Chem. Res. Toxicol. 1989,2, 301-306
30 1
2,3-Dithioerythritol, a Possible New Arsenic Antidote Victoria L. Boyd,*$+John W. Harbel1,t Richard J. O’Connor,t and Evelyn L. McGown*it Division of Biophysical Research, Chemistry Branch, Letterman Army Institute of Research, Presidio of S a n Francisco, California 94129-6800, and Microbiological Associates, Inc., 9900 Blackwell Road, Rockville, Maryland 20850 Received March 8, 1989
British antilewisite (2,3-dimercaptopropanol; BAL) has long been used as an arsenic antidote, but its therapeutic efficacy is limited by its inherent toxicity. We synthesized two less toxic derivatives of BAL and investigated their potential as antidotes to organic arsenic. The new compounds, 2,3-dithioerythritol (DTE) and 2,2-dimethyl-4-(hydroxymethyl)-1,3-dithiolane (isopropylidene derivative of BAL), react readily with phenyldichloroarsine (PDA) to yield the expected corresponding cyclic 1,3-dithioarsolanes. The BAL derivatives were compared to BAL in terms of their cytotoxicity and their ability to rescue PDA-poisoned mouse lymphoma cells in culture. The dithiolane was not a good antidote in the cultured cell system. In contrast, DTE was less toxic than BAL or DMSA and was superior at improving cell survival in PDA-exposed cells.
Introduction Trivalent arsenic is toxic because of its reactivity with crucial biological sulfhydryls (I). The only known antidotes are dithiol compounds which compete and extract the arsenical away from tissue sulfhydryls. The efficacy of dithiols is due to their ability to form cyclic adducts with arsenic which are more stable than the adducts formed with monothiols (2). British antilewisite (BAL;’ 2,3-dimercaptopropanol) was among the dithiol-containing compounds synthesized in the early 1940s (1, 2) in the search for a topical antidote to lewisite [trans-(2-chlorovinyl)dichloroarsine] (2, 3). BAL is an effective topical antidote to vesicant arsenicals. However, it is less than ideal as a systemic antidote because of its inherent toxicity and its unpleasant side effects. Nevertheless, BAL has been the recommended arsenic antidote in the United States for the past 40 years (4). Less toxic analogues of BAL have been receiving attention recently, in particular, dimercaptosuccinic acid (DMSA) and dimercaptopropanesulfonic acid (DMPS) (5). DMSA and DMPS have polar moieties (carboxylic or sulfonic acid) that make them less lipophilic, more water soluble, and considerably less toxic than BAL. Both have activity as arsenic antidotes in vivo and in vitro. However, we have observed that, on a mole for mole basis, neither competes as effectively for trivalent arsenic in cultured cells as does BAL (6). It is not known whether they are less competitive because of restricted intracellular access (due to polarity), or because their cyclic adducts are inherently less stable, or both. One goal of the present work was to determine if BAL could be modified to decrease its toxicity without compromising its efficacy. We chose to make a compound with polarity intermediate between that of BAL and DMSA, hoping it would have the efficacy of BAL and the low toxicity of DMSA. To this end, we synthesized 2,3-dithioerythritol (DTE; structure l),a derivative slightly more
* Authors to whom correspondence should be addressed. Letterman Army Institute of Research.
* Microbiological Associates, Inc.
0893-228x/89/2702-0301$01.50/0
polar than BAL by virtue of its additional hydroxymethyl group. Also, both sulfhydryl groups are secondary and should be less ionized (7) at physiological pH-a difference that could lower its toxicity. Our nuclear magnetic resonance (NMR) studies indicated that DTE reacts readily with phenyldichloroarsine (PDA) to form the expected stable five-membered cyclic chelate. We found DTE to be less toxic to cultured cells than BAL and superior to both BAL and DMSA at rescuing cells exposed to PDA. CH-OH
H-C-SH H-C-SH
CH,OH
1
We also considered the possibility of masking the sulfhydryls reversibly to lower their general reactivity but retain selective reactivity toward dichloroarsines. For example, synthetic chemists commonly protect ketones by forming the dithianes or dithiolanes with HS(CHz),SH (8) and then deprotect with HgClZ. We investigated the possibility that a dithiolane prepared from BAL would be “deprotected” in an analogous manner by PDA, resulting in the formation of the BAL-PDA chelate. For this purpose we investigated the isopropylidene derivative of BAL [2,2-dimethyl-4-(hydroxymethyl)-1,3-dithiolane; structure 21. We found that this latent form of BAL did readily react with PDA, but only under acidic and not physiological conditions.
