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DRAFT for review - do not cite or quote 1 2 3 4 5 6 7 IPCS EVALUATION OF 8 9 IN BY AND 10 11 12 13 14 15 16 17 18 Unithiol 19 (2,3-Dimercapto-1-propanesulphonic acid, DMPS) 20 21 22 23 24 25 26 27 28 April 2009 29 30 31 32

33 1. Introduction 34 35 Unithiol (2,3-dimercapto-1-propanesulphonic acid, DMPS) was developed and first 36 used in Russia in the 1950s (Petrunkin, 1956; Klimova, 1958), and later used in China 37 (He et al., 1984). It only became more widely used in America and Western Europe 38 since the mid-1970s (Hruby & Donner, 1987), and particularly since the late 1970s 39 when the Heyl Company in Germany began production (Aposhian, 1982; Aposhian et 40 al., 1984). 41 42 Both unithiol and succimer (2,3-, DMSA) are derivatives of 43 (2,3-dimercapto-1-propanol, British Anti-, BAL), and they are 44 replacing dimercaprol as the main used in the management of heavy 45 poisoning (Hruby & Donner, 1987; Aposhian et al., 1995; Andersen, 1999). These 46 derivatives have several advantages over dimercaprol including lower , 47 increased solubility in water and lower lipid solubility. It is due to these properties that 48 they are effective by oral administration (Hruby & Donner, 1987). Succimer is less 49 toxic than unithiol and where these two drugs appear to have similar efficacy as an 50 antidote for a particular metal, succimer is generally preferred. 51 52 Unithiol has been used in the management of acute and chronic poisoning with a 53 number of different metals and metalloids, and is particularly useful for , 54 and . Unithiol can be given parenterally or orally depending on the 55 clinical situation and severity of poisoning. It is well tolerated and adverse effects are 56 relatively rare. Most common adverse effects are skin reactions such as rashes, 57 pruritis and blistering which are allergic in origin. Most resolve within a few days and 58 generally no treatment is required, but antihistamines and/or corticosteroids may be 59 given if necessary. 60 61 62 2. Name and 63 64 International non-proprietary name: Unithiol 65 66 Synonyms: DMPS, sodium (DL)-2,3-dimercaptopropane-1-sulphonate, sodium 2,3- 67 dimercaptopropanesulphonate 68 69 IUPAC name: Sodium D,L-2,3-dimercapto-1-propanesulphonic acid 70 71 CAS No.: 4076-02-2 72 73 Chemical formula: H2C(SH)-HC(SH)-H2CSO-3Na.H2O 74 75 76 HS

SO3H

HS 77

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78 Relative molecular mass: 228.28 (monohydrate) 79 80 Commercial Names: Dimaval® 81 82 Conversion: 1 g = 4.4 mmol 83 1 mmol = 228.3 mg 84 1 g/L = 4.4 mmol/L 85 1 mmol/L = 0.228 g/L 86 87 88 3. Physico-chemical properties 89 90 Physical condition: White crystalline powder 91 0 92 Melting point: 235 C (decomposes) 93 94 Boiling point: Not applicable 95 96 Solubility: Readily soluble in water (350 mg/ml); not readily soluble in 97 ethanol; not soluble in apolar solvents 98 99 Optical properties: Not applicable as the racemate is used 100 101 Acidity: pH 4.5-5.5 of an 1% aqueous solution 102 103 pKa: Not known 104 105 Stability in light: No specific advice with respect to storage is necessary 106 107 Thermal stability: Stable (e.g., aqueous solution may be sterilized and the 108 substance may also be heated for drying) 109 110 Refractive index and 111 specific gravity: Not applicable 112 0 113 Loss of weight on drying: 6-8% when dried to constant weight at 100 C 114 115 116 4. Pharmaceutical formulation and synthesis 117 118 4.1 Routes of Synthesis 119 120 Procedures for the synthesis of unithiol were first described in the 1950s (Johary & 121 Owen, 1955; Petrunkin, 1956). A short description of possible ways of synthesis for 122 unithiol is also given in Hopkins (1981): sodium 2-propanesulphonate is brominated in 123 acetic acid with bromine which gives sodium 2,3-dibromopropanesulphonate. The 124 latter may either be treated with sodium hydrosulphide to give unithiol or may be 125 treated with acetylthiopropane sulphonate which is then hydrolysed with hot aqueous 126 acetic acid thus leading to unithiol. 127

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128 4.2 Manufacturing Process 129 130 The Heyl company uses a patented manufacturing process which involves 131 precipitation of unithiol as the salt, after which unithiol is released by addition of 132 hydrogen sulphide. The unithiol is subsequently recrystallised from (Ruprecht, 133 1997). 134 135 4.2.1 Parenteral Solution 136 137 The crystalline unithiol is diluted in freshly distilled water suited for injection. The 138 sterile filtered solution is then filled into ampoules. All steps have to be performed in 139 an atmosphere of sterile in order to protect the sensitive compound against 140 oxidation. For the same reason only non-metallic working materials should be used. 141 A complex-forming agent such as sodium edetate (1% of the amount of unithiol) may 142 be added to the solution (water for injection) in order to bind ions eventually released 143 from working materials. The ampoules are then sterilized. 144 145 4.2.2 Capsules 146 147 The active compound is thoroughly mixed with the filling aid until homogeneity is 148 achieved. Thereafter the mixture is filled into commercially available hard gelatine 149 capsules. 150 151 4.3 Presentation and formulation 152 153 At an analytical grade unithiol is available from several manufacturers. The 154 pharmaceutical product is available from Heyl Chemisch-pharmazeutische Fabrik 155 GmbH & Co. KG, Berlin, Germany, both for oral and parenteral administration. The 156 sodium salt of a racemic mixture is used for medical purposes. 157 158 Unithiol is available in a pharmaceutical preparation as capsules and a parenteral 159 injection. The capsules contain 100 mg unithiol and the ampoules 250 mg as a 5% 160 solution. The injection can be administered either intravenously or intramuscularly. 161 162 163 5. Analytical methods 164 165 5.1 Quality control procedures for the antidote 166 167 The quality control procedures listed below are oriented towards national and 168 supranational pharmacopoeial standards. Quality control parameters for the antidote 169 include 170 • Identity 171 • Purity 172 • By-products (mainly disulphides) are assayed by high performance liquid 173 chromatography (HPLC) and should not amount to more than 5% of total 174 peak area. 175 • Bromide content: maximum 0.5% (as potassium bromide). 176 • Heavy metals: maximum 20 ppm (as lead). 177 • Loss of weight on drying: 6.0-8.0%.

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178 • pH of an 1% aqueous solution: 4.5-5.5. 179 • The content may be assayed by iodometric titration. The assay by HPLC is 180 preferred however, because of its specificity. At least 95% is set for 181 requirements. The method has been validated formerly possessing a 182 variation coefficient of 1.5% and a recovery rate of 99.7%. 183 184 The pharmaceutical preparations are additionally controlled for 185 • Ampoules 186 • Sterility. 187 • Filling volume. 188 • Optical appearance (colour, clarity of the solution, particulate matter). 189 • Testing for leaks. 190 191 • Capsules 192 • Uniformity of filling weight. 193 • Disintegration time (maximum 30 minutes in water). 194 • Optical appearance. 195 196 5.2 Methods for identification of the antidote 197 198 Several methods are available to identify the antidote including HPLC, infrared 199 spectroscopy, colour reaction with sodium nitroprusside and flame spectroscopy 200 specifically for the sodium in the molecule. 201 202 5.3 Methods for identification of the antidote in biological samples 203 204 Methods for qualitative and quantitative determination of unithiol and its metabolites 205 are published by Maiorino et al. (1987; 1988; 1991) and Hurlbut et al. (1994). The 206 urine is treated with sodium borohydride and analysed by HPLC with fluorescence 207 detection. 208 209 5.4 Analysis of the toxic agent in biological samples 210 211 Heavy metals should be analysed in blood and urine before, during and after antidotal 212 therapy. Sensitive methods, such as atomic absorption spectroscopy (AAS) or 213 inductively coupled plasma-atomic emission spectroscopy (ICP-AES), can be used 214 (Berman, 1980; Bertram, 1983). 215 216 217 6. Shelf-life 218 219 The shelf life for the commercial available pharmaceutical preparations Dimaval® is 5 220 years for the capsules and 4 years for the ampoules. The expiry date is stated on 221 each package. Although no special advice for storage is given, it is recommended 222 that capsules are stored in a dry place. 223 224 225 7. General properties 226

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227 Unithiol, like succimer and dimercaprol, owes it metal-binding properties to the 228 presence of two adjacent groups. Unithiol is a water-soluble dithiol, a derivative 229 of dimercaprol and is capable of forming complexes with a number of metals and 230 metalloids. The advantages of unithiol over dimercaprol are: 231 • Lower local and systemic toxicity. 232 • Better solubility in water. 233 • Active by oral administration. 234 235 It should be noted that unithiol is not a true chelating agent; a chelator is a molecule 236 which binds a metal or ion by at least two functional groups to form a stable 237 ring complex known as a chelate. For mercury, it has been shown that unithiol (and 238 succimer) do not form a true chelate and as such both could be considered 239 suboptimal as metal antidotes (George et al., 2004). However, there are currently 240 no other substances available with the advantages of these two drugs (high water 241 solubility with relatively low toxicity). 242 243 The mechanism of action of unithiol has not been fully elucidated. Most studies on 244 this subject have focused on its interaction with arsenic and mercury. In addition to 245 increasing urinary elimination of arsenic, unithiol has also been shown to alter the 246 relative urinary concentrations of organoarsenic metabolites by interfering with 247 arsenic methylation (Aposian et al., 1997; Gong et al., 2002; Heinrich-Ramm et al., 248 2003). 249 250 Unithiol also promotes mercury excretion and is effective in inhibiting mercury 251 accumulation in renal proximal and distal tubular cells, and protecting against mercury- 252 induced renal damage. Studies in chickens (Stewart & Diamond, 1987) and rat 253 kidneys (Klotzbach & Diamond, 1988) have demonstrated that urinary excretion of 254 unithiol is blocked by both the substrate p-aminohippurate (PAH) and the inhibitor 255 probenecid of the organic transport process. In the in vitro study by Zalups et al., 256 (1998) using isolated perfused segments of rabbit proximal tubules the removal of 257 mercury from proximal tubules by unithiol was blocked by the addition of PAH. 258 Islinger et al. (2001) postulated on the transport of unithiol in renal proximal cells. 259 Unithiol enters the cells across the basolateral membrane from the blood, via the 260 organic anion transporter (OAT). Both oxidised and reduced unithiol interact with the 261 transporter, but the majority of unithiol entering cells is probably oxidised since this is 262 present in the blood in a greater concentration. Once in the cell the unithiol is reduced 263 by a -dependent thiol-disulphide exchange reaction in the proximal tubule 264 cell (Stewart & Diamond, 1988). Unithiol then binds mercury within the cell and the 265 complex exits the cell, presumably via an export pump. The unithiol-mercury complex 266 is not reabsorbed and is excreted in the urine. 267 268 Owing to its high hydrophilicity unithiol is relatively ineffective at clearing metals from 269 the brain (unlike its lipophilic parent compound, dimercaprol) and excretion by the 270 organic anion transporter (expressed in the apical membrane of the choroid plexus) 271 may further reduce the efficacy of unithiol in the brain (Islinger et al., 2001). Of the 272 organic anion transporters expressed in the basolateral proximal tubule cells 273 determined in animal studies (Kojima et al., 2002), recent work has identified (OAT1) 274 as the transporter involved in movement of unithiol into cells; it is a comparatively poor 275 substrate of OAT3 (Koh et al., 2002). 276

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277 Recently, multidrug resistance protein 2 (MRP2 or ATP-binding cassette, sub-family 278 C [ABCC2]) has been showed to be involved in the renal proximal tubular elimination 279 of unithiol complexes of methylmecury (Zalups & Bridges, 2009). 280 281 It is often assumed, that clinical effectiveness of a metal-binding agent is linked to the 282 stability constant in vitro where the greater the stability constant of a metal-binding 283 agent, the greater the mobilisation of that ion following administration of the metal- 284 binding agent. However, Jones et al. (1980) found no correlation in an in vivo study in 285 mercury-poisoned mice. Therefore data on stability constants are not given in this 286 monograph, but may be found elsewhere (Casa and Jones, 1979). 287 288 289 8. Animal studies 290 291 8.1 Pharmacodynamics 292 293 There are numerous studies on the effect of unithiol in animals with experimental 294 metal poisoning. Many studies compare the effect of a number of different metal- 295 binding agents and in some cases the antidotes are given immediately or sometimes 296 before dosing with the metal. Administration of the antidote with or before exposure 297 does not reflect the clinical situation in human metal poisoning. In addition, the effect 298 of antidotes are measured in a variety of ways including changes in survival rates, 299 urinary excretion, faecal excretion, metal concentrations in organs, body burden and 300 biochemical parameters in target organs. Many studies give conflicting results and 301 this is probably a reflection of a variety of factors including differences in doses of 302 metal and antidote, routes and times of administration and methods of determining 303 elimination and retention. Furthermore, direct extrapolation from animal studies to 304 humans is not possible because of the potential differences in the kinetics of both 305 heavy metals and metal-binding agents such as unithiol between animals and 306 humans. 307 308 8.1.1 309 310 Unithiol has been shown to reduce antimony toxicity in animals. 311 312 In a study comparing survival rates of different antidotes in antimony poisoning, mice 313 were given intraperitoneal antimony potassium tartrate 120 mg/kg (LD50 54.6 mg/kg). 314 The antidotes were given by the same route 1 hour later at a dose of 10:1 mole ratio 315 of antidote to antimony (except for dimercaprol which was given at a 1:1 ratio). 316 Succimer and unithiol were found to be the most efficacious antidotes for antimony 317 potassium tartrate poisoning, with succimer the superior of the two (Basinger & 318 Jones, 1981a). 319 320 In an earlier study administration of unithiol was shown to reduce the LD50 of 321 subcutaneous antimony potassium tartrate by a factor of 8 compared to controls 322 (Chih-Chang, 1958). 323 324 8.1.2 Arsenic 325 326 Unithiol has been shown to be of benefit in poisoning with several arsenic

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327 compounds including lewisite (β-chlorovinyl-dichloroarsine). Mückter et al. (1997) 328 argue that unithiol and succimer have advantages over dimercaprol in the treatment of 329 since they are more effective in preventing arsenic from crossing 330 epithelial boundaries and entering cells and they enhance the excretion of arsenic 331 more rapidly and completely. In addition, unithiol and succimer are less toxic than 332 dimercaprol. However, dimercaprol appears to be more effective in restoring cellular 333 function to tissues which are poorly penetrated by unithiol or succimer. Dimercaprol 334 also has the disadvantage of increasing arsenic concentrations in the brain; this is not 335 the case with unithiol or succimer. 336 337 Aposhian et al. (1982) demonstrated the effectiveness of unithiol in rabbits exposed to 338 subcutaneous lewisite. Unithiol increased survival when given orally or 339 subcutaneously. Similarly, in mice injected with sodium arsenite (0.14 mmol/kg 340 subcutaneously), intraperitoneal unithiol (0.25 mmol/kg) was a potent antidote, even 341 when given 2 hours later (Tadlock & Aposhian, 1980). 342 343 In rabbits poisoned with dermal lewisite dimercaprol, succimer and unithiol were all 344 shown to reduce the incidence and severity of changes. There was no difference 345 between the three agents at the dose of 40 µmol/kg. Compared to dimercaprol, 346 succimer and unithiol may have prolonged survival time and the relatively low toxicity 347 of unithiol and succimer allowed high doses (160 µmol/kg) to be given (Inns & Rice, 348 1993). In a study of intravenous lewisite poisoning in rabbits there was no difference 349 between the level of protection provided by the three antidotes (Inns et al., 1990). 350 351 In mice poisoned with sodium arsenite (0.129 mmol/kg subcutaneously) unithiol (0.8 352 mmol/kg intraperitoneally) given 90 minutes later was found to increase the LD50 by 353 4.2 fold (Aposhian et al., 1981). 354 355 In rabbits poisoned with sodium arsenite (1 mg subcutaneously) given either succimer, 356 unithiol or N-(2,3-dimercaptopropyl) phthalamidic acid (DMPA; 0.2 mmol/kg 357 intramuscularly) 1 hour later, the urinary excretion of total arsenic between 0 and 24 358 hours was elevated after antidote administration. However, urinary excretion of total 359 arsenic between 24 and 48 hours was significantly lower than controls. The three 360 antidotes differed in the proportion of arsenic metabolites in the urine. All increased 361 arsenite excretion by decreased dimethylarsinate excretion. Unithiol and DMPA 362 increased methylarsonate excretion but succimer did not. Both succimer and unithiol 363 increased arsenate excretion. Of the three antidotes used, unithiol was the most 364 effective at removing arsenic from the body (Maiorino & Aposhian, 1985). Unithiol and 365 succimer (both at 50 mg/kg) also significantly increased renal arsenic excretion in rats 366 with chronic arsenic poisoning (sodium arsenate 1 mg/kg orally 6 days a week of 3 367 weeks). Both also restored arsenic-induced inhibition of δ-aminolevulinic acid 368 dehydratase activity and hepatic glutathione concentrations. Although both antidotes 369 reduced arsenic-induced histopathological lesions, succimer was more effective (Flora 370 et al., 1995a). 371 372 Kreppel et al. (1989) compared the effectiveness of D-, dimercaprol, 373 unithiol and succimer as antidotes in acute arsenic intoxication using different 374 controlled experimental settings. In one study mice and guinea pigs were injected 375 subcutaneously with 8.4 mg/kg arsenic trioxide (containing a tracer dose of arsenic- 376 74). An antidote (0.7 mmol/kg intraperitoneally) was given 30 minutes later. As

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377 determined 4 and 12 hours after the arsenic injection, D-penicillamine was unable to 378 reduce the arsenic-74 content in any organ investigated (blood, liver, kidneys, lungs, 379 heart, brain, testes, spleen, skeletal muscl, and skin). In contrast, dimercaprol, 380 unithiol and succimer markedly reduced the tissue content of arsenic-74 compared 381 to controls. Finally, the ability of the antidotes to reverse biochemical effects of 382 arsenic was investigated in vitro using suspensions of isolated renal tubule cells. 383 The marked inhibition of gluconeogenesis induced by 30 μmol/L arsenic trioxide was 384 almost completely reversed upon addition of 90 μmol of dimercaprol, unithiol or 385 succimer. In this experimental model, too, D-penicillamine was ineffective. 386 387 In mice given arsenic trioxide, intraperitoneal administration of unithiol (0.7 mmol/kg) 388 0.5 minutes later was less effective than the same dose of succimer. When the 389 antidote was given 30 minutes after the arsenic, succimer and unithiol showed 390 reduced but similar efficacy. The efficacy of the antidotes for reducing arsenic 391 organ concentrations was investigated in mice and guinea pigs. Animals received 392 8.4 mg/kg (0.043 mmol/kg) of radiolabelled arsenic trioxide subcutaneously and an 393 antidote (0.7 mmol/kg intraperitoneally) 30 minutes later. Both unithiol and succimer 394 and the two in combination were more effective as reducing organ concentrations of 395 arsenic than dimercaprol. In addition, dimercaprol increased arsenic concentrations 396 in the brain, whereas succimer and unithiol did not. Succimer increased the arsenic 397 content of bile but unithiol and the two in combination did not (Kreppel et al., 1990). 398

399 In rabbits administered radiolabelled arsenic (1 mg/kg subcutaneously as sodium 400 arsenite), dimercaprol (0.2 mmol/kg intramuscularly) 1 hour later was shown to double

