gulf War fueled renewed fears of @the; use of toxic gases in battle, with the possible exposure of large civilian populations - while understnndi,tg of the mechanism of action of the alkykylatingsulfur mustards was sM1 quite restricted. In this arlicle Uri Wormser discusses the structureactivity studies f/rat are available, and the limited pharmacological measures that can be taken 10 protect against mustard gas attack. In addition to systemically administered sulfhydryl agents, new percutaneous preparations are being developed in the autllor’s laboratory which offer better protection tlrau is possible with simple adsorban: powders.
Mustard gas (sulfur mustard) is one of the most powerful vesicants used as a chemical weapon. Unlike the organophosphate nerve gases which instantly kill, mustard gas has delayed effects, and incapacitates far more pmple than it kills. Large amounts of the gas can rapidly be prepared for battlefield use because of the simplicity of its chemical synthesis. Mustard gas was employed for the first time in World War f in 1917 near Ypres, Belgium. This early use earned the gas two nicknames: Yperite, and Yellow Cross (due to the special sign on mustard gas-containing shells). Several exposures have since occurred. Following World War II, large stockpiles of chemical weapotls including mustard gas were dumped in the Baltic sea. Corrosion of the containers led to the liberation of shelfs which were accidently brought on board by fishing trawlers, resulting in exposure of fishermen to the vesicant’. Mustard gas was used by Iraqi forces during the Iran-Iraq conflice; the Iranian soldiers who were evacuated for treatment in hrrmpe showed classical symptoms of mustard gas poisoning? The possibility of large scale exposure of military and civilian populations to mustard gas must once again be seriously considered. This review describes the pathological effects of the vesicant, its cellular mechanism of action, and the available antidotes for prevention of skin bums caused by mustard gas.
Despite its name, mustard gas is a volatile oily liquid, slightly soluble (0.07%) in watefl, which rapidly penetrates human skin within 30 minutes of exposurd. Saturated vapor of mustard gas is absorbed at a rate of 1.4 pg cm-* min-’ at 22°C (Ref. 6). Most of the liquid form of mustard gas applied to the skin evaporates (80%) and of the 20% that penetrates, half is fixed in the skin and half is absorbed systemically’. Pathological effecta of muabrd gas lesions in humans Although there are great discrepancies in the sensitivity of individuals to mustard g&, the following description, based cn controlled human studies performed with 143 gas mask-protected volunteers in 1949 (Ref. 9), may be a typical picture of skin lesion development after exposure in times of emergency.
Unlike heat bums the typical chari&&r,ic of mustard gasinduced skin bums is its delayed effect. These start to appear at least several hours after poisoning. Exposure of humans to mustard gas vapor resulted in erythema which developed within 24 hours in the majority of subjects. In mild cases it was delayed for 3-4 days and in severely affected persons it appeared within 4 hours. In most cases the erythema gave place gradually to the pigmentation characteristic of mustard gas bums. Subcutaneous oedema was detected 3-9 days after exposure depending on the region of the body. Vesication of the skin occurred later, appearing as numerous blisters about the size of a pin’s head or smaller, and sometimes developing into large blisters. In most cases the mean time for first onset of v&cation in the various regions of the body was 7.2 daya; blistera invoked superficial layers which healed in a few days. By contraat, in severe cases (11% of the exposed individuals) deep veaicka appeared during the first four daya. Deep veaication or deaquamation produced breaches tf the epithelium surface leading to painful raw surfaces which were the main portals of infection. The mean time for the first appearance of raw surfaces was 8.8 days while maximal severity of the lesiona was obuemed after a mean of 10.9 days. The average time for healing was 19 days, while in severe cases it