Experimental Section Borane tetrahydrofuran complex (BH3-THF), dry THF, and M g S 0 4 were purchased from Aldrich Chemical Co., Milwaukee, Abbreviations: BAL, British antilewisite, DMPS, 2,a-dimercaptopropanesulfonic acid; DMSA, meso-2,3-dimercaptosuccinicacid; DTE, 2,3-dithioerythritol;PDA, phenyldichloroarsine; NMR, nuclear magnetic resonance;THF, tetrahydrofuran;GSH, glutathione; lewisite, trans-(2chloroviny1)dichloroarsine;LO, lewisite oxide; DMSO, dimethyl sulfoxide.
0 1989 American Chemical Society
302 Chem. Res. Toxicol., Vol. 2, No. 5, 1989
\
S. C H',
II
Boyd et al.
/CH3 L
CH,OH
2 2,3 - DTEePDA
WI. DMSA was purchased from Sigma, St. Louis, MO. Phenyldichloroarsine (PDA) was purchased from Research Organic/Inorganic Chemical Corp., Sun Valley, CA. Lewisite oxide was obtained through the courtesy of Susan F. Hallowell, US.Army Chemical Research Development & Engineering Center, Aberdeen Proving Ground, MD. Solvents were from Fisher Scientific, Santa Clara, CA. Deuterated NMR solvents came from Merck, Sharpe, and Dohme, West Point, PA. I 1 I I I I I I I I I I I I All spectra were recorded on a Varian XL-300 spectrometer la0 1310 120 1:O 100 90 80 70 60 at ambient temperature, and chemical shifts are referenced PPM from ( M e ) 4S i downfield relative to external Me4Si. 'H-13C shift-correlated Figure 1. 13C NMR spectra (75.429 MHz) of the PDA complex (HETCOR) 2D experiment, 'H-13C distortionless enhancement formed with 2,3-dithioerythritol (DTE-PDA, k1.5) in MeOH-d4. of polarization transfer (DEPT) experiments, and phase-sensitive (A) 13C NMR spectra with Waltz decoupling. (B) DEPT exdouble-quantum COSY (DQCOSY) experiments were performed periment showing methylene carbon signals inverted relative to by using Varian programs. Typically, windows of approximately methine carbon signals. 20 and 4 kHz were used for 13C and 'H spectra, respectively. The HETCOR experiments were optimized for 'JcH = 150 Hz. unreacted starting material. 'H NMR (MeOH-d4,300 MHz) 6 The f l (13C) and f 2 ('H) domains contained 1024 and 128 data points, respectively (128 transients per FID). A relaxation delay 4.0-3.9 (m, 1 H, CH), 3.38 (dd, 'JHH = 11.8 Hz, 'JHH= 5.0 Hz, of 10 s was used for both the HETCOR and DEPT experiments. 1H, HCH), 3.03 (dd, 'JHH= 11.8Hz, 3 J m = 8.0 Hz, 1H, HCH), 1.76 (6, 3 H, cH3), 1.74 (9, 3 H, cH3), 1.38 (d, 3 J =~6.5 ~Hz, 3 The pulse sequence chosen for the DEPT inverted the resonance H, CH3);''C NMR (MeOH-d4,75.429 MHz) 6 63.2 [C(CH3),], 53.5 signals for CH2 relative to CH3 and CH. For the DQCOSY experiment, the fi and f2domains contained 512 data points each. (CH), 47.5 (CH,), 35.8 and 34.6 (geminal CHis), 20.6 (CH3). The methylene derivative of BAL [4-(hydroxymethyl)-l,3Spin simulation programs were used to obtain data on the best dithiolane] was synthesized by reacting BAL (0.4 g) with 37% 'H chemical shift and coupling constants. The BAL-PDA adduct was synthesized according to the meformaldehyde (0.6 mL) in 2 mL of concentrated HCl for 1.5 h at thod of Dill et al. (9) by adding PDA to BAL in a 1:l ratio in room temperature. The product was extracted from the aqueous MeOH and recrystallizing in acetone-water. solution with CHzC12(3 X 30 mL), dried (MgSOJ, and concen2,3-Dithioerythritol(l, 2,3-DTE, 2,3-dimercapbl,4-butanediol) trated into an oil. The dithiolane was obtained in 15% yield following column chromatography (silica gel, 955 CH2C12-MeOH, was prepared by the reduction of meso-2,3-dimercaptosuccinic acid (DMSA). DMSA (0.45 g) was suspended in 16 mL of dry R, 0.65). 