401 the arsenic-74 concentration in the brain. In contrast, unithiol (0.2 mmol/kg 402 intramuscularly) was found to decrease the arsenic-74 concentration to about one-fifth 403 of that observed with dimercaprol administration (Hoover & Aposhian, 1983). Schäfer 404 et al. (1991) demonstrated that arsenic depots after injection of arsenic trioxide into 405 mice could be mobilised by oral administration of unithiol or succimer without 406 increasing the brain deposition, however, oral administration of dimercaprol 407 extensively increased the brain deposition or arsenic. 408 409 In mice given arsenic (5 mg subcutaneously as arsenic trioxide) immediately followed 410 by unithiol (100 mg/kg intraperitoneally) arsenic excretion in the faeces exceeded that 411 in the urine (Maehashi & Murata, 1986). In guinea pigs poisoned subcutaneously with 412 arsenic trioxide (2.1 mg/kg) the combination of unithiol (0.1 mmol/kg both 413 intraperitoneally and orally) and cholestyramine (0.2 g/kg orally) significantly enhanced 414 the faecal elimination of arsenic suggesting that interruption of enterohepatic 415 circulation of arsenic may be a valuable adjunct in the treatment of arsenic poisoning. 416 Increased faecal excretion was not observed with intraperitoneal unithiol plus oral 417 cholestyramine or oral plus intraperitoneal unithiol (Reichl et al., 1995). 418 419 Flora et al. (2005) compared the efficiacy of succimer, unithiol and monoisoamyl- 420 succimer in rats with chronic arsenic exposure (100 ppm sodium arsenite in drinking 421 water for 10 weeks). Antidotes were given after arsenic exposure at a dose of 50 422 mg/kg for 5 days. Succimer was not effective at resolving arsenic-induced oxidative 423 damage in cells or in reducing the arsenic burden. Unithiol was moderately effective 424 against generation of reactive oxygen species due to intracellular access. However, 425 monoisoamyl-succimer was the most effective antidote in reducing reactive oxygen 426 species in the blood and brain. It was also marginally better at restoring the activity

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427 of antioxidant enzymes. 428 429 An in vitro study of guinea-pig liver treated with arsenic trioxide demonstrated that 430 administration of unithiol (and other antidotes) resulted in a shift to faecal elimination 431 by increasing biliary excretion of arsenic. Unithiol was more effective than succimer 432 or dimercaprol but was not as effective as 2,3-bis-(acetylthio)-propanesulphonamide 433 (BAPSA) (Reichl et al., 1990). 434 435 8.1.3 Beryllium 436 437 Unithiol has been shown to enhance beryllium excretion and reduce beryllium- 438 induced toxic effects in experimental animals. 439 440 Unithiol administration (0.7 mmol/kg intraperitoneally) appeared to increase the lethality 441 of beryllium chloride in mice, but the result was not significant (Pethran et al., 1990). 442 443 In rats treated with beryllium (2.5 mg/kg intraperitoneally, as beryllium nitrate), 444 immediate administration of unithiol (50 mg/kg intraperitoneally) or succimer (same 445 dose) was shown to prevent most beryllium-induced biochemical alterations and 446 reduce tissue beryllium concentrations. Unithiol was relatively more effective and 447 resulted in significantly less marked lesions in the liver and kidneys (Mathur et al., 448 1994). 449 450 In another study beryllium nitrate was given to rats for 21 days (0.5 mg/kg, orally 451 daily for 5 days/week) and unithiol or succimer (25 or 50 mg/kg, twice daily for 5 452 days) was administered 24 hours after the last dose of beryllium. Unithiol was 453 effective at reducing beryllium in the liver, spleen and kidneys. The higher dose 454 marginally elevated the faecal excretion of beryllium but also resulted in 455 redistribution of beryllium into blood. Unithiol also reduced hepatic and renal lesions 456 compared to succimer (Flora et al., 1995b). 457 458 In studies comparing antidotal therapy combined with an antioxidant (sodium 459 selenite) supplementation, D-penicillamine with sodium selenite was found to be 460 more efficacious at reducing beryllium toxicity (as measured by glycogen and protein 461 concentrations in the liver, kidneys, lungs and uterus, with concentrations of liver 462 enzymes and beryllium) than unithiol and sodium selenite (Johri et al., 2002; Johri et 463 al., 2004). 464 465 8.1.4 Bismuth 466 467 Several studies comparing different antidotal agents have found unithiol to be an 468 effective antidote in bismuth poisoning. 469 470 In a study of several antidotes comparing efficacy in bismuth poisoning, mice were 471 given intraperitoneal bismuth citrate followed by an antidote 20 minutes later in a 10:1 472 molar ratio antidote:bismuth. All animals treated with unithiol survived and all in the 473 control group died (Basinger et al., 1983). 474 475 In a study of several antidotes comparing efficacy in bismuth poisoning, rats were 476 injected intraperitoneally with colloidal bismuth subcitrate (50 µmol/kg/day, for 14

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477 days). The antidotes were given twice daily (250 µmol/kg/day) for 3 days. The 478 animals were killed on the fourth day and tissue samples analysed. Unithiol, succimer 479 and dimercaprol were most effective in lowering bismuth concentrations in most 480 organs, particularly the kidney and liver, resulting from higher elimination in urine by 481 unithiol and dimercaprol. Dimercaprol was the only antidote effective in lowering 482 bismuth concentrations in brain tissue. It was concluded that unithiol and succimer 483 were the antidotes of choice with dimercaprol reserved for very severe bismuth 484 poisoning because of its own toxicity (Slikkerveer et al., 1992). 485 486 Unithiol was shown to reduce the whole-body burden of bismuth in mice given an 487 intraperitoneal injection of bismuth acetate. The unithiol was given in drinking water 488 (100, 300 or 600 µg/mL) two days prior to and three days after the bismuth. The 489 renal concentration of bismuth was reduced by up to 90% and unithiol also 490 significantly reduced deposition of bismuth in the femur (Jones et al., 1996). 491 492 8.1.5 493 494 Antidotal therapy for cadmium is particularly problematic because the absorbed 495 metal rapidly becomes strongly bound to metallothionein, a low-molecular weight 496 metal-binding protein, whose synthesis is induced by cadmium. Several studies 497 have compared antidote efficacy in and concluded that, although 498 unithiol is effective, other metal-binding agents appear to be more efficacious (Eybl 499 et al., 1984; Eybl et al., 1985; Andersen & Nielsen, 1988; Srivastava et al., 1996). 500 501 Unithiol administration (0.7 mmol/kg intraperitoneally) increased the LD50 of cadmium 502 chloride in mice from 9.1 mg/kg to 15.2 mg/kg (Pethran et al., 1990). Aposhian (1982) 503 also demonstrated that unithiol increases survival in mice injected with cadmium 504 chloride. 505 506 In a study comparing antidote efficacy, mice were given intraperitoneal cadmium 507 chloride at the previously determined LD50 (0.0267 mmol/kg). This was followed by 508 the antidote at a dose of 1:2 or 1:5 (cadmium:antidote) molar ratios by the same route. 509 The animals were killed after 14 days. Succimer and unithiol were found to be most 510 effective at reducing lethality and the cadmium burden of the liver, kidneys and brain 511 tissue. The and therapeutic efficacy was highest for succimer 512 followed by unithiol (Srivastava et al., 1996). 513 514 In a similar study mice were given radiolabelled cadmium chloride (0.53 mmol/kg by 515 stomach tube) followed by the antidote (2.12 mmol/kg) 15 minutes later. The 516 animals were killed after 10 days. Unithiol provided some protection against lethality 517 (whereas succimer provided complete protection). Penicillamine, succimer and 518 unithiol were all able to reduce peristaltic toxicity of cadmium chloride and all 519 reduced whole-body retention of cadmium; succimer was most effective at reducing 520 body retention. Succimer, and particularly unithiol, decreased hepatic deposition of 521 cadmium and increased relative deposition in the kidneys and lungs. It was 522 concluded that succimer was the most effective antidote for cadmium poisoning 523 (Andersen & Nielsen, 1988). 524 525 Another study in mice compared antidote efficacy when given intravenously 10 526 seconds, 1 or 3 hours after intravenous cadmium chloride (3 µmol/kg) administration.

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527 When given immediately after cadmium administration, all agents reduced the body 528 burden of cadmium but efficacy declined when dosing occurred at 1 or 3 hours after 529 administration. Unithiol was among the least effective of the antidotes investigated 530 (Planas-Bohne & Lehman, 1983). 531 532 Unithiol (50 mg/kg intraperitoneally) administered to rats 24 hours after injection of 533 radiolabelled cadmium (1 mg/kg intraperitoneally as cadmium chloride) did not have 534 any effect on the excretion or tissue distribution of cadmium. By the time the 535 antidote was given the cadmium was bound to metallothionein. Only dimercaprol 536 was effective in mobilising cadmium from metallothionein into bile (Cherian, 1980). 537 538 Unithiol (3.61 mmol/kg) was as effective as succimer (same dose) in promoting 539 survival in cadmium poisoned mice (1 mmol/kg cadmium chloride orally) when given 540 immediately after administration of cadmium. However, unithiol administration 541 resulted in a concentration of cadmium in the kidneys and liver that was approximately 542 four times greater than those in succimer-treated animals (Basinger et al., 1988). 543 544 Unithiol did not affect faecal excretion of cadmium and the increase in urinary 545 excretion was too small to affect body burden in rats given 0.4 mmol/kg of unithiol 546 following dosing with radiolabelled cadmium (3 µmol/kg as cadmium chloride). The 547 cadmium was given once; administration of the metal-binding agent started on the 548 third day and was given daily, 5 times a week for 2 weeks (Rau et al., 1987). 549 Administration of unithiol (0.1 mmol/kg) did not affect biliary excretion of cadmium in 550 rats 3 days after intravenous exposure to cadmium (Zheng et al., 1990). 551 552 Unithiol had no effect on survival rate in cadmium-poisoned mice when administered 553 intraperitoneally immediately after subcutaneous cadmium (20 mg/kg as cadmium 554 chloride) at antidote to metal molar ratios of 1:1 or 2:1. At a ratio of 5:1 the survival 555 rate was only 20%, whereas the survival rate with succimer was 100%. In another 556 study where cadmium (0.5 mg/kg intravenously) was immediately followed by 557 unithiol at a dose of 10:1, there was no increase in cadmium excretion or reduction in 558 body burden. Unithiol did, however, decrease the cadmium concentration in the liver 559 and gastrointestinal tract (Eybl et al., 1984). 560 561 In a study of antidotal efficacy in chronic cadmium poisoning in mice (2 mg of 562 cadmium chloride intraperitoneally at 48 hours intervals for 5 doses) unithiol 563 administration (225 mg/kg; 1.06 mmol/kg intraperitoneally) resulted in a significant 564 increase in liver cadmium concentrations (39%). However, interpretation of this 565 result is difficult since this group of mice were the only group to have a significant 566 loss in body weight. Therefore the apparent increase in cadmium concentration 567 could be due to changes in organ size rather than a reflection of the effect of unithiol 568 on cadmium distribution. Similarly there was also a slight increase in kidney 569 cadmium concentrations (Shinobu et al., 1983). 570 571 Unithiol had only a slight effect in reducing the cytotoxicity of cadmium in mammalian 572 cell culture (Fischer, 1995). In another in vitro study using human platelets, unithiol 573 was shown to protect against cadmium-induced stimulation of glutamate binding 574 (Borges & Nogueira, 2008) but unithiol increased cadmium-induced inhibition of δ- 575 aminolevulinate dehydratase (ALAD) in rat lung in vitro (Luchese et al., 2007). 576

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577 578 8.1.6 579 580 There is limited information on unithiol in chromium poisoning. Chromate-induced 581 cytotoxicity (as measured by the chromate content of cells and inhibition of cell 582 growth) was reduced in the presence of unithiol. This was only the case when cells 583 were incubated with unithiol and chromate. If the unithiol was added before or after 584 treatment with chromate it failed to restore chromate-induced cytotoxicity or reduce 585 cellular chromate concentrations (Susa et al, 1994). 586 587 Lethality was reduced in mice injected with chromate (40 mg/kg intraperitoneally as 588 potassium chromate) and then unithiol (500 mg/kg intraperitoneally) immediately 589 afterwards. At half the chromate dose and 300 mg/kg of unithiol the liver and kidney 590 content of chromium was reduced compared to controls. There was also increased 591 renal excretion of chromium and suppression of chromate-induced increase in serum 592 ornithine carbamyl transferase activity (Susa et al, 1994). 593 594 8.1.7 595 596 There is limited information on unithiol in cobalt poisoning. It has been shown to 597 reduce the lethality of cobalt in some studies (Cherkes & Braver-Chernobulskaya, 598 1958; Eybl et al., 1985). In the latter study, although unithiol increased survival it also 599 increased cobalt concentrations in the liver, gastrointestinal tract and carcass in mice 600 receiving 1 mmol/kg of cobalt chloride. The unithiol was given in a dose of 5:1, 601 antidote to metal ratio (Eybl et al., 1985). 602 603 8.1.8 604 605 Unithiol appears to be of benefit in copper-poisoned experimental animals. 606 607 Unithiol administration (0.7 mmol/kg intraperitoneally) increased the LD50 of copper 608 chloride in mice from 59 mg/kg to 143 mg/kg (Pethran et al., 1990). 609 610 Mice receiving copper sulphate (10 mg/kg intraperitoneally, the approximate LD50) and

611 then unithiol (132 mg/kg, 20 minutes later by the same route) were morphologically 612 free of hepatic or renal evidence of toxicity, whereas control animals have extensive 613 renal tubular (Mitchell et al., 1982). 614 615 In contrast unithiol has been shown to increase copper-induced haemolysis of 616 human red blood cells in vitro. At a concentration of 0.3 mM unithiol increased the 617 copper-induced haemolysis from 15% to approximately 25%. At 0.1 mM it was 618 ineffective (Aaseth et al., 1984). 619 620 8.1.9 621 622 Unithiol has been shown to be of benefit in experimentally induced gold toxicity. It is 623 able to reduce kidney gold concentrations and increase urinary excretion of gold. 624 625 In rats given 2 mg gold/kg intravenously (as Auro-Detoxin) and then oral unithiol (0.15- 626 3 mmol/kg) 30 minutes later, it was demonstrated that unithiol reduced the gold

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627 concentration in all organs except the liver and spleen. Similar effects were observed 628 when unithiol was administered 24 hours after injection of gold. In contrast to the 629 immediate treatment with unithiol delayed treatment with the lowest dose (0.15 630 mmol/kg) resulted in significantly decreased gold concentrations in the red blood cells, 631 plasma, femur, muscle and skin. However, the concentration in the kidneys was 632 significantly higher compared to controls (Gabard, 1980). 633 634 In another study rats were given 2 mg gold/kg intraperitoneally daily for 10 days then 635 unithiol 0.75 mmol/kg daily from days 11 to 20. Unithiol administration decreased the 636 gold concentration in the kidneys and in the skin, and increased it in the plasma. The 637 concentrations in the other organs remained unchanged (Gabard, 1980). 638 639 In rats injected with gold sodium thiomalate (up to 0.198 mmol/kg intravenously), 640 treatment with D-penicillamine, unithiol or succimer reduced renal toxicity, measured 641 by urinary concentrations of protein, aspartate aminotransferase and glucose, and 642 the blood urea nitrogen concentration. In addition all three antidotes increased 643 urinary excretion of gold and significantly decreased liver and renal gold 644 concentrations. Unithiol was the most effective antidote tested (Kojima et al., 1991). 645 Characterisation of the gold in urine following treatment with gold sodium thiomalate 646 and then unithiol has shown that it is present as a gold-unithiol complex. In the bile it 647 was present as a gold-unithiol complex, high molecular weight compounds (probably 648 proteins) and gold-L- (Kojima et al., 1992). 649 650 Takahashi et al. (1994) also demonstrated that administration of unithiol reduced 651 renal toxicity (using the same parameters as above) in rats given an intraperitoneal 652 dose (1.2 mmol/kg) immediately after intravenous injection of gold sodium thiomalate 653 (0.026 mmol/kg). Compared to the other gold antidotes tested (bucillamine, captopril 654 and tiopronin), only unithiol was able to significantly reduce the renal gold 655 concentration at the lowest dose used (0.2 mmol/kg). None of the antidotes reduced 656 the hepatic gold concentrations at doses of 0.2 or 0.4 mmol/kg. 657 658 Succimer and unithiol were the most effective antidotes at increasing survival in mice 659 given gold sodium thiosulphate (200 mg/kg intraperitoneally, the approximate LD99). 660 The antidotes were given by the same route 20 minutes after dosing at a ratio of 3:1, 661 antidote to gold (Basinger et al., 1985). 662 663 8.1.10 Lead 664 665 Unithiol has been shown to increase lead excretion, reduce lead tissue 666 concentrations (except in the brain) and to reduce lead-induced biochemical toxic 667 effects, such as inhibition of δ-aminolevulinic acid dehydratase, in experimental 668 animals. 669 670 In a study comparing dimercaprol and unithiol, rats received lead (2 mg/kg as lead 671 acetate) intraperitoneally daily for 7 days, and then antidotal treatment (50 µmol/kg) 672 daily for 3 days beginning 48 hours after the last lead dose. Bone and blood lead 673 concentrations were not significantly different from controls. The concentration of lead 674 in the kidneys was significantly reduced by both antidotes. In addition, both antidotes 675 reduced δ-aminolevulinic acid excretion and appeared to reactivate δ-aminolevulinic 676 acid dehydratase (Twarog & Cherian, 1983).

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677 678 In another study rats received lead (2 mg/kg as lead acetate) intraperitoneally daily for 679 7 days and 6 days after the last dose unithiol was administered (25-200 µmol/kg). The 680 highest dose of unithiol removed lead from kidneys, liver and bone, while the lower 681 doses (25 and 50 µmol/kg) decreased only the kidney concentrations. Urinary 682 excretion of lead was higher in animals given 100 or 200 µmol/kg of unithiol. Urinary 683 excretion of δ-aminolevulinic acid was unchanged by unithiol administration in this 684 study (Twarog & Cherian, 1984). 685 686 Administration of unithiol (0.5 mmol/kg subcutaneously daily for 4 days) was of benefit 687 in rats poisoned with oral lead (10 mg/kg 6 days a week for 6 weeks). The effect of 688 the antidotes was measured by changes in the concentrations of indicators of lead 689 toxicity (blood δ-aminolevulinic acid dehydratase, zinc , haemoglobin and 690 haemocrit and urinary δ-aminolevulinic acid). Unithiol was shown to reduce lead- 691 induced inhibition of δ-aminolevulinic acid dehydratase and increase blood zinc 692 porphyrin and haemoglobin concentrations. Unithiol decreased lead-induced urinary 693 excretion of δ-aminolevulinic acid. Unithiol also decreased blood, hepatic and renal 694 lead concentrations, but did not mobilise lead from the brain (Sharma et al., 1987). 695 Another study in lead-poisoned rats (20 mg/kg intraperitoneally for 5 days) 696 demonstrated that unithiol (25, 50 or 100 µmol/kg intraperitoneally daily 5 days a 697 week for 7 weeks, starting 3 days after last lead dose) increased urinary excretion of 698 lead (due to mobilisation of lead in bone), and reduced δ-aminolevulinic acid 699 excretion but did not have a beneficial effect on lead-induced anaemia. In addition, 700 the lethality of lead was unaffected by antidote administration (Hofmann & Segewitz, 701 1975). In the study by Llobet et al. (1990) unithiol administration at a dose of 2.90 702 mmol/kg given 10 minutes after intraperitoneal lead (0.58 mmol/kg of lead acetate 703 trihydrate) reduced lethality in mice from 55% to 40%. Unithiol also caused a 704 significant increase in urinary lead and a decrease in kidney lead concentrations. 705 706 In rats with chronic (lead acetate in drinking water, 50 mg/L for 86 707 days) administration of unithiol (0.27 mmol/kg intraperitoneally for up to 4 days) 708 failed to alter lead concentrations in the brain (Aposhian et al., 1996). In a study on 709 acute parenteral lead intoxication in mice antidotes (2 mmol/kg) were injected during 710 3 consecutive days after 7 daily injections of lead (50 mg/kg). Unithiol, sodium 711 calcium edetate (EDTA, calcium disodium edetate, calcium disodium versenate, 712 calcium EDTA) and D-penicillamine did not protect against lethality. Unithiol was, 713 however, more efficient than succimer and several other antidotes including sodium 714 calcium edetate in removing lead from the brain and kidneys (Xu & Jones, 1988). 715 716 Tandon et al. (1994) investigated the efficacy of combined metal-binding agents in 717 lead-poisoned rats (lead acetate 0.1% in drinking water for 8 weeks). Animals were 718 treated with calcium disodium ethylenediaminetetraacetic acid (EDTA sodium calcium 719 edetate, succimer, unithiol, and succimer or sodium calcium 720 edetate and unithiol. All metal-binding agents were given in the same dose (0.3 721 mmol/4 mL/kg intraperitoneally) for 5 days, followed, after a 5 day break by another 5 722 day course. Efficacy was measured by metal concentrations in tissues and 723 biochemical changes (blood δ-aminolevulinic acid dehydratase activity, zinc 724 protoporphyrin, urinary δ-aminolevulinic acid and total urinary proteins). The 725 administration of sodium calcium edetate or succimer resulted in more urinary lead 726 excretion than unithiol. In addition, the combination of sodium calcium edetate and