EPS - Apriil1991 /Vd. 121 could lengthen to months, particularly on the penis. Mustard gas vapor differentially affected various regions of the body. The most severely injured organs were the wet and warm areas, i.e. scrotum, axillae, penis and buttocks. The trunk, dorsum of hands (palms and soles were rarely affected) and forearms were also frequently injured to different degrees of severity9. Additional delayed symptoms such as epigashic distress and vomiting, appearing hours after exposure were ohsemd in soldiers attacked by mustard gas during World War I (Ref. 10). Occnlar damage such as conjunctivitis, lacrlmation, photophobia and eyelid oedema occurred in most of the affected individuals, while in severe cases cornea1 involvement, iritis and chemosis also appearedr”. The respiratory system was also highly vulnerable to mustard gas vapor. LaryngItls and bronchitis were primarily irritative and in Severe cases were complicated, 3-4 deys after exposure, by bronchopneumonia or pulmonary o&ma. The fatal cases (2-5s of exposed individuals) following ~~m~;;q~;~$;$?{ gone marrow toxicity was recorded in the most severely affected soldiers2 as well as in accidental exposure’*. The main long-term effect of mustard gas is its tumorigenic activity. The mutagenic and carcinogenic potential of the vesicant was proven by in vitro and in viuo experiments, as well as by epidemiologlcal studies’“. A 4%year follow-up of workers in British mustard gas manufacture during World War I1 has shown that the death rate from respiratory tract malignancies was higher (up to 5.5 times) than that of the onexposed populationr4. An even higher incidence of lung cancer (up to 35-fold) was observed in empIoyees in Japanese and German mustard gas factories”. Duration of employment significantly affected risk for certain cancers”. High mortality from chronic nonmalignant respiratory diseases was also observed”. Long-term keratopathy, including deposition of hyalin, Ca2* and crystals, was also detected 35-50 years after exposure to mustard gas during World War 1t5.
Cl-CH,-CH,-S-C&-CH,-Clmustard gas OH-C~J~~~-S-CH~-CH,-OH
h,,,lysis/ drtoxif&i&l
ion r-p
1 conjugation
excretion Cl-CH,-CH,-S-Cl-$-CH,-R, mono-alkylaticn R,-CH,-CH,-S-Ct$-CH,-R, di-alkylation
DNAALKYlAltoN
DNA,R?4AandPROlEtNALKYUVON
CELLUIAR PAwoLoQY
GROSS PAlMoLoGv
carcinogenicity
inhibition of DNA RNAand
pmtein synthesis, dwomatid aberrations, mduoed glycdysii, NADdepletion
erytherna
t basacelldmage, localewtraveaatkn, leukocyte
krfiltration,relsased inflammatorymediators
o&ma
J Miter formationand desquamation
Mechanismof action of mustard
gas It is established that the key reaction of mustard gas is the intramolecular cyclization to form the electrophilic ethylene episulfonium intermediate and the liberation of free chloride anion $I=PFig. 1). The conversion to this reactive deri-rative is temperature dependent”’ and is facilitated in the presence of aqueous solution’. This fact may explain the high vulnerability of warm and wet regions and the mucosal tissues of the eye and respiratory tract to mustard gas. Similar chemical cyclization occurs with the antineoplastic drug nitrogen mustard
(mechiorethamine) to form the ethylene immonium derivative”. The cyclic -onium intermediate alkylates the nucleophilic residues of macromolec4es. The major alkylation site of nucleic acids of mammalian origin is the N-7 of the guanine residue“‘. The second chloroethyl moiety of the mustard can attack additional I%7-guaninr to create inter&and” and intrastrandm crosdinklng. Additional targets for alkylation are N-3and b-g~anfne~‘-~. adeninc aberrations TllUS, chromatid occt~~‘*~,synthesis of DNA, RNA and proteins is inhibited and cells are blocked at the interfaceof the C&/Mphases of the cell cyclers.