'H NMR (MeOH-d4,300 M H z ) 6 3.85 and 3.86 (2 singlets, THF under N1.BH3-THF (7.8 mL of a 1M solution, i.e., a 24% 2 H, SCH2S), 3.78-3.70 (m, 1 H, methine), 3.65-3.45 (m, 2 H, CHzOH), 3.29 (dd, 'JHH= 12 Hz, 3 J = ~3.3 ~Hz, 1H, HCH), 3.09 excess) was added dropwise by syringe a t ice bath temperature (IO). The reaction mixture was stirred 4 h while warming to room Hz, 1 H, HCH); (ddd, 2 J ~ H= 12 Hz,3 J = ~5.2 ~Hz, 4 J = ~0.5 ~ 13CNMR (CDCl3, 75.429 MHz) 6 63.8 (CH20H), 55.6 (CH), 39.5 temperature. THF (40 ml) was added, and then the reaction was quenched by the slow addition of 1:l THF-H20 (10 mL). Solid (SCHZS), 34.0 (CHZ). Mouse lymphoma cells were used for the cytotoxicity and rescue K2COsand MgSO, were added; the reaction mixture was filtered, assays. These cells are commonly used in cytotoxicity assay and Et3N (1.5 mL) was added to break up any boron-sulfur systems because they grow rapidly and tend to be more sensitive complexes. The concentrated reaction mixture was purified by to toxic insult than slowly dividing cells. Also, because they grow column chromatography (silica gel, EtOAc, R 0.86), and the product obtained was a yellow oil in 15.8% yield. 'H NMR (300 in suspension,they c8n be counted easily. The cells, clone L5178Y MHz, acetone-d,) 6 4.33 (br s, 2 H, OH, exchanges in D20),4.0-3.7 3.7.2C, were the generous gift of Dr. Donald Clive, Burroughs (m, 4 H, CHz), 3.25-3.15 (m, 2 H, CH), 2.1 (apparent d, 2 H, SH, Welcome Co., Research Triangle Park, NC. They were maintained as an exponentially growing suspension culture in Fischer's meexchanges in D20); 13C NMR (75.429 MHz, acetone-d,) 6 65.6 (CHZOH), 47.2 (CHSH). dium for leukemic cells of mice (Irvine Scientific, Irvine, CA), The synthesis of compound 2 [ 2,2-dimethyl-4-(hydroxysupplemented with 0.22 mg/mL sodium pyruvate, 1 mg/mL Pluronic F68, and 10% horse serum (Hyclone Laboratories, Logan, methyl)-l,3-dithiolane] was similar to the procedure of Stocken UT). At the initiation of each assay, the cells were diluted to 3 (11). BAL (0.92 g) was dissolved in benzene (13 mL). Acetone (1.08 mL) and two drops of concentrated HCl were added, and X lo5 cells/mL in complete medium and distributed as 10-mL the reaction mixture was refluxed overnight. The mixture was aliquots into 15-mL polypropylene culture tubes. Phenyldidried (MgS04) and concentrated into an oil. The purified product chloroarsine was prepared immediately before use by serial diwas obtained in 8.2 % yield following column chromatography lutions, in Hanks' balanced salt solution, of the 50 mM ethanol stock solution. The antidotes were serially diluted in dimethyl (silica gel, CH2C12,Rr 0.9). Attempts to obtain crystalline material sulfoxide (DMSO) immediately before use. DMSO concentrations as reported by Stocken (11)were unsuccesaful. 'H NMR (CDCl,, 300 MHz) 6 4.20 (dd, 'JHH = 9.9 Hz, 'JHH= 3.4 Hz, 1 H,of in the final cultures did not exceed 0.5%. CH20H, diastereotopic effect), 4.13 (dd, 2 J = ~9.9 ~Hz, 3 J = ~ ~ Experiments were initiated by dosing the appropriate cultures 5.2 Hz, 1H of CH20H, diastereotopic effect), 3.64-3.55 (m, 1H, with PDA and then returning all of the cultures to the roller drum incubator for 1 h at 37 f 1 OC. During this time the antidote CH), 2.67 (s, 1H, HCH), 2.65 (br s, 1H, HCH), 1.63 (s,3 H, CH3), 1.55 ( ~ , H, 3 CH3); '3C NMR (CDC13,75.429 MHz) 6 92.8 [C(CH&], solutions were prepared. After the 1-h exposure period, the antidote solutions and DMSO controls were added to the ap72.7 (CHzOH),54.2 (CH), 31.8 (CH3),30.7 (CHJ,29.5 (CH2). (The assignments were aided by a DEPT experiment.) propriate tubes and the tubes returned to the roller drum incubator. Approximately 24 h after the initiation of the cultures, 2,2,4-Trimethyl-1,3-dithiolane was synthesized from 1,2-dithe cells were counted by using a Coulter ZM cell counter. Data thiopropane (0.74 g) and acetone (1 mL) in benzene/HCl as are expressed as percentage of the mean cell counts of the control described above for the synthesis of compound 2. The product was obtained in -80% yield and contained small amounts of the cultures.