15

727 succimer was more effective than sodium calcium edetate and unithiol. Both sodium 728 calcium edetate and succimer were more effective than unithiol in reducing lead 729 concentrations in blood, liver, kidney and femur. Only succimer reduced brain lead 730 concentrations. All the metal-binding agents reversed lead-induced inhibition of δ- 731 aminolevulinic acid dehydratase activity and increase in zinc protoporphyrin and 732 urinary excretion of δ-aminolevulinic acid, but the effect was greater with combined 733 therapy. Again the combination of sodium calcium edetate and succimer was more 734 effective than sodium calcium edetate and unithiol. 735 736 A study in rats demonstrated that antidotal therapy in combination with zinc and 737 copper supplementation was more effective at lowering blood lead concentrations 738 than the antidote alone. However, supplementation with unithiol administration (0.3 739 mmol/kg intraperitoneally, daily for 5 days) did not result in an increase in urinary 740 lead concentrations and unithiol was relatively less effective at promoting lead 741 excretion than sodium calcium edetate and succimer. The lead was administered at 742 a dose of 0.05 mmol/kg daily, 6 days/week for 6 weeks (Flora, 1991). 743 744 In mouse cortical cell cultures unithiol was shown to increase the toxicity of lead 745 chloride (Rush et al., 2009). In another in vitro study using human platelets, unithiol 746 was shown to protect against lead-induced stimulation of glutamate binding (Borges & 747 Nogueira, 2008). Unithiol increased lead-induced inhibition of ALAD in human blood 748 in vitro. In addition the inhibition of ALAD activity in the blood and liver of lead- 749 exposed mice was also increased by unithiol treatment (Santos et al., 2006). 750 751 8.1.11 Mercury 752 753 8.1.11.1 Inorganic mercury 754 755 Unithiol has been shown to increase mercury excretion and decrease tissue 756 concentrations in experimental animals. Most studies found that unithiol does not 757 increase faecal excretion of mercury (Aaseth et al., 1982; Kachru & Tandon, 1986). 758 However some studies have shown the opposite (Wannag A. & Aeaseth J., 1980), 759 but this may be an effect of decreased body burden of mercury with unithiol 760 treatment (Gabard, 1976a). In addition, although some studies have found that 761 unithiol mobilises mercury from the brain, others have found that unithiol is relatively 762 ineffective in reducing brain mercury concentrations. The former observation may be 763 due to contamination of brain tissue with blood which has a higher mercury 764 concentration (Buchet & Lauwerys, 1989). 765 766 In rats with mercury toxicity (5 µmol/kg as mercuric chloride) unithiol (400 μmol/kg 767 intramuscularly) was found to be the most effective antidote compared to sodium 768 diethyldithiocarbamate (DDC) and (diethylenetriaminepentaacetic acid, 769 DTPA) when given prior to exposure. Unithiol was shown to mobilise mercury in 770 the urine and reduce concentrations in the liver and spleen. It did not increase 771 faecal excretion (Kachru & Tandon, 1986). Treatment with unithiol (500 µmol/kg 772 intravenously) 24 hours after mercury administration (2 μmol/kg intravenously as 773 mercuric chloride) in mice reduced the mercury content of the kidneys to 60-70% of 774 controls by 48 hours after administration of the antidote (Wannag & Aaseth, 1980). 775 776 Immediate treatment with unithiol (500 µmol/kg intravenously) after mercury

16

777 administration (5 µmol//kg intravenously as mercuric chloride) in rats prevented 778 pathological changes in the kidneys and increased mercury excretion in the urine. 779 When administration was delayed 24 hours, unithiol was relatively ineffective in 780 reversing mercury-induced anuria. The kidney mercury concentration was significantly 781 reduced, but pathological changes could not be prevented. Mercury concentrations in 782 the blood, kidney and brain were reduced irrespective of whether the administration of 783 unithiol was immediate or delayed. Immediate unithiol treatment did not change faecal 784 mercury elimination whereas delayed administration resulted in increased faecal 785 excretion (Wannag & Aaseth, 1980). 786 787 Administration of oral unithiol (30 µmol/200 g) was shown to normalise renal excretion 788 of alkaline phosphatase in mercury-poisoned rats (0.75 mg/kg as intravenously 789 mercuric chloride). The dose of unithiol was given at 6 or 24 hours after mercury and 790 then once a day until the fifth day. Early administration of unithiol also abolished the 791 effect of mercury on renal excretion of leucine aminopeptidase, but there was no effect 792 on this enzyme if administration of unithiol was delayed until 24 hours after mercury 793 exposure. Furthermore, early treatment with unithiol reduced mercury-induced 794 lethality, whereas delayed treatment had no effect (Planas-Bohne, 1977). 795 796 In a study reported by Aaseth (1983), succimer or unithiol (1 mmol/kg daily for 4 797 days) given after a single intravenous dose of mercuric chloride (2 μmol/kg) to mice 798 reduced the renal mercury concentration almost by a factor of three. Brain mercury 799 concentrations were also reduced. Dimercaprol, in contrast, has long been known to 800 increase mercury deposition in the brain after exposure to methyl mercury in mice 801 (Berlin & Ullberg, 1963). 802 803 The efficacy of unithiol (220 mg/kg) and succimer (180 mg/kg) were compared in rats 804 exposed to mercury (0.5 mg/kg as mercuric chloride intraperitoneally 5 times a week 805 for 3 weeks). The antidotes were administered 7 days after the last mercury dose. 806 Both antidotes were ineffective in removing mercury from the brain and unithiol was 807 more effective in increasing urinary mercury excretion. This was also the case when 808 the same dose of mercury was administered as phenyl mercury (Buchet & Lauwerys, 809 1989). In a study of various antidotes given in mercury-poisoned mice (as mercuric 810 chloride 10 mg/kg), where the antidotes were given in doses of antidote:mercury molar 811 ratios of 10, 15, 20 and 30:1, unithiol reduced lethality and was significantly more 812 effective than succimer or dimercaprol. This was true for unithiol even at the smallest 813 dose ratio of 10:1 (Jones et al., 1980). 814

815 Comparing succimer (100 µmol/kg orally) and unithiol (300 µmol/kg orally) in mercury- 816 poisoned rats (0.5 mg as mercuric chloride) Planas-Bohne (1981) found that the rise in 817 urinary mercury excreted in the succimer treated animals corresponded to the mercury 818 content of the kidneys. In contrast, the urinary mercury concentration in the unithiol 819 treated group was higher indicating removal of mercury from other organs. The dose 820 of unithiol was three times that of succimer because only 30% of the unithiol was 821 absorbed from the gut compared with 100% of the succimer. However, Cherian et al. 822 (1988) reported that the increase in urinary excretion induced by unithiol (0.2-2.0 823 mmol/kg intraperitoneally) was almost equal to the amount of mercury lost from the 824 kidneys in mercury-poisoned rats (0.1-2 mg/kg intraperitoneally or mercury vapour 0.5- 3 825 2 mg/m ). 826

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827 In rats with chronic (0.5 mg/mercury/kg, as mercuric chloride, 828 intraperitoneally 5 days a week for 32 or 41 days) unithiol (0.27 mmol/kg 829 intraperitoneally for up to 4 days) failed to alter mercury concentrations in the brain 830 (Aposhian et al., 1996). 831 832 Both renal and biliary excretion of mercury were increased after administration of

833 unithiol (12.5 mg intramuscularly) in mercury-poisoned rats (120 µg intravenously as 203 834 mercuric chloride). The mercury content was decreased in all tissues, especially in 835 the kidneys and the brain. Treatment with the diuretic before 836 administration of mercury further increased the biliary excretion of mercury observed

837 with unithiol. However, the overall excretion of mercury was unchanged (Cikrt, 1978). 838 In a similar study comparing unithiol (15 mg/kg intramuscularly) and unithiol combined 839 with spironolactone and oral polythiol resin in mercury-poisoned rats (same dose as 840 above) Cikrt & Lenger (1980) found that urinary excretion was higher with unithiol 841 alone but the combination therapy resulted in increased faecal excretion. Both 842 regimens significantly decreased the mercury content of all tissues. 843 844 Of 15 antidotes studied in mercury-poisoned mice (0.5 mg mercury intravenously 845 mercuric chloride) only unithiol (50 µmol/kg) was shown to have a favourable effect 846 with increased urinary mercury excretion and reduced tissue concentrations 847 (erythrocytes, plasma, liver, kidneys, brain, femur, muscle, spleen and intestine). 848 Unithiol reduced the mercury concentration of the kidneys to approximately 40% of 849 controls and the other organs to 50-80%. Faecal mercury excretion was decreased 850 but this was due to a decreased body burden, and overall elimination was increased 851 due to a large rise in the urinary mercury concentration (Gabard, 1976a). Similarly, 852 oral unithiol (1 mmol/kg/day for 4 days) reduced the mercury concentration in the 853 kidney to about 30% of controls in mercury-poisoned mice. Dosing with unithiol was 854 started immediately after intravenous injection of mercury (2 µmol/kg as mercuric 855 chloride). There was increased urinary excretion of mercury but faecal excretion 856 was unchanged (Aaseth et al., 1982). 857 858 In a study in mercury-poisoned mice (5, 200, 300 or 400 µmol/kg of mercuric chloride 859 orally) unithiol (100, 800, 1200 or 1600 µmol/kg orally) given 15 minutes later was 860 most effective at reducing lethality. In addition oral administration was more effective 861 than parenteral, probably because of reduction in gastrointestinal absorption of 862 mercury. Unithiol also reduced mercury concentrations in the brain; intraperitoneal 863 unithiol reduced mercury brain concentrations to about one third of controls whereas 864 oral administration reduced concentrations to less than 15% of controls (Nielson & 865 Andersen, 1991). 866 867 Simultaneous administration of sodium selenite and mercuric chloride decreased the 868 efficacy of unithiol (300 µmol/kg orally) on mercury elimination in rats. The metal salts 869 were given at a dose of 1.5 µmol/kg by intraperitoneal injection. When both metal 870 salts were given together there was redistribution of mercury with reduced 871 accumulation in the kidneys (decreased by more than 94%) and an increased 872 concentration in the liver (increased about 6 times). Urinary excretion of mercury was 873 also reduced compared to elimination in the absence of (Jureša et al., 2005). 874 875 Kostial et al. (1984) investigated the influence of age on unithiol efficacy in rats (aged 876 2, 6 and 28 weeks old) with mercury toxicity (50 μg/kg intraperitoneally). Unithiol (50

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877 mg/kg 3 times, 1 day after mercury administration and then at 24 hour intervals) 878 decreased the body retention of mercury in all age groups, and was about twice as 879 effective in adults compared to suckling rats. The reduced effectiveness was due to 880 the reduced efficacy of unithiol in lowering kidney retention in young animals. This 881 age difference was also confirmed in a later study (Kostial et al., 1991). In addition 882 this study found that early treatment with oral unithiol in older rats given oral mercury 883 increased mercury retention. However, this was in contrast to succimer, where early 884 treatment while mercury was still in the gut decreased mercury retention. 885 886 An in vitro study on isolated perfused segments of rabbit proximal tubules exposed to 887 inorganic mercury found that mercury was rapidly taken up by the tubular epithelial 888 cells and resulted in cellular necrosis. The addition of unithiol provided complete 889 protection against this effect. This appeared to be due to a negligible rate of net 890 absorption of inorganic mercury ions from the lumen and low levels of ion 891 accumulation. Unithiol-mercury complexes are not readily transported into proximal 892 tubular cells and it is thought that unithiol reduces the renal mercury burden by 893 extraction of mercury during the trans-epithelial transport of unithiol. The therapeutic 894 efficacy of unithiol in mercury-induced renal damage may be linked to its transport at 895 the basolateral membrane by the organic anionic transporter (OAT) system and to 896 prevention of significant uptake of mercury by the proximal tubular cells due to the 897 formation of unithiol-mercury complexes (Zalups et al., 1998). Another in vitro study 898 confirmed that unithiol was effective at inhibiting mercury accumulation in renal 899 proximal and distal tubular cells, and protecting against mercury-induced renal 900 damage (Lash et al., 1998). 901 902 Unithiol, when incubated with mercuric chloride, partially restored cellular morphology, 903 viability, intracellular adenosine triphosphate (ATP) concentrations and mitochondrial 904 membrane potential in opossum kidney cells in vitro. Unithiol also had a protective 905 effect on mitochondrial morphology and showed potent antioxidant activity (Carranza- 906 Rosales et al., 2007). Similarly, unithiol was shown to reduce mercuric chloride 907 toxicity in mouse cortical cell cultures (Rush et al., 2009). In another in vitro study 908 using human platelets, unithiol was shown to protect against the inhibitory effect of 909 mercury on glutamate binding (Borges & Nogueira, 2008). 910 911 8.1.11.2 Organic mercury 912 913 Oral unithiol administration (1 mmol/kg) to rats with methyl mercury poisoning (0.23 914 mg/kg intravenous as methyl mercury) reduced the biological half-life of the mercury 915 body burden from 23.0 days to 4.3 days compared to controls. The mercury 916 concentration was decreased in all tissues, particularly in the kidneys and the brain 917 (Gabard, 1976b). 918 919 The effect of unithiol on motor impairment and cerebellar toxicity has been studied in 920 mercury-exposed mice. Methylmercury was given in drinking water (40 mg/L) for 17 921 days with unithiol given on days 15 to 17 (150 mg/kg by intraperitoneal injection). The 922 mice were tested for signs of toxicity 24 hours after the last dose of unithiol. The 923 mercury-induced motor deficit was reduced in unithiol treated mice and unithiol also 924 reduced mercury-induced lipid peroxidation in the cerebellum but did not prevent 925 mercury-induced reduction of cerebellar glutathione peroxidase. Unithiol significantly 926 reduced mercury deposition in the cerebellar cortex (Carvalho et al., 2007).

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927 928 Unithiol is able to increase elimination of both organic and inorganic mercury from 929 tissue, however with organic mercury the efficacy of unithiol is affected by the 930 relative capacity of tissue for dealkylation of organic to inorganic mercury. In rats 931 with chronic methyl mercury poisoning (10 ppm in drinking water for 9 weeks in a 932 single dose study and 6 weeks in a repeated dose study) a single dose of unithiol 933 (100 mg/kg intraperitoneally) was shown to reduce kidney inorganic and organic 934 mercury concentrations by 38% and 59%, respectively. In addition, urinary inorganic 935 and organic mercury concentrations increased by 7.2 and 28.3 fold, respectively, 936 compared with pre-treatment concentrations. A single dose of unithiol was relatively 937 ineffective at removing mercury from the brain and the mercury in this tissue was 938 predominantly in the organic form due to slow dealkylation to inorganic mercury. In 939 contrast, repeated doses of unithiol (100 mg/kg every 72 hours for 1, 2 or 3 doses) 940 significantly decreased mercury concentrations in the brain, kidney and blood, but 941 the decrease in the brain and blood was restricted to the organic mercury 942 component. This was thought to reflect the slow dealkylation of methyl mercury in 943 these tissues. This may be why unithiol is relatively ineffective at improving organic 944 mercury-induced neurotoxicity (Pingree et al., 2001). 945 946 Unithiol no effect on methylmercury and increased the toxicity of ethylmercury in 947 mouse cortical cell cultures (Rush et al., 2009). 948 949 8.1.12 950 951 Unithiol has been shown to be efficacious in experimental nickel poisoning in 952 animals, in terms of increased survival and reduced nickel-induced toxic effects. 953 954 Unithiol administered intraperitoneally at a 10:1 mole ratio of antidote to nickel, 955 increased survival rate in mice poisoned with intraperitoneal nickel acetate (62 956 mg/kg). Antidotes were administrated 20 minutes after injection of nickel. Unithiol 957 was less effective than D-penicillamine or sodium calcium edetate, although the 958 small sample size prohibited any significant differentiation between them (Basinger 959 et al., 1980). 960 961 Administration of unithiol (0.5 mmol/kg subcutaneously daily for 4 days) significantly 962 enhanced the urinary excretion of nickel in rats poisoned with intraperitoneal nickel 963 sulphate (4 mg/kg 6 days/week for 4 weeks). In unithiol treated animals there were 964 significant reductions in nickel concentrations in renal and hepatic tissue and 965 decreases in evidence of nickel-induced kidney and liver damage (as measured by 966 plasma concentrations of ceruloplasmin and amino acids, blood glucose and 967 glutathione, urinary amino acids, and renal, hepatic and cardiac concentrations of 968 mitochondrial malic dehydrogenase, which is inhibited by nickel). Unithiol reduced the 969 increase in plasma and urinary amino acids and blood glucose, but did not change the 970 decreases in plasma ceruloplasmin and blood glutathione concentrations. There was 971 no significant effect on mitochondrial malic dehydrogenase concentrations. Faecal 972 nickel excretion was unchanged and unithiol was ineffective at mobilising nickel from 973 the brain (Sharma et al., 1987). 974 975 Rats given nickel (1.5 mg/kg intraperitoneally as nickel sulphate daily, 6 days a week 976 for 30 days) then unithiol (0.3 mmol/kg intraperitoneally daily for 5 days) had their

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977 organs harvested 24 hour after the last injection. Unithiol reduced the nickel 978 concentrations in the liver, blood, heart and kidney but not in the brain. Unithiol also 979 reversed nickel-induced biochemical changes (decreased ceruloplasmin 980 concentration and blood glucose; increased plasma and urine concentrations of 981 amino acids) and increased faecal nickel excretion (Tandon et al., 1996). 982 983 8.1.13 Palladium 984 985 There is limited information on the effect of unithiol on palladium toxicity. It did not 986 influence toxicity or reduce lethality in mice with acute palladium chloride poisoning 987 (586 µmol/kg intraperitoneally). Unithiol was given subcutaneously at the same time 988 as palladium at a dose of 2.93 mmol/kg (5 times the molar dose of metal) (Mráz et 989 al., 1985). 990 991 8.1.14 Platinum 992 993 There is limited information on the effect of unithiol on platinum toxicity. A single 994 injection of unithiol (1 mmol/kg) produced no significant change in renal platinum 995 concentration in rats treated with cisplatin (4 or 6.5 mg/kg) 24 hours previously. 996 After four daily treatments unithiol and succimer caused a significant increase in 997 urinary excretion of platinum, but this was low, and represented only about 3% of 998 the injected dose of platinum. It was concluded that none of the antidotes studied, 999 unithiol, succimer or pentetic acid, were likely to be of benefit in the management of 1000 cisplatin-induced renal toxicity (Planas-Bohne et al., 1982). 1001 1002 8.1.15 1003 1004 Several animal studies have shown that although unithiol can remove polonium-210 1005 from most tissues it results in concentration of polonium in the kidneys, with the risk 1006 of renal damage. In addition, unithiol has been shown to cause renal damage when 1007 administered to polonium-poisoned animals (Poluboiarinova & Streltsova, 1964). 1008 1009 Rats were given approximately 0.3 µCi of polonium-210 intravenously followed 1.5 1010 minutes later by intraperitoneal or oral administration an antidote (1 mmol/kg). The 1011 α-activity of tissue was determined 48 hours later. Administration of an antidote 1012 produced a marked decrease in polonium-210 retention in the blood, spleen and 1013 bone and, to a lesser extent, in the plasma. Unithiol but did not decrease polonium- 1014 210 retention in the kidneys and overall retention of polonium-210 in the blood, liver, 1015 spleen, skeleton and kidneys was increased by 40%. The effect of oral unithiol was 1016 approximately one third of the intraperitoneal dose, and following administration by 1017 this route the overall retention of polonium-210 was increased by approximately 1018 50%. Retention of polonium-210 was particularly seen in the kidneys where it was 1019 transported but not excreted. In view of these findings it was concluded that 1020 although unithiol increased survival of rats given lethal doses of polonium-210, it was 1021 not a suitable antidote for polonium because it could potentiate the toxic effects of 1022 polonium on the kidneys (Volf, 1973). 1023 1024 In a study of rats given a lethal dose (40 µCi) of polonium-210 by intraperitoneal 1025 injection, administration of an antidote (0.2 mmol/kg) 1 minute, 90 minutes, 360 1026 minutes and twice daily on days 2, 3, 4, 12, 22 and 32 increased the mean survival