TiPS-April 1991[Vol. 121
166
The sulfoxide form of mustard
TABLE I. V~!dMng activityd IWSWd geSdSrivatiVSS coAIpound-----
Rob
VW ;+
VI VII
s(WCICHC!,)(CH,CH,CI) 6(CHClCHCM* 6(CHClCH,)P
VM
S(CH2CHZCHzCl)z s(CHBcHd(W,CHzCI) .%CaHJ(cH2CH2cI)
+.++’ ++ +
XI XII XIII
s(CycsHs)(CH&HzCI)
XIV XV
02s(CH2CH2cl)z OS(CH=CH&
+
XVI
og5(CH=CH&
+
os(cH?cHzC1)2
6.36,*1 41 17.36 41 41 41 17 ii 6 I7 6 17.37 37,41 17 17
++,slmng;+,moderate:-,t~vesicanl.’extentdvesicstingectMtynotde~
Nucleic acids are not the only target macromolecules attacked by sulfur mustard. Inhibition of various glycolytic and respiratory enzyme@ and impaired glucose uotake2b have also been observed f&owing exposure t0 sulfur mustard. Gross et nl. have shown reduced NAD+ levels after sulfur mustard exposure in human skin grafted to athymk nude mi@. This phenomenon has also been observed in other biological systemP2. In human keratinocyte cultures, prevention of NAD+ depletion did not reverse glucose uptake inhibition, which !Y a measure for cellular viabilip, mdicating that reduced NAD+ levels could be one of several mechanisms of sulfur mustardinduced cell injury in human skin. Dannenberg and coworkers developed an organ culture from rabbit skin exposed in vitro to mustard garP. This ex oiuo system was used to follow the inflammatory processes accompanying the various stages of sulfur mustardinduced skin lesions, including migration of polymorphonuclear, basophil and mononuclear teukocytesm, release of proteinases3’ and proteinase inhibitor!?*, and entry and turnover of serum proteins in developing and healing of skin lesionsm. Despite the extent of the nonspecific cellular processes impaired by its potent alkylating activity it appears that sulfur mustard primarily disrupts the normal homeostasis of cell proliferation particularly of the germinative (basal) cells of the ski#, resulting in separation of the epidermis from the dermis and blister formation.
Structu~vesicatlng activity relationships The key reaction of sulfur (~,fl’-dichlorodiethyl mustard sulfide) involves intramolecular cyclization to form the electrophilic ethylene episulfonium derivative. Compounds that do not produce this intermediate (see Table I) either because of the substitution of chloromethyl moieties (compound II) for chloroethyl groups (I), or replacement of Cl- by hydroxyl groups to form the urinary metabolite dihydroxydiethyl sulfides’ (Ill), have no vesicating activity. Similarly the introduction of a Cl- into the aposition of either one arm of the difunctional sulfur mustard (IV, V) or both arms (VI, VII) produced non-vesicating compounds. This might be a result of reduced electron density of the sulfur atom and decreased intramolecular cycliition. The presence of shlorlde in the y positions of the dichlorodipropyl sulfide derivative (VW) eliminated its blisterogenic activity whereas the S,@‘-dlchlorodipropyl sulfide (lx) was bund to have comparable activity to that of mustard gasas. In spite of its single alkylating group, chloroethykthyl sulfide (X) was shown to have strong veelcating actlvlty because of its rapid penetration of the skin, probably due to the presence of lipophilic moiety. Its penetration is more rapid than compound I (Ref. 6). Another monofunctional sulfur mustard in which the ethyl was replaced by a phenyl group (Xl) had blis nit activity, but introduction“gp0 benzyl moiety (XII) caused the molecule to be inactive.