Chem. Res. Toxicol., Vol. 2, No. 5, 1989 303
2,3-Dithioerythritol as Arsenic Antidote
30
S
i
50
70
PPM
90
from
(Me), Si 110
130
150
9
8
7
6
5
4
3
PPM from (Me), Si
Figure 2. laC-'H HETCOR contour plot of the DTE-PDA (k1.5) chelate in MeOH-d,.
Results 2,3-DTE (1) was characterized by 'H and 13C NMR spectroscopy in an anhydrous solvent, acetone-d6,in order to identify the deuterium-exchangeable protons (SH and OH, see Experimental Section). The reaction of DTE with PDA was observed in MeOH-d4in order to compare the results with previous studies of BAL and PDA (9). DTE reacted readily with PDA. The 13C NMR of the DTEPDA adduct is reproduced in Figure 1A. Only one resonance signal each for the methine (6 63.42) and methylene (6 62.42) carbons was observed. The chemical shift assignments were based on a DEPT experiment, seen in Figure 1B. The upfield portion of the 'H NMR spectrum of the DTE-PDA adduct was a symmetrical cluster of signals centered at 6 3.76. A lH-13C HETCOR correlated the two upfield carbon signals to the midpoint of all the nonaromatic proton signals (Figure 2). Therefore, this spectrum did not distinguish the correlation between the carbon and proton resonance signals. A COSY spectrum confirmed that the protons were all coupled to each other. The reaction of BAL with acetone resulted in the formation of the expected cyclic thioketal (11) (2). A 'H NMR of this dithiolane derivative of BAL in MeOH-d, (containing traces of water) is reproduced in Figure 3A. It is distinctive due to the diastereotopic methyl group signals at 6 1.63 and 1.55. These methyl groups, which originated from acetone, are diastereotopic by virtue of the chiral center contributed from BAL. In order to determine whether this dithiolane would react with a dichloroarsine, 1 molar equiv of phenyldichloroarsine was added. A 'H NMR spectrum was recorded immediately afterward and is reproduced in Figure 3B. The dithiolane was no longer present except for small traces due to a slight excess of the compound relative to PDA. Particularly noticeable was the almost complete absence of the diastereotopic methyl signals. Acetone was reformed quantitatively and detected as a signal at 6 2.11, labeled Z. BAL, which was simultaneously freed, reacted immediately with PDA to form the BAL-PDA adduct.
I'
I
C
I' 2'
AI
8
Y
A 4.0
3.5
3.0 2.5 PPM from (Me)4Si
2.0
1.5
1.o
Figure 3. 'H NMR spectra (300 MHz) in MeOH-d4of the reaction of 2,2-dimethyl-4-(hydroxymethyl)dithiolane with PDA (S= solvent). (A) Base-line spectra of the unreaded dithiolane. (B) Immediately after PDA was added to the dithiolane in (A). (C) Authentic spectrum of the BAL-PDA adduct (courtesyof Dill et al., ref 9). (D) After 2.5 h at 37 "C; note the loss of signal intensity for acetone (Z) and acetone hydrate (Z'). (E) Spectrum after the addition of 1 molar equiv of acetone to (D).
The resonance signals for this adduct lined up exactly with an authentic spectrum of BAL-PDA from a previous study (9),which is reproduced in Figure 3C. Because a new chiral center was created at arsenic, the BAL-PDA adduct existed as two diastereoisomers. Both diastereoisomers are present in Figure 3B,C, and both spectra show a pre-
304 Chem. Res. Toxicol., Vol. 2, No. 5, 1989
-
100-
0
-0-
11"
Boyd et al. BAL
DE
-fr- Dnhiolane
c)
$
60-
0
8 d
e a
60
BAL
40:
DTE
-me4
+ Dilhioiane 0
. . . . . , .. , 1
.