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1027 time from 39 days to 106 days. Unithiol significantly decreased the polonium-210 1028 content of all tissues studied except the kidneys. However, DMPA was found to be a 1029 more effective antidote and able to decrease polonium-210 in all tissues (Aposhian 1030 et al., 1987). 1031 1032 The study by Rencová et al. (1993) comparing a number of antidotes for polonium- 1033 210 also found that unithiol reduced retention in the bone, spleen and blood of rats 1034 but increased it in the kidneys. Indeed, the total body retention of polonium-210 1035 could not be reduced to less than 85% of controls with any of the 9 antidotes used. 1036 1037 The same investigators (Volf et al., 1995) looked at the use of antidotes in rats with 1038 simulated wounds contaminated with polonium-210. After 2 weeks of unithiol 1039 treatment (intramuscularly at injection site at 1 hour and 5 days, and systemic 1040 treatment with intramuscular injection on days 1, 7, 9 and 12) polonium-210 at the 1041 wound site was reduced to 12% of controls. The retention in the liver, spleen, 1042 muscle and skeleton was reduced to 14-40%, but the blood content was unchanged 1043 and retention in the kidneys was increased to 340% of controls. Unithiol was 1044 effective at removing polonium-210 from the wound site when given either locally or 1045 systemically, but it was not effective at removing polonium-210 from the body as a 1046 whole. In a series of other experiments it was concluded that treatment with unithiol 1047 combined with another antidote (particularly sodium N,N’-di-(2-hydroxyethyl)- 1048 ethylenediamine-N’N’-biscarbodithioate; HOEtTTC) was the most effective method of 1049 removing polonium-210 from the body. 1050 1051 8.1.16 Selenium 1052 1053 There is limited information on the effect of unithiol on selenium toxicity. Unithiol 1054 administration (60 mg/kg intraperitoneally) had no effect on selenium-poisoned rats 1055 (2.24 mg of selenium/kg by subcutaneously). The concentration of selenium in urine 1056 and faeces was unchanged (Paul et al., 1989). 1057 1058 8.1.17 Silver 1059 1060 There is limited information on the effect of unithiol on silver toxicity. Unithiol 1061 administration (0.7 mmol/kg intraperitoneally) increased the LD50 of silver chloride in 1062 mice from 13.6 to 74 mg/kg (Pethran et al., 1990). In dogs given intravenous silver 1063 nitrate unithiol prevented the development of toxic pulmonary oedema and death 1064 (Romanov, 1967). 1065 1066 In an in vitro study of silver inhibition of Na,K-ATPase, which was probably due to 1067 deposition of the metal on sulphydryl groups in the enzyme, it was found that the 1068 process was completely reversed by administration of unithiol (Hussain et al., 1994). 1069 1070 8.1.18 Strontium 1071 1072 There is limited information on the effect of unithiol on strontium toxicity. Unithiol did 1073 not affect survival rate in mice poisoned with strontium chloride (Domingo et al., 1990; 1074 Pethran et al., 1990). 1075 1076 8.1.19

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1077 1078 Unithiol is not of benefit in thallium poisoning in experimental animals. 1079 1080 In a study comparing the efficacy of (potassium ferric hexacyanoferrate 1081 II) and unithiol, rats were given an oral dose of 20 mg thallium (as thallium sulphate) 1082 and antidotal therapy was started 24 hours later. The rats were divided into 3 1083 treatment groups: Prussian blue 50 mg/kg for 4 days, unithiol 5 mg/kg 6 times a day on 1084 day 1, 4 times on day 2 and 3 times on days 3 and 4. In the third group the animals 1085 were given both antidotes. Animals were killed on day 5 and the thallium content of 1086 organs determined. Prussian blue administration limited thallium distribution into 1087 tissues, whereas unithiol did not decrease thallium concentration in any organ, 1088 although it did decrease thallium concentrations in blood. The two drugs in 1089 combination decreased the thallium content in all organs but no more than Prussian 1090 blue alone. It was concluded that unithiol is not a useful antidote in thallium poisoning 1091 (Mulkey & Oehme, 2000). 1092 1093 Similarly, unithiol did not affect survival rate in mice given thallium sulphate (Pethran et 1094 al., 1990). 1095 1096 8.1.20 Tin 1097 1098 There is limited information on the effect of unithiol on tin toxicity. Unithiol and 1099 succimer have been investigated as antidotes in rats following a single intravenous 1100 dose dibutyltin dichloride (27 µmol/kg). The antidotes were given at two doses, 100 1101 and 500 µmol/kg, orally and by intraperitoneal injection. Several parameters of 1102 organ toxicity were monitored from 6 hours to 8 weeks. Both drugs reduced 1103 dibutyltin dichloride-induced lesions of the bile duct, pancreas and liver. Unithiol was 1104 more effective than succimer in most measured parameters and the drugs were 1105 though to exert their protective effects on these organs by reducing biliary organotin 1106 excretion (Merkord et al., 2000). 1107 1108 In vitro studies with human erythrocytes incubated with tributyl tin have 1109 demonstrated that unithiol is unable to prevent the tributyl tin-mediated haemolysis of 1110 the cells (Gray et al., 1986; 1987). 1111 1112 8.1.21 Vanadium 1113 1114 There is limited information on the effect of unithiol on vanadium toxicity. However, it is 1115 unlikely to be useful since antidotes containing oxygen, rather than thiol groups such as 1116 unithiol, are more effective at binding vanadium. 1117 1118 Unithiol had no effect on lethality in mice poisoned with sodium vanadate (50 mg/kg 1119 intraperitoneally) or vanadyl sulphate (110 mg/kg intraperitoneally). Unithiol was 1120 given intraperitoneally 20 minutes after administration of vanadium, at a dose of 5:1 1121 antidote to metal compound (Jones & Basinger, 1983). 1122 1123 Unithiol had no significant effect on the death rate, body weight reductions, or 1124 reduction in weights of legs and toes in chick eggs incubated with vanadium 1125 (Hamada, 1994). 1126

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2+ 1127 A recent study, however, has shown that oxovanadium (VO ) is capable of forming a 4- 1128 [VO(DMPS)2] complex with unithiol in aqueous solution (Williams & Baran, 2008). 1129 The significance of this in vivo remains to be elucidated. 1130 1131 8.1.22 Zinc 1132 1133 Unithiol can increase excretion of zinc and reduced lethality in zinc-poisoned animals, 1134 but more effective antidotes are available. 1135 1136 In a comparison of several antidotes against the effects of acute parenteral zinc 1137 intoxication in mice, unithiol efficiently reduced acute lethality. Succimer (10:1 1138 antidote:zinc ratio) was given 20 minutes after a fatal dose of zinc (50 mg/kg) was 1139 administered. Survival was 73% with unithiol but other antidotes were equally or 1140 more effective (Basinger & Jones, 1981b). 1141 1142 In mice given zinc acetate (0.49 mmol/kg intraperitoneally) unithiol (2:1 or 5:1 molar 1143 ratios of antidote to metal) was the least effective of the six antidotes tested. Disodium 1144 calcium cyclohexanediaminetetraacetate (CDTA), sodium calcium edetate and 1145 pentetic acid and were the most effective (Llobet et al., 1988). 1146 1147 In a study comparing the efficacy of several antidotes mice were given intraperitoneal 1148 zinc acetate (66-330 mg/kg; LD50 108 mg/kg). Antidotal therapy was given 10 minutes 1149 later, also by intraperitoneal injection. Unithiol reduced the lethality in animals given 1150 66-241 mg/kg of zinc acetate. In those given 330 mg/kg of zinc acetate lethality was 1151 30% compared to 100% in control animals. Unithiol also increased renal excretion of 1152 zinc and reduced blood and heart tissue concentrations compared to controls. 1153 However, pentetic acid and CDTA were found to be more effective zinc antidotes in 1154 this study (Domingo et al., 1988). 1155 1156 8.2 1157 1158 There is information on unithiol pharmacokinetics in various species of experimental 1159 animals. 1160 1161 Following oral administration between 60% (dogs; Wiedemann et al., 1982) and 30% 1162 (rats; Gabard, 1978) of the dose is absorbed and plasma peak concentrations are 1163 reached after 30 to 45 minutes. Plasma protein binding was measured at 70% in dogs 1164 by equilibrium dialysis (Wiedemann et al., 1982). This is in agreement with the figure 1165 of 65-75%, determined in rats (Planas-Bohne & Lehmann, 1983). 1166 1167 After intravenous administration unithiol is mainly distributed in plasma and kidneys, 1168 only minor concentrations were measured in the brain and other organs. The 1169 apparent volume of distribution in dogs was calculated to be 0.160 mL/kg (Wiedemann 1170 et al., 1982). Most unithiol excreted in the bile of rats is in its altered form and 1171 recovery in bile was 40% of the administered dose (Zheng et al., 1990). 1172 1173 Although unithiol is mainly distributed to the extracellular compartment, studies have 1174 demonstrated uptake by cells. Planas-Bohne & Olinger (1981) studying the 1175 interaction of antidotes with methyl mercury bound to erythrocytes demonstrated that 1176 unithiol is lost from the extracellular fluid. Wiedemann et al. (1982) found that the

24

1177 distribution volume of radiolabelled unithiol exceeded the extracellular volume in dogs. 1178 In vitro studies have demonstrated that unithiol is taken up by cells of the basolateral 1179 membrane of the kidney and can bind mercury within cells (see section 7). 1180 1181 Unithiol is rapidly eliminated from the body. The serum half life is approximately 20 to 1182 60 minutes and plasma clearance is approximately 2.6 mL/min/kg. In dogs given 1183 radiolabelled unithiol 93% was eliminated within 3 days, with the bulk (98%) eliminated 1184 in the urine and the rest in the faeces (Wiedemann et al., 1982). 1185 1186 Gabard & Walser (1979) stated that unithiol is not involved in important metabolic 1187 pathways and a quantity is excreted unchanged; Maiorino et al. (1988) demonstrated 1188 the presence of several acyclic and cyclic oxidized metabolites in the urine of rabbits. 1189 1190 8.3 1191 1192 Unithiol is of low acute and chronic toxicity. Studies have determined the LD50 for 1193 various species and examined the effect of acute or chronic unithiol administration on 1194 trace element concentrations. 1195 1196 In order of decreasing sensitivity to unithiol, animal species can be ranked as follows: 1197 cat, dog, guinea pig, rabbit, rat and mouse (Klimova, 1958; Aposhian, 1983). The 1198 optical isomers of unithiol do not differ in their toxicity (Hsu et al., 1983). 1199 1200 8.3.1 Acute toxicity 1201 1202 Unithiol is of relatively low toxicity. By the parenteral route the acute LD50 of unithiol 1203 for various species is about 1 to 2 g/kg (Planas-Bohne et al., 1980; Aposhian et al., 1204 1981; Aposhian, 1982; Hruby and Donner, 1987; Pethran et al., 1990). After 1205 intraperitoneal injection of lethal doses rats are highly irritable for some minutes before 1206 they become apathetic followed by cessation of breathing and death within 12 hours. 1207 The LD50 for a single dose in rats was 5 mmol/kg (1.14 g/kg) and after dosing on 10 1208 consecutive days was 30.8 mmol/kg (Planas-Bohne et al., 1980). No acute toxicity 1209 was observed in mice receiving oral unithiol (100, 300 or 600 µg/mL) for 5 days in 1210 drink water. These doses are equivalent to 18, 55 and 109 mg/kg/day or 0.1, 0.3 1211 and 0.5 mmol/kg/day (Jones et al., 1996). 1212 1213 In rats treated with a single intraperitoneal dose of 1 mmol/kg of unithiol, urinary zinc 1214 and copper excretion was increased whereas the excretion of and manganese 1215 was unchanged (Gabard et al., 1979). 1216 1217 A single subcutaneous injection of unithiol (1.6 mmol/kg) in mice caused a significant 1218 increase in δ-aminolevulinic dehydratase activity in the blood and a decrease in the 1219 kidney, with no change in the liver or brain. Unithiol also significantly increased the 1220 zinc concentration of the kidneys but did not change liver or brain concentrations. 1221 There was also a significant increase in liver and kidney lipid peroxidation (Santos et 1222 al., 2005). 1223 1224 8.3.2 Chronic toxicity 1225 1226 Unithiol is of relatively low toxicity even in chronic administration. Hrdina et al. (1998)

25

1227 studied the effects of repeated injection of unithiol on the heart of rabbits. The animals 1228 were given intravenous unithiol, 50 mg/kg once a week for 10 weeks. There was no 1229 change in iron or selenium concentrations. There was a slight decrease in myocardial 1230 concentrations of calcium, potassium and magnesium, but only the later was 1231 significantly different. These changes were not associated with any haematological, 1232 histological or physiological changes. 1233 1234 Mice receiving unithiol 300 µg/mL in drinking water for 3 months showed no signs of 1235 toxicity. Haematological and biochemical parameters were also unchanged (Jones 1236 et al., 1996). 1237

1238 Rats receiving 600 µmol/kg/day orally (126 mg/kg/day) on 5 days per week for 66 1239 weeks did not show any adverse effects. Treatment for 36 weeks led to a reduction in 1240 copper-concentrations in the kidneys, liver and skin (Planas-Bohne et al., 1980). 1241 Beagle dogs treated for 6 months with doses up to 15 mg/kg/day intravenously or 45 1242 mg/kg/day orally showed no significant changes in blood-concentrations of glucose, 1243 uric acid, creatinine, total protein, sodium, potassium, calcium, magnesium and iron, 1244 and activity of liver enzymes and in the serum. In addition, the red and 1245 white blood picture as well as gain in body weight remained unchanged. A dose- 1246 dependent decrease in the copper content was found in the serum, liver, kidney and 1247 spleen. The macroscopic and microscopic examination of several organs revealed no 1248 pathological changes. After treatment for 10 weeks at a dose of 2 x 75 mg/kg/day 1249 intravenously the following changes were noted: a depletion of copper in the serum 1250 and in various organs, an increase of the iron content of the liver and spleen, and a 1251 decrease in haemoglobin, haematocrit, red blood cells, alkaline phosphatase activity 1252 and zinc content in the blood (Szincicz et al., 1983). 1253 1254 8.3.3 Reproductive toxicity and teratogenicity 1255 1256 Unithiol does not appear to produce reproductive toxicity or teratogenicity. 1257 1258 No teratogenic effects were reported in the offspring of rats given 600 µmol/kg/day 1259 orally (126 mg/kg/day) on 5 days per week. Female rats were mated with untreated 1260 males after 14, 26 or 60 weeks of treatment with unithiol. Treatment was continued 1261 during and nursing. The number of pregnant animals and the litter size 1262 was smaller in the treated group but the difference was not significant (Planas-Bohne 1263 et al., 1980). 1264 1265 No adverse effects were observed in mothers or offspring in mice given up to 630 1266 mg/kg/day in two dosing regimens: from gestation day 14 until birth or from gestation 1267 day 14 until post-natal day 21. The no observed effect level (NOEL) of 630 1268 mg/kg/day is much higher than that used in the treatment of human heavy metal 1269 poisoning (Domingo et al., 1990). 1270 1271 Mice given unithiol, up to 300 mg/kg, on days 6 to 15 of gestation showed no 1272 maternal or reproductive effects. Unithiol had a minor effect on maternal and fetal 1273 metabolism which was variable and not dose related. These effects did not 1274 produce maternal or embryofetal toxicity (Bosque et al., 1990). 1275 1276 No teratogenic effects were reported in rabbits given unithiol (up to 100 mg/kg

26

1277 intravenously daily) from days 6 to 18 of gestation (Anon, 1992/1993). 1278 1279 Unithiol has been shown to protect against the developmental toxicity of arsenic 1280 (Domingo et al., 1992) and mercury (Gomez et al., 1994) in experimental animals. 1281 Mice were given a single intraperitoneal injection of sodium arsenite on day 9 of 1282 gestation followed by immediate injection of dimercaprol or unithiol with further doses 1283 at 24, 48 and 72 hours. Dimercaprol did not protect against arsenic-induced 1284 developmental toxicity, whereas unithiol was protective at 150 and 300 mg/kg/day 1285 and was able to prevent embryotoxicity and fetotoxicity. The higher dose also 1286 prevented maternal arsenic toxicity (Domingo et al., 1992). Pregnant mice were 1287 given a single oral dose of 30 mg/kg of methyl mercury chloride on day 10 of 1288 gestation followed by dimercaprol (by subcutaneous injection) or unithiol (by gavage) 1289 at 24, 48 and 72 hours. Dimercaprol administration did not prevent maternal or 1290 developmental toxicity, whereas unithiol in doses up to 360 mg/kg/day significantly 1291 reduced maternal lethality. Treatment with the higher doses, 180 and 360 1292 mg/kg/day, also protected against mercury-induced embryotoxicity and teratogenicity 1293 (Gomez et al., 1994). 1294 1295 8.3.4 Genotoxicity 1296 1297 Unithiol has been evaluated for mutagenicity in the Ames test with negative results 1298 (Aposhian et al., 1983; Ruprecht, 1997). Normal DNA synthesis was maintained with 1299 unithiol concentrations up to 8 µg/mL in an in vitro study using 3 murine tumour cell 1300 lines. Above 8 µg/mL unithiol inhibited DNA synthesis, and above 30 µg/mL inhibition 1301 exceeded 80% (Jones et al., 1996). 1302 1303 An in vitro study found that unithiol increased the incidence of nickel-induced DNA 1304 breaks in a human leukaemia cell line. There was also an increase in DNA breaks in 1305 bacterial plasmids (a simpler system). Succimer and dimercaprol also increased 1306 DNA breaks in plasmids in the presence of nickel, but the effect was strongest with 1307 succimer. These metal binding agents all generate hydrogen peroxide in solution 1308 but succimer is the most potent. Free radicals are throught to be involved in the 1309 DNA damage observed. For the most potent compound, succimer, the breakage of 1310 DNA was completely prevented by the presence of mannitol and partially reduced by 1311 antioxidants. This protective effect was not investigated for unithiol (Lynn et al., 1312 1999). 1313 1314 1315 9. Volunteer studies 1316 1317 There are three main studies of unithiol pharmacokinetics, all conducted by the same 1318 group, consequently some volunteers took part in more than one study. The 1319 characteristics of each study are as follows: 1320 • The study by Maiorino et al. (1991) involved 10 male volunteers, aged 24-34 years, 1321 weighing 68-98 kg. 1322 • The study by Hurlbut et al. (1994) involved 5 volunteers (4 male, 1 female), aged 24 1323 to 32 years, weighing 49-93 kg. 1324 • The study by Maiorino et al. (1996) involved 4 male volunteers, aged 23 to 27 1325 years, weighing 86-91 kg. 1326