gas (XIII) had no toxic activity. However, the sulfone mustard (XIV), whose conjugated form was
found as a urinary metabolitr, did act as a vesicant although less potently than sulfur mustard itself. Although neither compound is able to cyclize to form the isulfonium ion it electrophilic was assumed 2 that they are converted in vioo to the corresponding non-active divinyl &oxide (XV) and to the toxic vesicant divinyl sulfone (XVI), respectively. The possibility of divinyl sulfone activation via epoxide formation cannot be excluded. These assumptions need to be further investigated. Neutralizing agenln for mustard grs The ele&ophilic nature of the ethyleneepi&onium intermediate can be neutralilntd by nucleophilic compounds. Sulfhychyl agents such aa dlthiothwitd
and
mercaptoethylamlne reduce sulfur mustard toxicity in in titro systemP. In in ufuo studies in fmimalt3 sodium thlosulfate’ combined with various drugs such as cortlcosteroids, antlmddanb, anticoagulants and antlbistunines was able to prevent, to smno extent, mustard gas t&cl@‘. HowS ever, neither these amt nor numerous others7 JFZ? completely protective against systemic or topical toxic&y of mustard gas. Skin lesions caused by percutaneous appliattion of sulfur mustard can be prevented by adsorbent powders such as Fuller% earth, charcoal and t&urn* provided they are used within 10 minutes of exposure. Recently we have patented a non-toxic and non-irritant iodlne/povidoneiodine-containing preporation which efficiently protects guineapig skin from must& gas even when applied 20 minutes after exposu#. Preliminary results of similar newly developedprepar. ations obtained by us indicate that even longer intervals such as 40 and 60 minutes between exposure and treatment achieve high degrees of protection against mustard gas. cl
cl
cl
Although mustard gas has been known for decades as a powerful
TiPS - April 1992 [Vol. 121 chemical weapon to which mass populations might be exposed, understanding of its mechanism of action is quite limited. A combination of detailed structureactivity zrudies with inves’igation of the molecular and cellular pathology of mustard gas may lead to practical solutions for the current military medical problems.
References 1 Aasted, A., Dane, E. and Wulf, H. C. (19S7)Ann. Plasf. surg. 19, X30-333 2 Suhrabpuur, H. (1987) Med. J. Islamic Rep. irau 1,32-37 3 Requena, L. et al. (1988) J. Am. Acad. Dcmutol. 19,5=6 4 Hopkins, E. F. (1919)J. PJ~anacoI. Exp. 77Icr.lz35?uu3 5 Culltunbine, H. (1946) Br. 1. Dmalol. 5B,291-294
6 Nagy, S. hi., Golumbic, C., Stein, W. H., Frutun, J. S. and Bergmann, M. (1946) /. Cen. Physfol. 29,441&9 7 Sumani, S. M. and Babu, S. R. (1989) 1. C&I. Pharatacal. Thcr. Toxicof. 27, 419-435 8 Mamhali, E. K., Lynch, V. and Smith, H. W. (1918)J. Pharmrcul.Exp. Thcr. 12, 291301 9 Sfndair, D. C. (1949) Br. I. Derwtol.
Six volumes on medicinal chemistry Comprehensive Medicinal Chemistry edited by Cotwin Hansch, Pergamon Press, 1990. $1995.00/E1145.00 kii + 5494 pages in six volumes) ISBN 0 08 032530 0
It is not insignificant that the first chapter of Comprehensive Mcdicinal Chemistry, written by a team of over 250 authors with Corwin Hans& chairing the board of editors, comes from Alfred Burger. Indeed, this book may be regarded as a successor to Burce; Medicinal Chemistry, ’ appeared in l%O. In 1980 three volumes sufficed, but now an impredvese&es of six volumes is needed in order to be comprehensive. The new book contains a wealth of information but also has some drawduplications occur; some subjects are treated in dif-
167 Syphilis 61,113-125 10 Derby, G. S. (1918)Am. I. Med. Sri. 156, 733-736