, ,
. . .., , 10
. .
. . . . .., too
,
,
,
,, ,
,,
1000
,
,
.
0 10000
Antidote (pMol)
Figure 4. Cytotoxicity of BAL and derivatives to mouse lymphoma cells during 24-h exposure. Data are expressed as percent of cell concentration of parallel control culture, and each plot contains the combined results of four separate experiments.
dominance of the same isomer. Under the reaction conditions, PDA released HCI. The acidic conditions catalyzed the formation of the hydrate of acetone seen as a signal at 6 1.26 and labeled Z'. Acetone was evaporated from the reaction mixture by heating at 37 "C for 2.5 h. The resulting spectrum, Figure 3D, shows the absence of the signals assigned to acetone and its hydrate. These signals reappeared upon the addition of 1 molar equiv of acetone to the same sample (Figure 3E). Both acetone (Z) and its hydrate (Z') are present in the same approximate ratio in spectra B and E of Figure 3. In a control experiment, 1equiv of PDA added to a sample of acetone in MeOH-d4also catalyzed the formation of the hydrate of acetone. The reaction of the dithiolane and PDA could not be observed in D20 because neither was sufficiently soluble. As a compromise, the reaction was repeated in D20DMSO-d6 (25:75), which gave results identical with those observed in MeOH-d,. Similar results were obtained for the reaction of the dithiolane with lewisite oxide (LO), which had been solubilized by the addition of 2 equiv of DC1, in MeOH-d4. The BAL-lewisite adduct formed instantly, and its spectrum was identical with that formed from the reaction of BAL with LO (12). The isopropylidene derivative of BAL did not react with the water-soluble form of PDA [PhAs=O or PhAs(OH),] in a MeOH-H20 solution that had been adjusted to -pH 7.0. NMR studies in MeOH-d4 also showed no reaction of the dithiolane with phenylarsonic acid, sodium arsenite, or the 2:l GSH-PDA adduct. Figure 4 illustrates the relative cytotoxicity of BAL, DTE, dithiolane, and DMSA to mouse lymphoma cells in culture, and Figure 5 illustrates their relative abilities to rescue arsenical-poisoned cells.2 BAL displayed some Figures 4 and 5 contain the composite results of four separate experiments which were conducted several weeks apart. All experiments were qualitatively similar; Le., BAL was consistently the moet toxic, DTE was consistently the best antidote (above 1 @MI,and DMSA was consistently a poor antidote in the cultured cell system. However, the cultured cells differed considerably between experiments in the magnitude of their response to the treatments, which yielded large variability when all experiments were pooled for statistical evaluation. Even so, we applied a two-way ANOVA model to look for differences between compounds. To increase the number of data points per treatment, the data were collapsed into four concentration ranges (4-20,25-50,100-250, and 500 pM). When differences occurred, Dunnett's t test was applied to determine which compound differed from BAL (at P 5 0.05). In Figure 5, DMSA and dithiolane differed from BAL over the first three Concentration ranges (4-250 FM), and DTE was significant at the 0.05 level of confidence only at 100-250 rM. The compounds were not compared t o BAL at 500 rM due to insufficient numbers.
. . . . . . ..,
. . . .. . ..,
1
1
01
. . . . ,., ,,
,
, , , ,,.
.,
,
loo
10
, ,
.., , 1000
.
,
.. 10000
Antidote (@I) Figure 5. Efficacy of BAL and derivatives in reversing the cytotoxic effects of PDA exposure. Mouse lymphoma cells were exposed to PDA for 1 h and then cultured in the presence of antidote for 24 h. Data are expressed 8s percent of parallel control (no PDA or antidote) culture cell concentration. PDA-treated tubes that had received no antidote had cell concentrations 10-15% of controls. Each plot contains the combined results of four separate experiments.
cytotoxicity at concentrations as low as 4 pM and thus was the most toxic of the four antidotes. In particular, DTE was approximately 1 order of magnitude less toxic than BAL. DTE also was the most effective at rescuing arsenical-poisoned cells (Figure 5). At concentrations less than 5 pM, BAL and DTE had comparable antidotal efficacies. With increasing antidote concentration, the efficacy of BAL rapidly dropped off due to its inherent toxicity, whereas DTE showed maximal efficacy over a much larger concentration range (5 to approximately 500 pM). DMSA and the dithiolane were less effective than BAL, except at the highest concentrations where the results were complicated by the inherent toxicities of the antidotes.