27

1327 9.1 Absorption 1328 1329 Unithiol is rapidly absorbed. In volunteers given 3 x 100 mg capsules, unithiol was 1330 detected in blood within 0.5 to 4 hours after ingestion. Maximal concentrations were 1331 reached within 3 to 4 hours (Maiorino et al., 1991). 1332 1333 In 4 male volunteers the oral bioavailability of unithiol was calculated to be 39%, with a 1334 range of 19-62% (Hurlbut et al., 1994). 1335 1336 9.2 Distribution 1337 1338 In volunteers given 3 x 100 mg capsules, metabolites (altered unithiol) were confined 1339 to the plasma portion of the blood suggesting that they were bound to plasma 1340 proteins (Maiorino et al., 1991). Plasma protein binding of unithiol was approximately 1341 90% when measured by equilibrium dialysis in human plasma samples from 3 1342 volunteers (Wiedemann et al., 1982). However in a further study by Maiorino et al. 1343 (1996), less than 1% of unithiol was present in an unaltered form in the plasma 5 1344 hours after a single oral dose of 300 mg. The protein-bound unithiol and non-protein- 1345 bound unithiol disulphides were present as 62.5% and 36.6% of the total unithiol, 1346 respectively. The protein-bound unithiol was present as a unithiol-albumin complex 1347 (84%) and a higher molecular weight complex (16%). 1348 1349 In volunteers given unithiol 3 mg/kg intravenously over 5 minutes the volume of 1350 distribution varied from 2.67 to 15.4 L/kg (Hurlbut et al., 1994). 1351 1352 Although unithiol is mainly distributed to the extracellular compartment, studies have 1353 demonstrated uptake by cells. An in vitro study investigating the interaction of 1354 antidotes with methylmercury bound to erythrocytes demonstrated that unithiol is lost 1355 from the extracellular fluid (Planas-Bohne & Olinger, 1981) and uptake by human 1356 erythrocytes has been demonstrated in vitro (Wildenauer et al., 1982). Uptake of 1357 unithiol by the renal proximal cells is thought to be mediated by the organic anion 1358 transporter (Islinger et al., 2001; see section 7). 1359 1360 9.3 Elimination 1361 1362 Unithiol is rapidly metabolised and subject to renal elimination. 1363 1364 In volunteers given 3 x 100 mg unithiol capsules the elimination half-life in blood of 1365 unithiol and metabolites (altered unithiol) was 4.4 and 9.6 hours, respectively 1366 (Maiorino et al., 1991). Maiorino et al. (1996) determined the half-lives of unithiol and 1367 metabolites to be 1.8 and 20 hours, respectively. The long half-life of altered unithiol 1368 was thought to reflect the stability of the unithiol-albumin complex and since unithiol is 1369 released slowly, albumin may act as a reservoir for unithiol. 1370 1371 In volunteers given unithiol 3 mg/kg intravenously over 5 minutes blood concentrations 1372 declined rapidly with an apparent elimination half-life of 1.8 hours. By 96 hours 12% of 1373 the total unithiol found in the urine was excreted as the parent compound, 1374 representing 10% of the administered dose; 88% was present as disulphide 1375 metabolites (74% of the administered dose). The metabolites are eliminated more 1376 slowly than the parent compound, with an elimination half-life of 23 hours (range 19.8-

28

1377 37.5 hours). The elimination half-life in urine is 20 hours (Hurlbut et al., 1994). 1378 1379 The apparent difference in blood and urine half-lives in the Maiorino et al. (1991) oral 1380 and Hurlbut et al. (1994) intravenous study may be due to different metabolites 1381 produced following administration by different routes. In addition the total unithiol 1382 concentration was determined at different time points (Hurlbut et al., 1994). 1383 1384 9.4 Metabolism 1385 1386 Unithiol is extensively metabolised and unchanged drug is present as only a small 1387 concentration in blood and urine. In volunteers given unithiol 3 mg/kg intravenously 1388 over 5 minutes only 12% of unchanged unithiol was detected in the blood after 15 1389 minutes (Hurlbut et al., 1994). In volunteers given 3 x 100 mg unithiol capsules 3.7% 1390 was excreted as unchanged unithiol and 38.7% as metabolites by 15 hours. Of the 1391 total unithiol found in the urine by 15 hours, unchanged unithiol and metabolites 1392 represented 9% and 91%, respectively. The metabolites are thought to be 1393 disulphide compounds (Maiorino et al., 1991). In a later study (Maiorino et al., 1996) 1394 the unithiol disulphide metabolites were determined to be cyclic polymeric unithiol 1395 disulphides (97%), unithiol-cysteine mixed disulphide (2.5%) and acyclic unithiol 1396 disulphide (0.5%). 1397 1398 9.5 Effect of DMPS on the excretion of metals 1399 1400 Few volunteer studies on the effect of unithiol on metal excretion are available. 1401 1402 9.5.1 Arsenic elimination 1403 1404 In addition to increasing urinary elimination of arsenic, unithiol has also been shown 1405 to alter the relative urinary concentrations of organoarsenic metabolites. 1406 1407 The arsenic-antidote complex was determined in the urine of Romanian subjects 1408 exposed to high concentrations of arsenic in drinking water (up to 16 µg/L). Samples 1409 were collected before and after oral administration of 300 mg of unithiol. Subjects 1410 had been asked to avoid for three days prior to and during the collection 3 1411 period. The presence of a unithiol-monomethylarsenous acid (As ) complex was 1412 demonstrated in the urine samples of subjects given unithiol. Administration of 5 1413 unithiol resulted in a decrease in dimethylarsenic acid (As ) and an increase in 5 3 1414 monomethylarsonic acid (As ) concentrations. Monomethylarsenous acid (As ) is a 5 1415 substrate for the biomethylation of arsenic from monomethylarsonic acid (As ) to 5 1416 dimethylarsenic acid (As ); the formation of the unithiol-monomethylarsenous acid 3 1417 complex reduces the availability of monomethylarsenous acid (As ) for 1418 biomethylation and inhibits further methylation (Gong et al., 2002). 1419 1420 A similar change in urine concentrations of dimethylarsinic acid and 1421 monomethylarsonic acid was also found in the study by Aposhian et al. (1997) 1422 investigating arsenic excretion in two populations in Chile. In one town the drinking 1423 water contained 593 µg/L of arsenic, and in the other 21 µg/L (controls). Subjects 1424 were asked to exclude seafood from their diet for 3 days prior to the study and 1425 bottled water (arsenic <0.5 mg/L) was drunk throughout. Subjects were excluded if 1426 they had a history of previous antidotal therapy, hypersensitivity to similar metal-

29

1427 binding agents or administration of other investigational drugs, serious renal or 1428 psychiatric disease, abnormalities in blood or urine analysis that could 1429 interfere with evaluation, pregnancy, lactation or abuse of alcohol or recreational 1430 drugs. There were 13 subjects in the high arsenic exposure group and 11 controls. 1431 Urine samples were collected before and after oral administration of 300 mg unithiol. 1432 In the 2 hour period after unithiol administration the urinary concentration of the 1433 metabolites monomethylarsonic acid and dimethylarsinic acid represented 42% and 1434 37-38%, respectively, with an inorganic arsenic concentration representing 20-22% 1435 of the total urinary arsenic. The normal range of monomethylarsonic acid is 10-20% 1436 and the percentage increase was almost the same for the two groups. The rise in 1437 monomethylarsonic acid was accompanied by a decrease in dimethylarsinic acid. 1438 The percentage of inorganic arsenic also increased with unithiol treatment. 1439 1440 9.5.2 Bismuth elimination 1441 1442 Two groups of 12 volunteers (age 26-65 years), who had been treated with colloidal 1443 bismuth subcitrate for 28 days because of Helicobacter pylori-associated gastritis, took 1444 part in a study of bismuth elimination with unithiol and succimer. Each subject 1445 received a single oral dose of succimer or unithiol 30 mg/kg in a randomised single 1446 blind study. The succimer or unithiol was given 7 to 14 days after the last dose of 1447 bismuth. Both antidotes produced a 50-fold increase in urinary bismuth excretion 1448 compared to control urine samples. The highest concentration was excreted within 1449 the first 4 hours after dosing. No significant difference was observed in bismuth 1450 elimination between succimer and unithiol and both were well tolerated (Slikkerveer et 1451 al., 1998). 1452 1453 9.5.3 Cadmium elimination 1454 1455 Of the 6 metal-binding agents investigated in an in vitro study using human cell 1456 cultures, unithiol, succimer and mercaptosuccinic acid were found to the most 1457 effective at increasing cadmium movement from cells. Unithiol produced the most 1458 rapid elimination from cells in the first two hours, but the other two agents mobilised 1459 more cadmium than unithiol (Bakka et al., 1981). 1460 1461 9.5.4 Mercury elimination 1462 1463 Significant increases in urinary mercury elimination have been demonstrated with 1464 unithiol administration. 1465 1466 The mercury elimination rate was determined in 5 healthy volunteers (4 male, 1 1467 female, aged 24 to 32 years, 49-93 kg) given unithiol 3 mg/kg intravenously over 5 1468 minutes. Unithiol increased mercury excretion by a factor of 24.2 in the 11 hours 1469 after administration. No relationship was observed between the dose of unithiol and 1470 the quantity of mercury excreted in the urine (Hurlbut et al., 1994). In the 4 male 1471 subjects who had taken part in a previous oral study (Maiorino et al., 1991) the 1472 quantity of mercury excreted in the 12 hours after intravenous administration was 1473 less than that excreted in the same period after oral unithiol (Hurlbut et al., 1994). 1474 1475 Mercury elimination was examined after a unithiol challenge test in 12 male (66-96 1476 kg), former chloralkali workers exposed to metallic mercury vapour for 2-18 years.

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1477 The investigation was undertaken 18-56 months after exposure has ceased. A 1478 single 300 mg dose of unithiol was given and this increased 24 urinary excretion of 1479 mercury by a factor of 7.6. A high proportion (62%) was excreted within the first 6 1480 hours and this probably reflects mercury stored in the kidney (Sallsten et al., 1994). 1481 1482 The clinical efficacy of unithiol was investigated in 10 male volunteers (aged 19-45 1483 years) with occupational mercury exposure (with a urine mercury concentration 1484 equal to or greater than 50 μg/g of creatinine). Each subject received unithiol 100 1485 mg orally three times a day for 5 days. They had been asked to omit seafood from 1486 the diet for one month and to take no iron-containing preparations or 1487 for 2 weeks prior to the study. One subject developed a macular rash 1488 which resolved in two days. Otherwise, all the subjects remained well with no 1489 changes in renal or liver function, blood biochemistry or vital signs. In 9 of the 10 1490 subjects mercury elimination was significantly enhanced during the first 24 hours 1491 after unithiol administration, and in all subjects the mean increase in mercury 1492 excretion was higher over the 5 day period compared to the baseline (Torres- Alanís 1493 et al., 1995). 1494 1495 Gonzalez-Ramirez et al. (1998) also studied the effect of unithiol on mercury 1496 elimination in volunteers with occupational exposure (5 males, 3 females, aged 21- 1497 57 years). The unithiol was given in 3 cycles: 3 days after an initial challenge test, 1498 unithiol was given for 8 days with 5 subsequent days with no treatment. This was 1499 followed by second cycle of 7 days of unithiol, a 5 day period of no treatment and 1500 then 6 days of unithiol. The unithiol was given orally 1 hour before breakfast, lunch 1501 and dinner in doses of 100 mg, 100 mg and 200 mg, respectively, on each treatment 1502 day. One subject developed a maculopapular rash and raised liver enzymes after 1503 the first course and did not receive further doses. Prior to treatment the mean total 1504 urinary mercury excreted in 24 hours was 504 µg (range 140-1692 µg) and during 1505 the first course of unithiol this rose to 1754 µg (range 657-2880 µg). The figure for 1506 the two subsequent courses became progressively lower (314 µg [range 152-658 µg] 1507 and 173 µg [range 74-443 µg]) but in both cases was higher than the period of no 1508 treatment which preceded it (106 µg [range 66-212 µg] and 48 µg [range 30-97 µg]). 1509 Unithiol was effective in lowering the body burden of mercury and increasing the 1510 urinary mercury concentration. 1511 1512 The elimination of mercury following unithiol administration was examined in 75 1513 mercury-exposed volunteers from a gold mining area in the Philippines. Urine 1514 samples were collected before and 2 to 3 hours after unithiol administration (200 mg 1515 orally). In the first urine the mean concentration of inorganic mercury was 15.7 µg/g 1516 of creatinine and for organic mercury it was 2.2 µg/g of creatinine. In the samples 1517 after unithiol dosing the concentrations were 262 µg/g of creatinine and 14.5 µg/g of 1518 creatinine, respectively. Unithiol increased the inorganic urinary concentration by a 1519 factor of 16 and the organic mercury concentration by a factor of 5.1 (Drasch et al., 1520 2007). 1521 1522 Mercury clearance during dialysis was investigated in an in vitro experiment using 1523 pooled plasma samples to which mercury and metal-binding agents had been added. 1524 Of the agents investigated was the most effective in clearing mercury 1525 after 90 minutes of in vitro dialysis, reducing the mercury concentration in the 1526 perfusate by 73%, whereas unithiol removed almost 70% of mercury in the perfusate.

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1527 Unithiol was more effective than succimer but the results were not statistically 1528 different. In contrast the quantity of mercury removed from plasma without the 1529 presence of a metal-binding agent was very low at 5% (Ferguson & Cantilena, 1992). 1530 1531 9.5.5 Palladium elimination 1532 1533 There is limited information on palladium elimination with unithiol. In a study of 50 1534 volunteers the elimination of palladium increased from 0.3 to 38 µg/g of creatinine 1535 and there was no difference between oral and intravenous administration of unithiol 1536 (Runow, 1996). 1537 1538 9.5.6 Trace element elimination 1539 1540 Unithiol has been shown to increase elimination of some trace elements in volunteer 1541 studies. 1542 1543 Trace element elimination was examined after a unithiol challenge test in 12 male 1544 (66-96 kg), former chloralkali workers exposed to metallic mercury vapour for 2-18 1545 years. The investigation was undertaken 18-56 months after exposure has ceased. 1546 A single 300 mg dose of unithiol was given and this increased 24 hour urinary 1547 excretion of copper by a factor of 12 and zinc by 1.5 (Sallsten et al., 1994). 1548 1549 Eleven patients presenting with concerns over exposure to mercury in dental amalgam 1550 were given a unithiol challenge test, and the urinary elimination of trace elements 1551 examined. The patients (8 female, 3 male) had no known occupational exposure to 1552 mercury. The dose of unithiol was 3 mg/kg intravenously and urine samples were 1553 taken 1 hour before and 1 hour after. There was a significant increase in mercury 1554 excretion (3-107 fold) in all subjects. The elimination of chromium and manganese 1555 was unchanged and the urinary concentrations of cobalt, aluminium and molybdenum 1556 were too low for reliable measurement. The urine copper (2-119 fold), selenium (3- 1557 43.8 fold), zinc (1.6-44 fold) and magnesium (1.75-42.7 fold) concentrations were 1558 increased in most patients (Torres- Alanís et al., 2000). 1559 1560 Trace element blood concentrations were examined in 80 volunteers (51 females, 29 1561 males) given 2 mg/kg unithiol intravenously. An increase in urinary copper and zinc 1562 concentrations were observed 30 and 120 minutes after injection. The selenium 1563 concentration was unaffected. There was also a decrease in blood concentrations of 1564 copper, zinc and selenium. However, this may have been due to a dilution effect since 1565 the concentrations returned to normal within 120 minutes of the injection and a similar 1566 effect was observed within iron concentrations (Høl et al., 2003). 1567 1568 1569 10. Clinical studies – clinical trials 1570 1571 Few controlled clinical trials on unithiol are available, and most concern mercury. 1572 1573 10.1 Arsenic and unithiol clinical trials 1574 1575 Unithiol was investigated in a randomised placebo-controlled trial in the management 1576 of chronic arsenicosis due to contaminated drinking water (arsenic >50 μg/L) in

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1577 India. All the patients weighed 40-56 kg and had been drinking the water for more 1578 than 3 years. Subjects were excluded if they had ceased drinking arsenic- 1579 contaminated water for more than 3 months, had been treated with other antidotes, 1580 had a history of smoking, alcoholism, were taking hepatotoxic drugs or were serum 1581 positive for hepatitis B virus surface antigen. Pregnant or lactating women were also 1582 excluded. 1583 1584 A total of 21 patients were randomly assigned to 2 treatment groups: 11 patients (9 1585 males, 2 females, mean age 30.63 years) received unithiol and 10 patients (5 males, 1586 5 females, mean age 34.4 years) received the placebo. The unithiol dosage regimen 1587 was 100 mg 4 times a day for 1 week, repeated in the third, fifth and seventh week. 1588 Further ingestion of arsenic-contaminated drinking water was also stopped. Therapy 1589 with unithiol resulted in significant clinical improvement as evaluated by an objective 1590 clinical scoring system. Cessation of exposure and placebo also reduced clinical 1591 scores but the post-treatment scores were significantly lower for unithiol-treated 1592 subjects. Improvement in clinical scores for the placebo group was attributed to 1593 cessation of exposure, rest and provision of an adequate hospital diet. The most 1594 significant improvement was seen in the clinical score of weakness, pigmentation 1595 and lung disease. There were also significant increases in total urinary arsenic 1596 excretion with unithiol treatment, compared to no increase in the placebo group. 1597 Unithiol was well tolerated with no adverse effects reported (Guha Mazumder et al., 1598 2001). 1599 1600 10.2 Copper (Wilson’s disease) and unithiol clinical trials 1601 1602 Wang et al. (2003) studied 28 patients with Wilson’s disease (18 males, 10 females 1603 aged 14-20 years). Patients were included on the basis of presence of 1604 extrapyramidal symptoms and signs, presence of characteristic corneal Kayser- 1605 Fleischer ring observed with a slit lamp, a serum ceruloplasmin <200 mg/L and 1606 copper oxidase concentration <0.21 units and urinary copper >100 μg (1.56 µmol)/24 1607 hours. Group A received captopril, 1 mg/kg orally daily in 3 divided doses, Group B 1608 unithiol 20 mg/day intravenously and Group C both. Group D was the control group 1609 and did not receive either drug. Serum sulphydryl concentrations and 24 hour 1610 urinary copper concentrations were the markers of anticopper efficacy. Unithiol had 1611 a more potent anticopper effect than captopril and increased urinary copper 1612 concentrations and serum sulphydryl concentrations. Only 1 patient developed an 1613 with transient elevation of alanine aminotransferase. 1614 1615 10.3 Lead and unithiol clinical trials 1616 1617 There are few clinical trials on the use of unithiol in lead poisoning. 1618 1619 One of the earliest reports of unithiol use in the treatment of lead poisoning was by 1620 Anatovskaya (1962). Sixty men with chronic lead toxicity were given 250 mg 1621 intramuscularly for 20 days. Blood concentrations of lead fell and urinary excretion 1622 increased. The clinical signs and symptoms improved subjectively and objectively. 1623 Haematological parameters and liver function improved in more patients in the 1624 unithiol-treatment group compared to controls given only supportive care. In addition 1625 patients treated with unithiol could be discharged from hospital 6 weeks earlier than 1626 controls.