11 Mandel. M. and Gibson, W. S. (1917) J. Am. Med. Assoc. 69,1978-1971 12 Hobbs, E. 8. (1944)Br.Med. j. 2,306-y17 13 IARC Monographs (1975) Vol. 9, pp. i81-i92, WHG, Geneva 14 Easton, D. F., Pete, J. and Doll, R. (1988) Br. 1. Ind. Med. 45.652-659 15 Blodl, F. C. (197l) InL Ophthalmol. Clin. ll, l-13 16 Ward, J. R. and Seiders, R. P. (1984) Thcrmochim. Acfa 81.343-348 17 Dixon, hi. and Needham, D. M. (1946) Nature 158,432-438 18 Wheeler, C. P. (1962) Cancer Rts. 22, 651-688 19 Ball, C. R. and Roberts, J. J. (1972) Chrm-tliol. Interact.4.297303 20 Walker, 1. C. (197l) Con. 1. Biochcm.49, 332-336 21 Ludhun, D. B., Kent, S. and Mehta, J. R. (1986)Carcinogenesis7,1203-12% 22 Habraken, Y. and Ludhun, D. B. (1989) carcfnogcnesii 10,489-492 23 Kirchner, M. and Brendei, M. (1983) Chem-Biol. Interacf. 44, n-39 24 Savage, J. R. K. and Breckon, C. (l%l) Mutat. Rn. 84,37u87 25 Rub&s, J. J., Friedlos, F., Scott, D., Cbmerud, M. C. and Rawlings, C. 1. (1986)Mulot. Res. 166,169-181 26 Md, M. A. E., Van De Ruit, A. M. 5. C. and Kluivers, A. W. (1989)Toxicol.Appl. Pharmacol.98,159-165 27 Gross, C. L,, Meier, H. L., Papirmeis~er,
ferent volumes, which makes it difficult to get complete information quickly; the style differs substantially tinm chapter to chapter but, more importantly, the difkrent subjects are not all pitched at the same level. The presentation of the book is excellent, but this is to be expected as the price of the book is extraordinary. Very few printing errors appear throughout the book. In the prefaace of the book the aim is presented as ‘. . . to present the subject, the modem role of which is the understanding of relationships structure-activity and drug design from the mechanistic view-point, as a field of its own right, integrating with its central chemistry all the necessary ancillary disciplines’. The first volume covers general principles. Several chapters are very general and offer familiar information, and the presentation makes the volume very useful for educational purposes (particularly the chapter by Sneaders on the
B., Brinkley, F. B. and Johnson, J B. (1985) Toxicof. Appt. fharmoml. 81, 85-90 28 Petrali, I. P., ogksby, S. 8. and Meier, H. L. (1990) Ultrostruct. Parhol. 14. 253-262 29 Ku, W. W. and Bernstein, I. A. (1988) Toxicol.Appl. Phermacol. 95,397+1 30 Darwten& A M. Jr et al. (1985)AM. 1. Pufhol. 121.15-27 31 Higuchi, K. et al. (1988)lnflummrtion12, 311-334 32 Harada,5. et al. (1987)Am. 1. Pathol.l26, 148-163 33 Handa, S. et al. (1985)Am. 1. Pathol. 121, 28-38
34 Davison, C., Rozman. R. S. and Smith, P. K. (l%l) Biochem. I%wmucol. 7,65-14 35 Lynch. V., Smith, H. W. and Marshall, E. K.Jr(1918)1. Pharmacol.Exp. ntr. 12, 265-290 # Renshaw, B. (1947)1. Invest. Dennatol. 9, 7S-85 37 Banks, T. E., BoursnelL J. C., Francis, C. 8.. Hupwuud. F. L. and Wonnall, A. (1946)Biochrm.J. 40,734-736 38 Walker, I. G. and Smith, J. F. (1969)Can. J. Physiol. Pharmaco~.47,143-151 39 Vojvudic, V., Milusavljwic, Z., Buskovic, B. and Bojanic, N. (1985) Fundurn. A@. ToticoI. 5, S165S168 40 Sohnan, T. (1919) J. Pharmrcol. Exp. Ther. 12,303-318 41 Peters, R. A. and Walker, E. (1923) Biochm. I. 17,~W6 42 Wonnser, IJ. (1990) Israeli Patent No. 95087
chronology of drug introductions and the chapter on sources of information). The chapters on physiological aspects are very condensed, but informative. Some of the chapters are too limited (even superficial) to be of interest (e.g. the chapter on selectivity by Lipnick, which seems to be an abstract of Albert’s famous book). Too much attention is paid to aspects of recombinant DNA technology procedures. Information on organization and funding of medical research and on healthcan? systems is included, but only for the USA and UK. The next four volumes treat medicinal chemistry subjects impressively and concord with the promises made in the preface. Volume 2 covers enzymes and other molecular targets; a lively, factua! and fascinatig account but again there is variation in presentation, e.g. treatment with fi-lactams begins with chemical aspects, while in another case biological mechanisms open the chapter. Volume 3. Membranes and Receptors, also shows remarkable differences between chapters; the information on p-adrenoceptors