Discussion DTE, a derivative of BAL and DMSA, was synthesized for the first time, for investigation as a possible antidote to organic arsenic. NMR spectroscopy was used to confirm DTE's ready reactivity with arsenicals and the stability of the adducts. DTE was less toxic to mouse lymphoma cells in culture than BAL or DMSA and was superior at improving survival of PDA-exposed cells. 2,3-DTE differs from BAL by the presence of the additional hydroxymethyl group. The 'H NMR spectrum of 2,3-DTE in anhydrous acetone is very similar to that of BAL. DTE reacted readily with PDA, and the product consisted of only one of the two possible diastereomers. The hydroxymethyl groups from the (meso-) 2,3-DTE must both be on the same face of the cyclic DTE-PDA product. By analogy to the major isomer observed for BAL-PDA, the single diastereomer that formed from DTE and PDA is tentatively structure 3, in which the hydroxymethyl groups are trans to the phenyl ring of arsenic.
I HI
Pil
H
3
Chem. Res. Toxicol., Vol. 2, No. 5, 1989 305
2,3-Dithioerythritol as Arsenic Antidote Table I. ‘HNMR Chemical Shifts and Coupling Constant Values for DTE-PDA (Structure 3) chemical 2J~(H), proton assignment shift, ppm Hz VW(H),Hz methylene (diastereotopic) 3.87 -11.34 6.19 methylene (diastereotopic) 3.64 -11.34 5.47 methine (enantiomeric) 3.75 6.19, 5.47
A DEPT experiment aided in the 13CNMR assignment of the methine (6 63.42) and methylene (6 62.42) signals of DTE-PDA (Figure 1B). The presence of only one methine and one methylene signal further supported the presence of only one diastereomeric product. A ‘H spectrum of the DTE-PDA adduct was analyzed by a spin simulation program. The symmetry and coupling constants (Table I) were consistent with the presence of a single diastereomeric product. The chemical shift of the two enantiomeric methine protons of DTE-PDA was 6 3.75. This is similar to the major isomer of BAL-PDA (6 3.97), in which the methine proton is syn to the phenyl group. The chemical shift therefore supports the assigned structure (structure 3). DTE-PDA was more stable in solution (MeOH) than BAL-PDA. The latter turned cloudy after several hours, whereas solutions of DTE-PDA were stable for weeks. The cloudiness of the BAL-PDA solution was likely due to gradual polymerization/dimerization of free BAL which was present due to the microscopic reversibility of the reaction with PDA. We suspect that DTE-PDA is more stable because DTE itself is more stable in solution (unpublished observations) and because the DTE-PDA chelate has a higher association constant. The second hydroxymethyl relative to BAL could increase the stability of DTE-PDA by providing an additional chelating oxygen atom. The second compound we synthesized, the isopropylidene derivative of BAL, readily reacted with PDA, but not at neutral pH. Thus, it would seem to have no value as a systemic arsenic antidote. Certainly, in our cultured cell system, it showed no antidotal activity below 100 pM. We suspect that the small amount of activity above 100 pM was due to traces of underivatized BAL and not due to hydrolysis of the dithiolane because it is hydrolytically stable (13). However, the isopropylidene derivative is still potentially useful. It reacts with neat dichloroarsines and is considerably less toxic than BAL. It could conceivably be better than BAL for certain purposes, e.g., as a topical cutaneous or ophthalmic antidote. In methanol, the reaction between the isopropylidene derivative and PDA readily occurred, with the formation of the BAL-PDA adduct and acetone. The mechanism for this reaction is shown in Scheme I and is analogous to mechanism for HgC12 deprotection of dithianes (24). In order to substantiate that the reaction mechanism for PhAsC12is similar to that for HgC12,we investigated the reaction of PDA with the methylene derivative of BAL. No reaction was observed. This (anticipated) lack of reactivity was due to the decreased stability of the primary carbonium ion intermediate in the methylene case relative to the tertiary carbonium ion intermediate formed from the isopropylidene compound. This chemistry parallels the slower rate of reaction of HgC12with dithianes formed from aldehydes. Additional studies were done to determine whether PDA could serve as a general deprotecting agent for dithiane and dithiolane compounds. However, the dichloroarsine failed to react with other “protected” dithiol compounds. As a result of this observation, 2,2,4-trimethyl-1,3-dithiolane, which differs from BAL by the absence of the
Scheme I
I CHzOH
I
H,C-C,H2
CHzOH
I
Ph
L
hydroxyl group, was investigated to serve as a probe into the reaction mechanism. It also failed to react with PDA. Evidently, the unique reactivity of the isopropylidene derivative of BAL with PDA involves some stabilization of the carbonium ion intermediate by way of the hydroxyl group (neighboring group participation).