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1627 1628 A more recent study involved children attending a lead poisoning treatment clinic in 1629 Baltimore, USA, where lead paint was the main source of poisoning. The criteria for 1630 inclusion were a blood lead concentration of 400-600 µg/L, normal hepatic, renal and 1631 haematological function, no history of antidotal therapy within the previous 3 months, 1632 no concurrent disease and an age of 30 to 72 months. Six children (5 males, 1 2 1633 female, aged 31 to 53 months) received a 5 day course of unithiol, 200 mg/m daily. 2 1634 Another 6 children (1 male, 5 females, aged 38-69 months), received 400 mg/m daily 1635 for 5 days. The drug was well tolerated in all cases. Administration of either dose 1636 reduced the blood lead concentration to approximately 78% of its pre-treatment 1637 concentration by 48 hours. After 96 hours the blood lead concentration was 76% and 1638 68% of its pre-treatment concentration in the low and high dose group, respectively. 1639 Blood lead concentrations did not increase until 48 hours after cessation of therapy. 1640 Urinary excretion of both copper and zinc were increased, 2-30 fold and 1.2-9.1 fold, 1641 respectively with higher concentrations of both in the higher treatment group. There 1642 were no significant changes in plasma copper or zinc concentrations. The urinary 1643 concentration of lead increased 1.3-15 fold between the last pre-treatment day and the 1644 first treatment day (Chisolm & Thomas, 1985). This trial was later terminated following 1645 the occurrence of at least one case of Stevens-Johnson syndrome and succimer was 1646 used instead (Chisolm, 1990). 1647 1648 10.4 Mercury and unithiol clinical trials 1649 1650 There are a small number of clinical trials investigating the effect of unithiol on 1651 mercury toxicity. An early study undertaken in Iraq, with preliminary results reported 1652 by Bakir et al. (1976) was limited in scope due to various circumstances (Clarkson et 1653 al., 1981). A more recent study examined patients with chronic mercury exposure in 1654 the Philippines (Böse-O’Reilly et al., 2003). Another study in China examined the 1655 effects of antidotal treatment in acutely poisoned patients treated 5 months after 1656 exposure (Zhang, 1984). A Mexican study examined the effect of unithiol on 1657 mercury excretion in patients with high mercury concentrations due to use of 1658 mercury-containing facial cream (Garza-Ocañas et al., 1997). 1659 1660 The early study investigated antidotal therapy in patients with methyl mercury 1661 poisoning. The source was homemade bread made from wheat contaminated with a 1662 methyl mercury fungicide over the winter of 1971-1972. Antidotal agents where in 1663 limited supply and were not available until February 1972 after exposure had ceased. 1664 The conditions of the time did not allow for the implementation of a clinically controlled 1665 study but it was possible to obtain data on the effects of antidotes on blood mercury 1666 concentrations. D-penicillamine (12 patients), N-acetyl-DL-penicillamine (17), unithiol 1667 (10) and a thiolated resin (8) were used. There were 27 females and 20 males, aged 1668 from 18 months to 55 years. Ten other patients received a placebo and 6 received no 1669 specific treatment. The duration of treatment was variable but for unithiol was 4 to 15 1670 days. All agents reduced blood mercury concentrations but unithiol was the most 1671 effective. However, the patients did not show any immediate clinical improvement, 1672 presumably because the duration of therapy was too short (Clarkson et al., 1981). 1673 1674 Unithiol was evaluated in the management of chronic mercury exposure, including 1675 exposure from mercury vapour, inorganic mercury and organic methyl mercury, in the 1676 gold mining area of Mount Diwata, in the Philippines. A total of 95 patients (no details

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1677 given) were included in the study. There was no control group. Unithiol was given 1678 orally at a dose of 200 mg twice daily for 14 days for adults and 5 mg/kg/day for 1679 children. Patients were assessed by questionnaire, medical, including neurological, 1680 examination and neuropsychological testing before and after therapy. Blood, urine 1681 and hair samples were analysed for mercury concentrations before therapy and blood 1682 and urine after therapy. Two study populations were identified: those living near 1683 Mount Diwata and exposed to metallic and/or inorganic mercury (60 patients), and 1684 those living downstream in Monkayo who were exposed to methyl mercury (35 1685 patients). Exposure to mercury continued during therapy with unithiol. The mercury 1686 concentrations in hair and blood did not differ between the two populations before 1687 unithiol therapy. The urine concentration was significantly higher in the Mount 1688 Diwata population (due to the differing pharmacokinetics of organic and inorganic 1689 mercury). There was therefore a higher renal excretion of mercury in the Mount 1690 Diwata population when treated with unithiol. However, the Monkayo population 1691 demonstrated a relative increase in renal excretion of mercury almost as high as the 1692 Mount Diwata population when given unithiol. Some patients (number not specified) 1693 did not respond to unithiol administration and showed no increase in mercury 1694 excretion. In the Monkayo group there was only a modest decrease in blood 1695 mercury concentrations, indicating the duration of treatment was too short to have a 1696 long-term effect on mercury stored in tissues. More than two-thirds of patients 1697 reported an improvement in subjective complaints after therapy and objective 1698 neurological parameters also showed significant improvement. Significant 1699 improvement was also demonstrated in two neuropsychological tests. The authors 1700 concluded that unithiol could increase mercury excretion but that a 14 day regimen 1701 was too short to have a permanent effect on mercury concentrations (Böse-O’Reilly 1702 et al., 2003). This study has a number of limitations including the absence of a control 1703 group, lack of details of patients (age, weight) and continued exposure to mercury 1704 during therapy. 1705 1706 Forty-one patients (aged 2 to 65 years) from 8 families were poisoned with mercury 1707 following ingestion of rice contaminated with an ethyl mercury seed dressing. One 1708 patient died soon after onset of symptoms and 8 were admitted in the initial acute 1709 phase of intoxication. The remaining 40 patients demonstrated a variety of clinical 1710 features 5 months after ingestion and 27 were treated with unithiol (250 mg daily by 1711 intramuscular injection) and/or succimer (500 mg twice daily by intravenous injection). 1712 The drugs were given for 3 days followed by a 4 day break and then another 3 day 1713 course, if required. Patients were given 1 to 8 courses until the urinary mercury 1714 concentration was normal. The 13 untreated patients showed little improvement in 1715 clinical features of toxicity but all those on therapy had some relief and 19 became 1716 asymptomatic. In 2 cases there was only slight improvement. In patients where the 1717 urinary concentration was measured before therapy, all but one had increased 1718 mercury excretion during antidotal administration. Side effects were mild and 1719 generally resolved within 30 minutes to 4 hours. Unithiol was found to be more 1720 effective, although no data distinguishing between the two drugs is given. In addition, 1721 the two antidotes were used interchangeably in some patients (Zhang, 1984). 1722 1723 Unithiol was also used in 12 females (aged 19-45 years) with mercury toxicity 1724 following use of a facial cream containing 5.9% mercurous chloride (calomel) for 2 to 1725 10 years. Two patients were symptomatic (exanthema and tremor) and all had 1726 elevated urinary mercury concentrations. Subjects were given a 5 day course of

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1727 unithiol (200 mg/day) as outpatients. Only 8 subjects took the unithiol as prescribed 1728 and supplied 24 hours urine samples for analysis. In all cases there was a significant 1729 increase in urinary mercury concentrations after 24 hours of unithiol. One 1730 symptomatic patient has complete resolution of effects and the other had persistent 1731 tremor. Unithiol was well tolerated in all patients (Garza-Ocañas et al., 1997). 1732 1733 1734 11. Case reports - clinical studies 1735 1736 In many clinical case reports of unithiol use in poisoning the authors do not determine 1737 a balance between the quantity of metal absorbed and excreted; this can be difficult 1738 where the dose ingested, injected or inhaled cannot be quantified. Clinical efficacy of 1739 unithiol is often only stated in terms of enhancing excretion and/or decreasing blood 1740 concentrations in the absence of severe adverse effects. In many cases, the authors 1741 are unable to distinguish between the effect of unithiol administration and the effect of 1742 supportive therapy (including removal from the source). 1743 1744 The efficacy of a metal-binding agent may be difficult to determine. After 1745 discontinuation of metal exposure (and absorption) a decrease in the blood 1746 concentration will occur without any therapy. Clinical efficacy should not be judged 1747 only by the amount of metal excretion or the decrease of blood concentrations. The 1748 reduction of the tissue content in the target organ and the restoration of pathological 1749 alterations also need to be considered. It is important to note that enhancement of the 1750 metal excretion by mobilisation may increase the metal burden of the target organ by 1751 redistribution, and conversely the body burden may be reduced without a striking 1752 decrease of the blood concentrations. However, in some case reports severe toxicity 1753 usually associated with a demonstrated high blood or urine concentration does not 1754 occur, and this may reasonably be assumed to be due to antidotal therapy. With 1755 these reservations in mind, it is clear that administration of unithiol can prevent 1756 development of toxicity and in symptomatic patients it can reduce recovery time and 1757 improve clinical signs and symptoms of toxicity. 1758 1759 Reference values for the metal and metalloids (Walker, 1998) are given as a guide at 1760 the start of each section to aid interpretation of the concentrations given in the case 1761 reports. 1762 1763 11.1 Use in antimony poisoning 1764 1765 Reference values for antimony (Walker, 1998): 1766 Children Serum/plasma 0.18 µg/L 1767 Blood 0.26 µg/L 1768 Urine 0.05 µg/L 1769 Adults (unexposed) Urine 0.8 µg/L 1770 1771 There is limited information on the use of unithiol in antimony poisoning, but it appears 1772 to be of benefit in the management of poisoning with trivalent antimony compounds. 1773 There is no information on its use in the management of poisoning with the less toxic 1774 pentavalent antimony compounds. 1775 1776 A 2 year 11 month old child was treated with unithiol after ingestion of an unknown

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1777 quantity of tartar emetic (antimony potassium tartrate). The clinical features, 1778 particularly massive fluid loss and dehydration, were consistent with antimony 1779 poisoning. She was given unithiol 65 mg intravenously then 3 x 100 mg daily for 10 1780 days followed by 3 x 50 mg daily for another 10 days. She was also treated with 1781 exchange transfusion at 39 hours post-ingestion. The serum antimony concentration 1782 at 4.5 hours post-ingestion was 0.6 mg/L and the urine concentration at 7 hours 31 1783 mg/L. Unithiol was considered effective in reducing the antimony concentration in the 1784 early stages of poisoning (Iffland & Bösche, 1987). 1785 1786 Unithiol has also been used with apparent success in other paediatric cases of 1787 antimony potassium tartrate poisoning (Kemper et al., 1989; Jekat & Kemper, 1990) 1788 and is recommended by others for the management of antimony poisoning (Chzhi- 1789 Tysan, 1959; Lauwers et al., 1990). 1790 1791 11.2 Use in arsenic poisoning 1792 1793 Reference values for arsenic (Walker, 1998): 1794 Blood <0.13 µmol/L (<10 µg/L) 1795 Urine <0.13 µmol/24 hours (<10 µg/24 hours) of inorganic arsenic 1796 Urine <40 nmol/mmol creatinine (unexposed) 1797 <173 nmol/mmol creatinine (occupational exposure) 1798 1799 Unithiol has been used successfully in a number of cases of acute and chronic 1800 arsenic poisoning and is considered the antidote of choice for arsenic toxicity (Adam 1801 et al., 2003). 1802 1803 A 33-year-old female (62 kg) was treated with unithiol after accidental ingestion of 1804 arsenic paste (approximately 1.85 g of arsenic trioxide). She presented 100 hours 1805 after ingestion and was started on unithiol (250 mg intravenously every 6 hours for 1806 48 hours, then 250 mg orally 3 times a day for 48 hours, followed by 250 mg every 1807 12 hours for 23 days). Her condition improved after 2 days and she made a full 1808 recovery. The highest urinary arsenic concentrations occurred on hospital days 1-3 1809 (Kruszewska et al., 1996). 1810 1811 Two brothers, aged 21 and 19 years, were treated with unithiol after ingestion of 4 g 1812 and 1 g, respectively, of a powder which was later identified as arsenic trioxide. 1813 Treatment with unithiol was started 32 and 48 hours after ingestion. In the older 1814 brother the blood arsenic concentration at 26 hours was 400 µg/L. He suffered a 1815 cardiac arrest just before unithiol was started but was successfully resuscitated. At 1816 36 hours the blood arsenic concentration of the younger brother was 98 µg/L. Both 1817 men made a full recovery with no clinical or electrophysiological signs of arsenic 1818 neuropathy (Moore et al., 1994). 1819 1820 A 21-year-old male was treated with unithiol after intentional ingestion of 0.6 g of 1821 arsenic trioxide. He presented to hospital seven hours after ingestion and was 1822 dehydrated following diarrhoea, intestinal colic and . He was given a gastric 1823 lavage, rehydrated (on average 5.5 L of fluid/daily over 5 days) and given high dose 1824 unithiol. The parenteral regimen was 250 mg/hour on day 1, 125 mg/hour on day 2 1825 and then 62.5 mg/hour on days 3-5. From days 6-12 he was given oral doses of 600- 1826 700 mg/day. The total dose given was 15.225 g. The serum arsenic concentration

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1827 on day 1 was 143 mg/L (urine 210,000 µg/L), on day 2, 29 mg/L (3800 µg/L), on day 1828 3, 10 mg/L (1675 µg/L) and by day 8 was <1 mg/L (115 µg/L). He developed mild 1829 increases in transaminase concentrations but remained otherwise well (Horn et al., 1830 2002). Heinrich-Ramm et al. (2003) examined the arsenic compounds excreted in this 1831 patient. In urine sampled on days 2-8 he excreted about 50 mg of arsenic of an 1832 estimated total ingested dose of 230 mg of arsenic. In the first urine sampled after 1833 unithiol therapy (11 hours post-ingestion) the arsenic concentration was 215 mg/L. 1834 This fell by a 1000-fold to 169 µg/L after 8 days of unithiol administration. As reported 1835 in other studies (see section 9.5.1), administration of unithiol resulted in a decrease 1836 in the urinary concentration of dimethylarsinic acid. In this patient the urinary 1837 concentration of dimethylarsinic acid did not exceed that observed in a reference 1838 population where it had accounted for 95% of the total arsenic excreted. In contrast, 1839 the urinary dimethylarsinic acid concentration in this patient accounted for <5% of 1840 the total arsenic excreted. This supports the theory that unithiol interferes with 1841 methylation of arsenic. 1842 1843 A 27-year-old female developed gastrointestinal effects, ECG changes and liver 1844 damage after ingestion of 9 g of arsenic trioxide. She was treated with intravenous 1845 fluids, activated charcoal and continous alkaline irrigation of the stomach over 36 1846 hours. She was given succimer (15 mg/kg every 8 hours for 26 days) and 1847 intramuscular dimercaprol (4 mg/kg every 4 hours) for 24 hours. A second course of 1848 dimercaprol was administered on day 5 due to continued deterioration. She was 1849 also started on a regimen designed to enhance methylation of arsenic derivatives 1850 which are less toxic and more readily excreted. She was given , 1851 , , sodium bicarbonate, glutathione and intravenous unithiol 1852 (250 mg every 4 hours for 5 days). She began to improve within 48 hours with 1853 improved pulmonary function and ECG. Liver function tests began to resolve but 1854 were elevated for more than a month. She also received unithiol from days 15 to 18. 1855 Urinary arsenic concentrations fell rapidly in the first 10 days. At follow up one year 1856 later she had mild polyneuropathy. The role of succimer in this patient’s recovery 1857 was not evaluated. Unithiol, with and without the methylating regimen, increased the 1858 proportion of methylated arsenic metabolites (monomethylarsonic acid and 1859 dimethylarsenic acid) in the urine (Vantroyen et al., 2004). 1860 1861 A 33-year-old female with chronic arsenic poisoning from an unknown source was 1862 treated with unithiol. She presented with a 1.5 year history of episodes of peripheral 1863 neuropathy, pancytopenia, ventricular tachycardia, gastrointestinal symptoms, skin 1864 rash and nail changes. Her blood arsenic concentration was 56 µg/L and the 24 1865 hour urine concentration 130 µg/L. Analysis of well water demonstrated an arsenic 1866 concentration of 78 µg/L. Electromyography revealed moderate demyelinating 1867 neuropathy with axonal involvement. She was started on succimer 10 mg 3 times a 1868 day but serial urinary arsenic determination did not show any elimination. Her 1869 neuropathy continued to progress and she required ventilation. She was then 1870 started on unithiol at 250 mg/kg intravenously every 4 hours. During the first 24 1871 hours of treatment the urinary arsenic concentration rose from 101 to 300 µg/L. 1872 There was also improvement in her neuropathy and she was continued on unithiol 1873 for 12 days. She was much improved, extubated and discharged in a wheelchair 10 1874 days later. By 3 months she was walking on her own with some residual 1875 paraesthesiae and weakness in distal lower extremities. At follow up one year later 1876 she had only mild weakness and residual paraesthesiae controlled with amitriptyline

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1877 (Wax & Thornton, 2000). 1878 1879 Adam et al. (2003) reports 6 cases of arsenic poisoning, with retrospective 1880 comparison of the use of dimercaprol and unithiol. Three patients were treated with 1881 dimercaprol. Two of these patients with blood arsenic concentrations of 540 µg/L 1882 and 620 µg/L died within 34 hours and 6 days of exposure, respectively. One patient 1883 had a maximal blood arsenic concentration of 558 µg/L and developed paraplegia as 1884 a result of arsenic poisoning. The three patients treated with unithiol recovered fully. 1885 Two of these patients (including one with anuric renal failure) had very high blood 1886 arsenic concentrations of 2240 µg/L and 4469 µg/L. The third patient had a 1887 relatively mild course, after the early use of unithiol, despite a blood arsenic 1888 concentration of 245 µg/L. The authors concluded that unithiol is the treatment of 1889 choice for arsenic poisoning and that dimercaprol is obsolete. 1890 1891 11.3 Use in beryllium poisoning 1892 1893 No well documented case reports of unithiol use in beryllium poisoning could be 1894 found. 1895 1896 11.4 Use in bismuth poisoning 1897 1898 Reference values for bismuth (Walker, 1998): 1899 Blood <0.5 nmol/L (<0.1 µg/L) basal 1900 Up to 240 nmol/L (up to 50 µg/L) acceptable therapeutic 1901 >480 nmol/L (>100 µg/L) risk of toxicity 1902 Urine <0.5 nmol/L (<0.1 µg/L) 1903 1904 Unithiol has been used in both acute (Stevens et al., 1995; Bogle et al., 2000; 1905 Dargan et al., 2001; Dargan et al., 2003b; Ovaska et al., 2008) and chronic (Playford 1906 et al., 1990) bismuth poisoning, although clinical deterioration has been reported in 1907 one chronic case due to redistribution of tissue stores of bismuth (Teepker et al., 1908 2002). Unithiol is considered an effective antidote for acute bismuth poisoning 1909 (Andersen, 1999). 1910 1911 Unithiol was started 24 hours after intentional ingestion of 2.88 g of bismuth 1912 subcitrate in a 13-year-old girl. She was given 30 mg/kg orally for 10 days and 10 1913 mg/kg for 9 days. The serum bismuth concentration at 4 hours was 300 µg/L, 14 1914 μg/L at 48 hours and 8 µg/L at 72 hours. At 10 days post-ingestion the serum 1915 bismuth concentration was 1.8 µg/L. She remained well throughout (Bogle et al., 1916 2000). 1917 1918 In a similar case a 30-year-old male was started on unithiol after ingestion of 4.8 g of 1919 tripotassium dicitratobismuthate. He was admitted 2 hours post-ingestion with 1920 vomiting, but was otherwise well. The initial bismuth blood concentration was 424 1921 µg/L and urine 10,000 µg/L. Oral unithiol was started on day 2 (200 mg four times 1922 daily for 10 days, then 200 mg twice daily for 10 days). He remained well and after 1923 antidotal therapy the blood concentration was 17 µg/L and urine 37 µg/L (Dargan et 1924 al., 2001). 1925 1926 A 21-year-old male developed bismuth toxicity after ingestion of 50 to 60 tablets of

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1927 tripotassium dicitratobismuthate. The blood bismuth concentration was 590 µg/L 1928 and he was initially given dimercaprol 150 mg intramuscularly. Four hours later he 1929 was started on haemodialysis. He was subsequently given unithiol 250 mg 1930 intravenously 4 hourly for 48 hours and then 250 mg 6 hourly for 48 hours, then 500 1931 mg orally in two divided doses for 14 days (days 6 to 19 after ingestion). He 1932 received haemodialysis for the first 6 days and on days 8, 9 and 12. Each session 1933 lasted 4 hours and started 1 hour after unithiol administration. No significant 1934 clearance of bismuth occurred with dimercaprol. In contrast significant bismuth 1935 clearance was obtained with unithiol and despite improving renal function cessation 1936 of unithiol on day 20 resulted in a decrease in urinary bismuth clearance (Stevens et 1937 al., 1995). 1938 1939 Unithiol has been used in poisoning resulting from the use of bismuth iodoform 1940 paraffin paste used to pack a large wound of the sacrum following tumour excision. 1941 The patient began to develop neurological effects 5 days later and blood and urine 1942 bismuth concentrations were raised (340 µg/L and 2800 µg/L, respectively). The 1943 packing was removed and he was started on unithiol (intravenously 5 mg/kg four 1944 times daily for 5 days, 5 mg/kg 3 times daily for 5 days, and 5 mg/kg twice daily for 1945 17 days, then orally 200 mg 3 times daily for 10 days, 200 mg twice daily for 14 1946 days; 61 days in total). His neurological effects improved over the next month and 1947 the bismuth concentrations were normal after 55 days (Dargan et al., 2003b; Ovaska 1948 et al., 2008). 1949 1950 A 68-year-old male with renal impairment developed encephalopathy from ingestion 1951 of double the dose of tripotassium dicitratobismuthate (864 mg of bismuth daily) for 2 1952 years. He was given unithiol 100 mg 3 times daily for 10 days. Renal clearance of 1953 bismuth increased 10-fold from 0.24 mL/minute to 2.4 mL/minute during this time. 1954 The whole blood bismuth concentration decreased from 880 µg/L to 46 µg/L over a 1955 50 day period and he had marked improvement in cerebral function. There were no 1956 adverse effects (Playford et al., 1990). 1957 1958 In a 49-year-old female with encephalopathy from 5 years of chronic oral abuse of 1959 bismuth, unithiol had to be discontinued due to deterioration in her clinical condition. 1960 She was given 100 mg daily and there was a marked decrease in plasma bismuth 1961 concentrations with an increase in urinary bismuth concentrations. However, she 1962 deteriorated with cluster-like myoclonic jerks and stupor and the unithiol was stopped 1963 after 3 days. Her clinical condition improved over the next few weeks and lagged 1964 behind the plasma bismuth concentrations. They deceased from 550 µg/L to 30.4 1965 µg/L by day 21 when she demonstrated marked improvement. The unithiol may 1966 have caused redistribution of bismuth from tissue stores and caused the clinical 1967 deterioration in this patient (Teepker et al., 2002). 1968 1969 11.5 Use in cadmium poisoning 1970 1971 Reference values for cadmium (Walker, 1998): 1972 Blood <27 nmol/L (<3 µg/L) non smokers 1973 <54 nmol/L (<6 µg/L) smokers 1974 Urine 44.5 nmol/L (5 g/L) 1975 0.4-1.3 nmol/mmol creatinine (unexposed) 1976