Acknowledgment. We thank Drs. Gerald Zon and Kilian Dill for helpful suggestions during the course of this work, Dr. Virginia Gildengorin for statistical analyses, and Angela M. Stanislao for the preparation of the manuscript.
References (1) Webb, J. L. (1966) Arsenicals. In Enzyme and Metabolic In-
hibitors, Vol. 3, Academic Press, New York. (2) Waters, L. L., and Stock, C. (1945) BAL (British Anti-Lewisite). Science 102,601-606. (3) Peters, R. A., Stocken, L. A., and Thompson, R. H. S. (1945) British Anti-Lewisite. Nature 156, 616-619. (4) Klaassen, C. D. (1985) Heavy Metals and Heavy Metal Antagonists. In The Pharmacological Basis of Therapeutics (Gilman, A. G., Goodman, L. S., and Gilman, A., Eds.) pp 1605-1627, Macmillan, New York. (5) Aposhian, H. V. (1983) DMSA and DMPS-Water Soluble Antidotes for Heavy Metal Poisoning. Annu. Rev. Pharmacol. TOX~C 23,O193-215. ~. (6) McGown, E. L., Harbell, J. W., Dumlao, C. R., and OConnor, R. J. (1985) Use of In Vitro Cell Systems to Study Cytotoxic Effects of Vesicant Arsenicals and Efficacy of Antidotes. Proceedings of USAMRDC Fifth Annual Chemical Defnese Bioscience Review, pp 291-301.3 (7) Brown, E. D., Iqbal, S. M., and Owen, L. N. (1966) The Reductive Fission of Methyl Sulfides, 1,3-dithiolanes and 1,3-0xathiolane by Sodium in Liquid Ammonia. J. Chem. SOC.C, 415-419. (8) Barros, M. T., Geraldes, C. F. G. C., Maycock, C. D., and Silva, M. I. (1984) Formation and NMR Study of some Cyclic B-Ketodithioacetals. Tetrahedron 44, 2283-2287. (9) Dill, K., Adams, E. R., O’Connor, R. J., and McGown, E. L. (1987) 2D NMR Studies of the Phenyldichloroarsine-British Anti-Lewisite Adduct. Magn. Reson. Chem. 25, 1074-1077. (10) Yoon, N.M.,Pak,C. S., Brown, H. C., Krishnamurthy, S.,and Stocky, T. P. (1973) Selective Reductions. XIX. The Rapid Reaction of Carboxylic Acids with Borane-tetrahydrofuran. A remarkably Convenient Procedure for the Selective Conversion of Carboxylic Acids to the Corresponding Alcohols in the Presence of Other Functional Groups. J. Org. Chem. 38, 2786-2792. Copies may be obtained by writing to Commander, U S . Army Medical Research Institute of Chemical Defense, Attn: SGRD-UV-CM, Aberdeen Proving Ground, MD 21010-5425.
306 Chem. Res. Toxicol., Vol. 2, No. 5, 1989 (11) Stocken, L. A. (1947) 2,3-Dimercaptopropanol ('British Anti-
Lewisite") and Related Compounds. J. Chem. SOC.,592-595. (12) OConnor, R. J., McGown, E. L., Dill, K., and Hallowell, S. F. (1989) 2D-NMR Studies of Arsenical Sulfhydryl Adducts. Magn. Reson. Chem. 27, 669-675.
Boyd et al. (13) Miles, L. W. C., and Owen, L. N. (1950) Dithiols. Part IX. Synthesis of Isopropylidene and Benzylidene Derivatives of 2,3Dimercaptopropanol. J. Chem. SOC.,2938-2946. (14) Seebach, D. (1969) Nucleophile Acylierung mit 2-Lithium-1,3dithiamen bzw-l,3,4-trithiamien. Synthesis, 17-36.