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1977 There is limited information on the use of unithiol in cadmium poisoning. In a woman 1978 with chronic occupational exposure to cadmium and signs and symptoms of toxicity, 1979 administration of unithiol increased the urinary concentration of cadmium from 1.8 to 1980 4.2 μg/L (Daunderer, 1995). 1981 1982 11.6 Use in chromium poisoning 1983 1984 Reference values for chromium (Walker, 1998): 1985 Blood <10 nmol/L (<0.5 µg/L) 1986 Urine <5 nmol/L (<0.25 µg/L) non-occupational exposure 1987 Up to 98 nmol/mmol creatinine, depending on type of occupation 1988 1989 There is limited information on the use of unithiol in chromium poisoning. An adult 1990 who fell in a pool of chromic acid was started on unithiol within an hour of exposure. 1991 He became anuric and chromium was detected in the dialysate. Urine excretion 1992 returned 2 days after exposure and the chromium concentration was 5859 μg/L. The 1993 maximum urinary chromium concentration, 13,614 μg/L, was obtained 12 hours after 1994 the start of unithiol therapy. The serum chromium concentration was 1,983 mg/L 24 1995 hours after exposure. The patient recovered but no further details are given (Donner 1996 et al., 1986). 1997 1998 A 20-year-old male died after ingestion of 10-30 g of potassium dichromate. He was 1999 treated with haemodialysis (commenced at 2.5 hours) and continuous arterio-venous 2000 haemofiltration (at 16 hours). Unithiol (250 mg intravenously every 4 hours for 24 2001 hours then 6 hourly) was started 13 hours after admission. The initial plasma 2002 concentration of chromium was 5.8 mg/L and urine 159 mg/L. He had a cardiac 2003 arrest and died 48 hours after admission (Pudill et al., 1989). 2004 2005 11.7 Use in cobalt poisoning 2006 2007 There is limited information on the use of unithiol in cobalt poisoning. Unithiol was 2008 used in 2 paediatric patients with ingestion of an unspecified cobalt compound from 2009 a chemistry set. The children were initially treated with penicillamine and then on the 2010 fifth day started on the less toxic unithiol (3 x 50 mg orally). There were slight 2011 increases in serum cobalt concentrations during unithiol therapy and it was stopped 2012 once the urine cobalt concentrations were normal. No further details are given 2013 (Müller et al., 1989). 2014 2015 11.8 Use in copper poisoning 2016 2017 There is limited information on the use of unithiol in acute copper poisoning. A 3- 2018 year-old child who ingested more than 3 g of copper sulphate received a gastric 2019 lavage within 30 minutes of ingestion and was commenced on unithiol therapy within 2020 an hour. The serum copper concentration did not reach a toxic concentration and 2021 urinary copper excretion was more than 10 times normal; no further details are given 2022 (Donner et al., 1986). 2023 2024 A 33-year-old female ingested an unknown quantity of copper sulphate and developed 2025 severe haemorrhagic gastroenteritis, dehydration, metabolic acidosis, renal failure, liver 2026 damage, intravascular haemolysis and methaemoglobinaemia. She was given unithiol

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2027 (250 mg intravenously every 4 hours) within 24 hours of ingestion until she developed 2028 anuria (more than 48 hours later). This patient survived with 10 days of intensive 2029 therapy and 5 weeks of hospitalisation. The serum copper concentration was normal 2030 on admission (5 hours post-ingestion) and as there were no measurements of copper 2031 excretion, the effectiveness of unithiol administration cannot be determined (Sinković et 2032 al., 2008). 2033 2034 11.8.1 Use in Wilson’s disease 2035 2036 Unithiol has been used with some success in patients with Wilson’s disease, usually 2037 those who fail to respond to penicillamine. 2038 2039 Walshe (1985) reported the use of unithiol (200 mg twice daily) in an adult patient with 2040 Wilson’s disease. He had initially been managed with penicillamine and trientine but 2041 had become intolerant to them. He developed proteinuria but remained well with 2042 unithiol. His copper excretion was maintained at 31.5-47.2 µmol (2000-3000 µg) daily. 2043 Two other patients were also tried on unithiol. One developed fever and a 2044 decreased leucocyte count which also occurred with a second test dose and further 2045 treatment was not given. The other patient took it for 10 days but then refused 2046 because of intense nausea. Other patients (number not specified) were given test 2047 doses and the resulting cupruresis was comparable with that obtained with 2048 penicillamine and trientine in most cases. 2049 2050 11.9 Use in gold poisoning 2051 2052 Unithiol has been used in iatrogenic gold poisoning and although the patient died 2053 from heart failure the unithiol was thought to be effective in removing gold from the 2054 body. No further details are given (Ashton et al., 1992a). 2055 2056 11.10 Use in lead poisoning 2057 2058 Reference values for lead (Walker, 1998): 2059 Blood <0.5 µmol/L (<10 µg/L) environmental exposure 2060 Urine <100 nmol/24 hours (<10 µg/24 hours) normal adults 2061 2062 Unithiol was first used in the management of chronic lead poisoning in Russia 2063 (Anatovskaya, 1962), and although unithiol has been used in subsequent cases of 2064 lead poisoning and has been shown to be of benefit in animal studies, succimer is 2065 usually the preferred antidote for lead poisoning, particularly in children (Angle, 2066 1993) as it is the less toxic of the two antidotes (Andersen, 1999). 2067 2068 An adult with lead poisoning due to the use of a lead-containing ointment was treated 2069 with oral unithiol (400 mg then 200 mg 2 hourly). She had a blood lead concentration 2070 of 1150 µg/L with gastrointestinal effects and a paraesthesia. Within 36 hours the 2071 concentration has decreased to 570 µg/L and the maximum urinary lead concentration 2072 in 24 hours was 73000 µg/L. The blood concentration rose to 890 µg/L during an 2073 interruption of therapy due to lack of drug supply, but decreased rapidly once unithiol 2074 was recommenced. Therapy was continued over 2 months without adverse effects 2075 (Donner et al., 1987; Hruby & Donner, 1987). 2076

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2077 A 24-year-old female developed lead poisoning after ingestion of instant lemon tea 2078 from a lead-glazed cup, over a period of 2.5 months. The reason for her illness was 2079 difficult to establish at first due to the unusual source, but once lead toxicity was 2080 confirmed she was treated with oral unithiol (5-10 mg/kg 3 times a day for 2 days, then 2081 2.5 mg/kg twice daily) until the blood and urine lead concentrations were normal. Her 2082 initial whole blood and urine lead concentrations were 600 µg/L and 1700 µg/L, 2083 respectively. She recovered over a 4 month period (Autenrieth et al., 1998). 2084 2085 Unithiol treatment (parenteral and then two 5 day cycles) improved optic neuropathy 2086 caused by lead deposition in the eye. There was improvement in visual acuity, visual 2087 field and dark adaptation (Dambite, 1966). 2088 2089 11.11 Use in mercury poisoning 2090 2091 Unithiol is the antidote of choice in patients with mercury poisoning. It has been used 2092 successfully in the treatment of acute and chronic poisoning, involving metallic 2093 mercury, inorganic mercury salts and organic compounds. Andersen (1999) suggests 2094 that unithiol is the optimal antidote for inorganic mercury poisoning and that succimer 2095 is more effective in organic mercury intoxication. 2096 2097 In acute poisoning with mercuric salts the initial doses usually have to be given by the 2098 parenteral route because of damage to the gastrointestinal tract. Unithiol should be 2099 used in combination with haemofiltration in patients with mercury-induced renal failure. 2100 2101 Reference values for mercury (Walker, 1998): 2102 Blood <20 nmol/L (<4 µg/L) 2103 Urine <50 nmol/24 hours (<10 µg/24 hours) 2104 2105 11.11.1 Inorganic mercury compounds 2106 2107 Campbell et al. (1986) reported two patients with high mercury concentrations, due to 2108 occupational exposure, treated with unithiol. One patient aged 22 years was 2109 asymptomatic despite a urinary mercury concentration of 832 µg/24 hours. The other, 2110 aged 23 years, had clinical features of mercury toxicity (weight loss, muscle twitching, 2111 excessive salivation, night sweats). His urinary mercury concentration was 429 µg/24 2112 hours. Both were given unithiol, 100 mg 3 times a day then 400 mg daily, for 2 2113 months. The drug was well tolerated and during therapy the elimination half-life of 2114 mercury deceased from 33 days to 11 days. After 2 months the symptomatic patient 2115 had improved and the urinary mercury concentration was within acceptable limits. 2116 2117 Ashton & House (1989) describe two patients treated with unithiol after ingestion of 2118 inorganic mercury. The first patient, aged 19 years, ingested approximately 29 g of 2119 mercuric nitrate and presented to hospital within 1.5 hours. He was treated with 2120 dimercaprol initially and he developed acute renal tubular necrosis and hypotension. 2121 He was then given high dose intravenous unithiol, haemodialysis, haemofiltration and 2122 plasma exchange. The initial blood mercury concentration was very high but renal 2123 function returned at 10 days post-ingestion and he made a full recovery. 2124 Haemodialysis, haemofiltration and plasma exchange were ineffective as promoting 2125 mercury removal. The second patient, aged 42, ingested approximately 1 g of 2126 mercuric chloride. He was treated with high dose intravenous unithiol and then oral

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2127 administration. He recovered without developing renal impairment, despite an initial 2128 mercury blood concentration of 600 µg/L. 2129 2130 Prompt administration of unithiol (at 8 hours post-ingestion) with intravenous fluid 2131 therapy was thought to be responsible for the lack of renal impairment in a 53-year-old 2132 man who had ingested approximately 50 g of mercuric iodide. He had vomited 2133 repeatedly and this may also have decreased absorption, however, the blood and 2134 urine mercury concentrations were high, at 1197 nmol/L and 159 nmol/L, respectively. 2135 He was given unithiol intravenously 250 mg every 4 hours for 60 hours and then orally 2136 twice daily for 18 days (Anderson et al., 1996). 2137 2138 A 19-year-old female vomited 30 minutes after ingestion of 3g of mercuric chloride and 2139 was given a gastric lavage. She developed anuria one hour later and was started on 2140 peritoneal dialysis and haemodialysis. She was given unithiol and dimercaprol and 2141 urine excretion returned after 10 days with a polyuric phase at 20 days. Creatinine 2142 clearance was normal by 100 days after ingestion (Nadig et al., 1985). 2143 2144 A 38-year-old male intentionally ingested 100 ml of mercuric chloride solution (of 2145 unknown concentration) and developed vomiting, haematemesis and bloody diarrhoea 2146 shortly afterwards. He was given a gastric lavage and once the history of ingestion 2147 had been determined was stated on dimercaprol. However, he rapidly developed 2148 oliguria and acute tubular necrosis. By 8 hours post-ingestion the urine output was 2149 less than 10 mL/hour. The blood mercury concentration was 14,300 µg/L. The urine 2150 mercury concentration before onset of anuria was 36,000 µg/L. He was started on 2151 intravenous unithiol 10 hours after ingestion: 250 mg every 4 hours for 48 hours, then 2152 250 mg every 6 hours for 48 hours and 250 mg 8 hourly. He required intravenous 2153 fluids for hypovolaemic shock and haemodialysis for renal failure. The blood mercury 2154 concentration remained high (at least 2 mg/L for the first 10 days) but kidney function 2155 returned within 10 days and haemodialysis was no longer required. Administration of 2156 unithiol was continued by the parenteral route because of ulceration of the 2157 oesophagus and stomach. After about 4 weeks oral unithiol was given (300 mg 3 2158 times a day), and unithiol was given for a total of 7 weeks until blood and urine 2159 concentrations were considered to be non-toxic. The average mercury half-life is 40- 2160 60 days and in this patient it was 2.5 days in the initial distribution phase and 8.1 days 2161 in the terminal phase of metabolism and elimination. The patient made a full recovery 2162 (Toet et al., 1994). 2163 2164 Haemodialysis is ineffective in enhancing mercury elimination (Toet et al., 1994; Pai 2165 et al., 2000) even in patients treated with unithiol (Toet et al., 1994). However, 2166 continuous venovenous haemofiltration was successful in enhancing elimination of 2167 the mercury-unithiol complex in a patient with elevated blood mercury concentrations 2168 (initially 5200 µg/L) following ingestion of an inorganic mercury salt (Pai et al., 2000). 2169 2170 Continuous venovenous haemofiltration in combination with unithiol was also 2171 effective in the patient reported by Dargan et al. (2003a). A 40-year-old male 2172 ingested approximately 1 g of mercuric sulphate and soon after presentation required 2173 intubation and ventilation due to respiratory distress (his pharynx and epiglottis were 2174 oedematous and haemorrhagic). He was started on intravenous unithiol (250 mg 2175 every 4 hours for 4 days) 4.5 hours after ingestion. However he developed an 2176 erythematous maculopapular rash with blistering on the lower legs and the dose of

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2177 unithiol was reduced to 250 mg every 8 hours. After another 6 days he was started 2178 on oral unithiol (200 mg/day for 9 days). Anuria developed by 12 hours post-ingestion 2179 and he was commenced in continuous venovenous haemofiltration 7 hours after 2180 ingestion. He was anuric for 11 days with oliguria for days 12 to 43. Continuous 2181 venovenous haemofiltration was continued for 14 days with 8 sessions of 2182 haemodialysis for renal support between days 16 and 37. He was discharged on 2183 day 50 asymptomatic with no neurological signs or symptoms. Continuous 2184 venovenous haemofiltration removed 12.7% of the ingested dose, mostly over the 2185 first 72 hours. 2186 2187 11.10.2 Organic mercury compounds 2188 2189 A 20-year-old male was given a gastric lavage and activated charcoal within 2 hours of 2190 ingesting haloperidol, benztropine and 2-3 mouthfuls of a fungicide containing 0.69% 2191 methyl mercury and ethanol. The estimated dose of methyl mercury was 800 mg. He 2192 was started on oral D-penicillamine within 4 to 5 hours of ingestion. The whole blood 2193 mercury concentration 2 hours after ingestion was 1930 μg/L and at 24 hours was 2194 1007 μg/L. Approximately 36 hours after ingestion he was started on acetylcysteine 2195 and haemodialysis. D-penicillamine was discontinued 3 days after ingestion and oral 2196 unithiol (200 mg every 6 hours) was started. He received unithiol for 14 days but due 2197 to mild anorexia and nausea elected not to continue taking it as an outpatient. The 2198 whole blood mercury concentration at this time was 355 μg/L and he remained well. 2199 Serum zinc and copper concentrations remained normal during unithiol therapy. 2200 Neurological examination was normal at 6 weeks and 1 year post-ingestion. 2201 Haemodialysis and D-penicillamine were relatively ineffective in clearing the mercury. 2202 The unithiol was also relatively ineffective but this may have been due to 2203 administration of a and mineral preparation containing copper and zinc at 2204 the same time, which may have reduced efficacy (Lund et al., 1984). 2205 2206 In a 49-year-old female who ingested 125 g of fungicide containing 3.5% mercury in 2207 the form of methoxyethyl mercury treated with penicillamine and unithiol alternating 2208 every 2 weeks, unithiol reduced the protein binding of methoxyethyl mercury from 93% 2209 to 83%. Antidote therapy was continued for 12 weeks and she developed no renal or 2210 neurological effects (Köppel et al., 1982). 2211 2212 A 44-year-old man was treated with unithiol and succimer after ingestion of a solution 2213 of thiomersal. The ingested dose was 83 mg/kg although he vomited about 15 2214 minutes later. He was given a gastric lavage just over 1 hour after ingestion and 300 2215 mg of unithiol was instilled into the stomach via a nasogastric tube. This dose was 2216 repeated on days 2, 3, 9 and 10, and he was given 250 mg of intravenous unithiol on 2217 days 3, 8 and 17, with 750 mg on days 4, 5 and 11, and 1000 mg on days 12-16 and 2218 23-29. He was also given oral succimer on days 17-23, 33-46 and 51-70. He 2219 received no antidotal therapy on days 6, 30-32 and 47-50. He developed renal failure 2220 on day 1 which persisted until day 40. He also developed gastritis, dermatitis, 2221 gingivitis, polyneuropathy and coma. He made a full recovery but the decline in 2222 mercury concentrations in the blood, urinary mercury excretion and renal mercury 2223 clearance were not influenced by antidotal therapy to a great extent. There was 2224 minimal or no increase in renal and blood clearance and no effect was detected after 2225 day 30 (Pfab et al., 1996). 2226

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2227 11.11.3 Metallic mercury 2228 2229 In a patient with intrabronchial aspiration of metallic mercury the urine and plasma 2230 mercury concentrations remained high despite intermittent therapy with unithiol and 2231 penicillamine over a 14 month follow up period. However, the patient remained well 2232 apart from vomiting and faintness on admission and intermittent mild arthralgia 2233 thereafter (Batora et al., 2001). In a similar case, a 35-year-old male remained 2234 asymptomatic after rupture of a Miller-Abbot tube and subsequent aspiration. The 24 2235 hour blood mercury concentration was 940 µg/L. He was treated with unithiol and the 2236 blood concentration decreased rapidly (Kummer & Michot, 1984). 2237 2238 There are a number of case reports of unithiol use in children with mercury toxicity 2239 (Bertram et al., 1989; Jekat & Kemper, 1990; Ruprecht, 1997). Unithiol was used in 2240 three children, aged 33 and 20 months and 6 years 10 months, with mercury toxicity 2241 from a broken thermometer. The thermometer had been broken on the carpet of the 2242 children’s room, which had under floor heating, about 8 months previously. Unithiol 2243 (50 mg 3 times daily) was given for up to 4 months and all the children recovered (von 2244 Mühlendahl, 1990). 2245 2246 Unithiol was used in the management of mercury poisoning in a 14-year-old girl with 2247 . For 3 months her clinical features were diagnosed as a neurotic anxiety 2248 disorder but once the diagnosis of mercury toxicity was made she was treated with 2249 unithiol, 100 mg every other day and made a slow recovery. The source was metallic 2250 mercury spilt on a carpet that had been vacuumed up (Böckers et al., 1983). 2251 2252 11.11.4 Dermal mercury exposure 2253 2254 Unithiol was used in a 21-year-old diabetic male who developed mercury intoxication 2255 following use of a mercury-containing ointment for eczema for about 3 weeks. He 2256 developed classic signs of mercury poisoning with tiredness, sweating, mild 2257 fasciculation of extremities, ataxia, hand tremors, weight loss, proteinuria, anxiety 2258 and behavioural changes. The urinary mercury concentration was 0.252 mg/L and 2259 he was given oral unithiol for 12 days. The highest urine mercury concentration 2260 during therapy was 2.1 mg/L. He improved rapidly over a 2 week period, however, 2261 signs of neurological and renal damage, not typical of diabetes persisted (Pelclová et 2262 al., 2001). 2263 2264 Use of a mercury-containing cosmetic bleaching cream for 4 years caused nail 2265 dyschromia in a 56 year female. The nails were discoloured greenish-black and she 2266 had features of mercury toxicity with night sweats, insomnia and nervousness. 2267 Treatment with unithiol reduced the serum mercury concentration from 64 to15 µg/L; 2268 the urinary mercury concentration rose and peaked at 1660 µg/L on the 10th day. 2269 After the initial course of unithiol the serum mercury concentration increased and she 2270 was given a second course, lasting 3 weeks. The unithiol was well tolerated (Böckers 2271 et al., 1985). 2272 2273 11.11.5 Parenteral mercury exposure 2274 2275 Long-term unithiol administration has been used in the management of parenteral 2276 mercury poisoning. A 23-year-old male injected about 20 ml of metallic mercury

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2277 intravenously and mercury was visible on x-ray in the right ventricle with multiple 2278 emboli in the lung. Mercury was also seen in the abdomen and right forearm. Only 2279 0.2 ml was removed from the heart by cardiac catheterisation and the blood mercury 2280 concentration rose to 294 µg/L. He was started on oral unithiol 300-800 mg a day and 2281 this continued for at least 4.5 years. The blood mercury and urine concentrations 2282 peaked at 1608 µg/L and 73,500 µg/L, respectively. He remained well except for 2283 intermittent chest pain. He developed no adverse effects to unithiol, and no reduction 2284 in plasma zinc, copper or selenium concentrations, although the urine copper 2285 concentrations were elevated (Ashton et al., 1992b). Batora et al. (2000) also used 2286 unithiol in a patient with intravenous mercury injection with elevated blood and urine 2287 concentrations (21.4 µg/L and 183.3 μg/L, respectively). They used only two courses 2288 of treatment over 17 days, and 6 weeks after therapy the blood concentration had 2289 fallen to 8.1 µg/L and the urine concentration increased to 397.6 µg/L. Mercury 2290 deposits were visible on both lung fields but by 1 year had almost completely 2291 disappeared. He remained well with only a temporary enzymuria (N-acetyl beta 2292 glucosaminidase) indicating subtle renal tubular damage. 2293 2294 A 35-year-old male presented with a history of gingivitis, weakness, pyrexia, anorexia 2295 and weight loss 6 weeks after intravenous injection of metallic mercury. Mercury was 2296 visible in the lungs, abdomen and injection site on X-ray and the urinary concentration 2297 was 500 µg/L. He was started on unithiol 2 days after admission on a 6 day course of 2298 600 mg/day in 3 divided doses. After the first 24 hours of treatment the urinary 2299 mercury concentration rose to 7750 µg/L and he developed a moderate 2300 hypersensivity-type skin reaction with resolved within 2 days. He was discharged 2301 one week after completion of unithiol therapy and had urine concentration 1100 µg/L 2302 with irritability and weakness. He did not return for a year when he presented with 2303 irritability, weakness, insomnia and tremor. The urinary mercury concentration was 2304 2206 µg/L. Mercury was still visible on X-ray and he was given another course of 2305 unithiol. Over the next 4 years he was reviewed every 6 months and the urinary 2306 mercury concentration was 800-1000 µg/L. Two years after the original presentation 2307 he was given a third course of unithiol. At 5 years the concentration was 807 µg/L 2308 and a fourth course of untihiol produced a less dramatic increase in urinary 2309 elimination. Tremor and weakness persisted. Although unithiol increased 2310 elimination there appeared to be no change in the radiographic deposits mercury in 2311 the lungs and abdomen. It was not clear that the infrequent administration of unithiol 2312 was of any benefit to this patient (Torres-Alanís et al., 1997). 2313 2314 In another case of elemental mercury injection, a 27-year-old male, 65 kg, injected 2315 1.5 mL (20 g) into his left cubital vein. Within 12 hours he developed pyrexia, 2316 tachycardia and dyspnoea and mercury was visible on chest X-ray. The serum 2317 mercury concentration was 172 µg/L on admission and peaked on day 6 at 274 µg/L. 2318 At 37 hours he was started on unithiol (200 mg orally every 8 hours) for 5 days, 2319 during which time he eliminated 8 mg of mercury. Three days later he was started 2320 on succimer (500 mg daily) for another 5 days. This resulted in the excretion of 2321 another 3 mg of mercury. Neither unithiol nor succimer were effective in enhancing 2322 elimination of mercury after intravenous injection (Eyer et al., 2006). 2323 2324 Unithiol was shown to be effective in enhancing mercury elimination and reducing 2325 renal irradiation in patients given radioactive chlormerodrin, an obsolete mercurial 2326 diuretic, for renal scintigraphy (Ogiński & Kloczkowski, 1973; Kloczkowski & Ogiński,

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2327 1973). 2328 2329 11.12 Use in nickel poisoning 2330 2331 There are no case reports of unithiol administration in nickel poisoning but it has been 2332 recommended (Daunderer, 1982). 2333 2334 11.13 Use in palladium poisoning 2335 2336 No well documented case reports of unithiol use in platinum poisoning could be 2337 found. 2338 2339 11.14 Use in platinum poisoning 2340 2341 No well documented case reports of unithiol use in platinum poisoning could be 2342 found. 2343 2344 11.15 Use in polonium poisoning 2345 2346 Experience with use of unithiol in polonium exposure is limited. Shantyr et al. (1969) 2347 reported 10 children who were contaminated with polonium-210 from a damaged 2348 polonium-beryllium neutron source. They had body burdens of 0.2-7.0 µCi, far 2349 above the maximal permissible burden. Some were treated with unithiol (number 2350 unknown) and all remained well with no changes in general health, blood or renal 2351 function over the 46 month period of monitoring. However, most of the children 2352 developed impairment of protein formation in the liver, manifested as an increase in 2353 albumin and a decrease in globulin. This was observed from 21 months and 2354 persisted through the remaining period of observation. 2355 2356 11.16 Use in selenium poisoning 2357 2358 No well documented case reports of unithiol use in selenium poisoning could be 2359 found. 2360 2361 11.17 Use in silver poisoning 2362 2363 There is limited information on the use of unithiol in silver poisoning. Unithiol was 2364 used in a patient with after penicillamine had failed to increase the urinary 2365 excretion of silver. The patient was 60 years old and had developed argyria after 15 2366 years use of silver nitrate to treat gingivitis due to ill-fitting dentures. Unithiol was 2367 given at a dose of 1-5 x 500 mg for 5 days then 2.5 g/day for another 5 days with a 5 2368 day period in between. Even though the renal excretion of silver was increased by 2369 unithiol administration the total amount of excreted silver was low, only 1 µmol of silver 2370 in total. This was estimated to be approximately 1% or less of the total body burden of 2371 silver and it was concluded that unithiol was of no benefit in argyria (Aaseth et al., 2372 1986). 2373 2374 In a 55-year-old patient with argyria, use of unithiol (3 x 100 mg/day) increased the 2375 urinary excretion of silver by approximately 100 fold. However, the total quantity of 2376 silver excreted was low. Administration of penicillamine did not affect silver excretion

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2377 (Kemper et al., 1989; Jekat and Kemper, 1990). 2378 2379 11.18 Use in strontium poisoning 2380 2381 No well documented case reports of unithiol use in strontium poisoning could be 2382 found. 2383 2384 11.19 Use in 2385 2386 There is limited information on the use of unithiol in tin poisoning. Unithiol was used 2387 in dental assistant with tin exposure due to kneading of amalgam in the unprotected 2388 palm of the hand. The urinary concentration of tin was increased to 1094.4 µg/L with 2389 unithiol administration and clinical features (tiredness, dizziness and tremor) improved 2390 (Hruschka, 1990). 2391 2392 2393 12. Summary of evaluation 2394 2395 12.1 Indications 2396 2397 Unithiol appears to be effective (in terms of accelerating metal excretion without 2398 causing severe adverse effects) in most cases of: 2399 • acute and chronic intoxication by organic and inorganic mercury 2400 • acute and chronic intoxication by bismuth 2401 • chronic lead poisoning 2402 • acute and chronic arsenic poisoning. 2403 2404 It has also been used with some success in cases of human poisoning with the 2405 following, but data are limited: 2406 • trivalent antimony (there is no information on the less toxic pentavalent 2407 antimony compounds) 2408 • chromium 2409 • cobalt 2410 • copper, including patients with Wilson’s disease 2411 • gold 2412 2413 Animal studies have demonstrated apparent benefit but experience of unithiol use in 2414 human poisoning is lacking for the following: 2415 • beryllium 2416 • cadmium 2417 • nickel 2418 • tin 2419 • zinc 2420 2421 On the basis of animal or human case reports unithiol does not appear to be useful for 2422 the following: 2423 • palladium 2424 • platinum 2425 • polonium

49

2426 • silver (argyria) 2427 • strontium 2428 • thallium 2429 • vanadium 2430 2431 See Table 5 (section 16.2) for a summary of the available information on unithiol use 2432 in metal and metalloid poisoning. 2433 2434 12.2 Advised routes and dose 2435 2436 Unithiol may be administered both orally and parenterally. In cases of acute heavy 2437 metal ingestion the parenteral route is strongly recommended, because given orally 2438 the drug may bind residues of the metal in the gut, promote absorption and decrease 2439 body burden. 2440 2441 High dose therapy should be avoided in patients with chronic poisoning without severe 2442 clinical symptoms because of the risk of sudden mobilisation of the metal from tissue 2443 stores with resultant clinical deterioration. This occurred, for example, in a patient with 2444 chronic bismuth toxicity, necessitating cessation of unithiol therapy (Teepker et al., 2445 2002). 2446 2447 12.2.1 Oral administration 2448 2449 There is no standard regimen for unithiol administration; dosing and duration depends 2450 on clinical condition and blood and urine concentrations of the metal. High doses 2451 have been used and are well tolerated, but are not advised in chronically poisoned 2452 patients without severe clinical effects (see section 12.2). Unithiol should be taken on 2453 an empty stomach. 2454 • Adults: The usual initial dose is 100-200 mg every 6-8 hours (3-4 times/day); this is 2455 then tapered over the following days or weeks. 2456 • Children: The usual initial dose 50-100 mg every 6-8 hours (3-4 times/day); this is 2457 then tapered over the following days or weeks. Alternatively 5 mg/kg daily in 2 to 4 2458 divided doses has been used (Willig et al., 1984; Chisholm and Thomas, 1985; 2459 Karpinski & Markoff, 1997; Böse-O’Reilly et al., 2003). 2460 2461 12.2.2 Parenteral administration 2462 2463 Unithiol may be administered parenterally by the intramuscular or slow intravenous 2464 injection over 5 minutes (1 mL/minute). The parental doses of unithiol for adults and 2465 children are given in Table 1. From day 4 frequency of dosing should be based on the 2466 patient’s clinical condition; parental unithiol may be continued or oral treatment may be 2467 started. Unithiol solution must be administered immediately after opening the vials, 2468 and any remaining after dosing must be discarded. Parenteral unithiol must not be 2469 mixed with other infusion solutions (as this may reduce antidotal efficacy). 2470

50

2471 Table 1: Parenteral unithiol dosage 2472 Day of Adults Children treatment Dose IM or Total daily Dose IM or slow IV Total daily slow IV dose dose 1 250 mg every 3- 1.5-2.0 g 5 mg/kg every 3-4 30-40 mg/kg 4 hours hours 2 250 mg every 4- 1.0-1.5 g 5 mg/kg every 4-6 20-30 mg/kg 6 hours hours 3 250 mg every 6- 0.75-1.0 g 5 mg/kg every 6-8 15-20 mg/kg 8 hours hours 4,5… 250 mg every 8- 0.50- 0.75 g 5 mg/kg every 8-12 10-15 mg/kg 12 hours hours 2473 2474 12.3 Supportive therapy 2475 2476 Other supportive therapy, and gut decontamination, rehydration or cardiovascular 2477 support may be required. Haemofiltration in conjunction with unithiol administration is 2478 the renal replacement method of choice in patients with renal failure because it may 2479 enhance elimination of heavy metals (particularly mercury). 2480 2481 In cases of chronic poisoning identification and removal from source should also be 2482 undertaken. 2483 2484 12.4 Controversial issues 2485 2486 There are indications from animal studies that unithiol may be useful in the treatment of 2487 acute poisoning by beryllium, cadmium, nickel and tin and from a small number of 2488 human case reports for trivalent antimony, chromium, cobalt, copper and gold. 2489 However, there is a lack of good quality clinical data. 2490 2491 Diagnostic use of unithiol in cases of asymptomatic or suspected poisoning cannot be 2492 recommended. The metal-binding agent mobilises the metal resulting in redistribution 2493 which could increase the concentration at the target organ, despite an increased 2494 excretion. 2495 2496 It is still controversial, whether concurrent administration of more than one metal- 2497 binding agent is harmful or beneficial. If adequate doses of unithiol are given, are well 2498 tolerated and the duration of therapy long enough, then use of more than one metal- 2499 binding agent is not necessary. 2500 2501 12.5 Proposals for further studies 2502 2503 The mode of action of unithiol has not been fully elucidated and recent work has 2504 tended to focus on arsenic and mercury. Further study is required to determine the 2505 interaction of unithiol with individual metals and metalloids but also with the target 2506 organs. Modern tools, such as computer molecule modelling for complex formation, 2507 should be used to provide a better understanding of the biological processes involved.

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2508 There is also a clear need for more clinical trials on the role of unithiol in the treatment 2509 of poisoned patients, in particular to define the optimum dose, duration of therapy and 2510 route of administration of unithiol and compare it to other metal-binding agents such as 2511 succimer and sodium calcium edetate. There are large populations of individuals 2512 poisoned with metals and metalloids, particularly arsenic and mercury, from 2513 environmental sources. For the metals which are less commonly involved in poisoning 2514 and where clinical trials are unlikely to be practical unless a mass incident occurs, there 2515 is a need for well documented case reports with biochemical and chemical analyses 2516 used to determine the efficacy of the antidote used. 2517 2518 Evaluation of the risks and benefits of combined antidotal therapy is also an area that 2519 warrants father investigation. 2520 2521 The suggestion that unithiol is the optimal antidote for inorganic mercury poisoning 2522 and that succimer is more effective in organic mercury intoxication requires verification 2523 (Andersen, 1999). 2524 2525 12.6 Adverse effects 2526 2527 Unithiol is generally well tolerated and the incidence of adverse effects is low. 2528 2529 Administration of unithiol also increases elimination of some trace elements, particularly 2530 zinc and copper (Bertram, 1977; Mant, 1985; Aaseth et al., 1986; Sallsten et al., 1994; 2531 Torres- Alanís et al., 2000; Høl et al., 2003), but also selenium and magnesium 2532 (Torres- Alanís et al., 2000). This effect is only likely to be of clinical significance in 2533 patients on chronic unithiol therapy. 2534 2535 Skin reactions including rashes, pruritis and blistering have been reported (Dubinsky & 2536 Guida, 1979; Mant, 1985; Ashton et al., 1992a; Hla et al., 1992; Toet et al., 1994; 2537 Torres- Alanís et al., 1995; Torres-Alanís et al., 1997; Gonzalez-Ramirez et al., 1998; 2538 Böse-O’Reilly et al., 2003; Dargan et al., 2003a). Erytheme multiforme with buccal 2539 ulceration (Ashton et al., 1992a) and depigmentation (Pagliuca et al., 1990) has been 2540 reported. Stevens-Johnson syndrome has been reported in a small number of cases 2541 (Chisholm, 1990; Chisholm, 1992; Van der Linde et al., 2008). Anaphylactic shock has 2542 not been reported (Ruprecht, 1997). In most cases allergic reactions have resolved 2543 within 3-5 days and generally no treatment is required. However, antihistamines and/or 2544 corticosteroids may be given if necessary. 2545 2546 Nausea may occur from oral administration (Lund et al., 1984; Walshe, 1985; 2547 Stevens et al., 1995; Gonzalez-Ramirez et al., 1998), and body fluids usually have a 2548 sulphur odour for 6-8 hours after unithiol administration. Mild elevation of liver 2549 enzymes (Chisolm & Thomas, 1985; Gonzalez-Ramirez et al., 1998; Wang et al., 2550 2003), diuresis (Glukharev, 1965), fever and leucocytosis (Walshe, 1985) have 2551 been reported. 2552 2553 With the parenteral preparation cardiovascular reactions may occur, particularly if 2554 injected too rapidly. These effects are hypotension (Hurlbut et al., 1994), nausea 2555 (Stevens et al., 1995), dizziness and weakness (Dubinsky & Guida, 1979; Zhang, 2556 1984). Necrosis and ulceration may occur at the injection site, but this is associated 2557 with high doses e.g. 100 mg/kg (Sanotsky et al., 1967).

52

2558 12.7 Restrictions of use 2559 2560 Unithiol should not be administered in acute arsine poisoning (AsH3), because it is 2561 ineffective and can increase arsine toxicity (Mizyukova & Petrunkin, 1974). 2562 2563 The administration of unithiol in cases of asymptomatic or suspected poisoning (a 2564 challenge or mobilisation test) cannot be recommended, because the metal-binding 2565 agent mobilises the metal from tissue stores resulting in redistribution and potentially 2566 increasing the concentration in the target organ, despite increasing excretion. 2567 2568 Renal impairment is not a restriction of use; haemofiltration is the renal replacement 2569 method of choice in patients with renal failure and in conjunction with unithiol 2570 administration may enhance elimination of heavy metals (particularly mercury). 2571 2572 2573 13. Model information sheet 2574 2575 13.1 Uses 2576 2577 Unithiol is a derivative of dimercaprol (2,3-dimercapto-1-propanol, British Anti-Lewisite, 2578 BAL), and is replacing dimercaprol as one of the main antidotes used in the 2579 management of heavy metal poisoning. Unithiol has several advantages over 2580 dimercaprol including lower toxicity, increased solubility in water and lower lipid 2581 solubility. It is due to these properties that it is effective by oral administration. 2582 2583 Unithiol has been used in the management of acute and chronic poisoning with a 2584 number of different metals and metalloids, and is particularly useful for arsenic, 2585 bismuth and mercury. It has been used for other metals and metalloids. Unithiol can 2586 be given parenterally or orally depending on the clinical situation and severity of 2587 poisoning. 2588 2589 13.2 Dosage and route 2590 2591 Unithiol may be administered both orally and parenterally. In severe cases and/or 2592 acute poisoning parenteral administration is recommended. In cases of acute heavy 2593 metal ingestion the parenteral route is strongly recommended, because given orally 2594 the drugs may bind residues of the metal in the gut, promote the absorption and 2595 increase the body burden by this way. 2596 2597 High dose therapy should be avoided in patients with chronic poisoning without severe 2598 clinical symptoms because of the risk of sudden mobilisation of the metal from tissue 2599 stores with resultant clinical deterioration. 2600 2601 13.2.1 Oral administration 2602 2603 There is no standard regimen for unithiol administration; dosing and duration depends 2604 on clinical condition and blood and urine concentrations of the metal. High doses 2605 have been used and are well tolerated, but are not advised in chronically poisoned 2606 patients without severity clinical effects (see section 12.2). Unithiol should be taken on 2607 an empty stomach.

53

2608 • Adults: The usual initial dose is 100-200 mg every 6-8 hours (3-4 times/day); this is 2609 then tapered over the following days or weeks. 2610 • Children: The usual initial dose 50-100 mg every 6-8 hours (3-4 times/day); this is 2611 then tapered over the following days or weeks. Alternatively 5 mg/kg daily in 2 to 4 2612 divided doses has been used. 2613 2614 13.2.2 Parenteral administration 2615 2616 Unithiol is given by IM or slow IV injection over 5 minutes (1 mL/minute). See table for 2617 dosing regimen. From day 4 frequency of dosing should be based on the patient’s 2618 clinical condition; parental unithiol may be continued or oral treatment may be started. 2619 Unithiol must not be mixed with other infusion solutions, as this may reduce its 2620 efficacy. 2621 2622 Table: Parenteral unithiol dosage 2623 Day of Adults Children treatment Dose IM or slow Total daily Dose IM or slow IV Total daily IV dose dose 1 250 mg every 3-4 1.5-2.0 g 5 mg/kg every 3-4 30-40 mg/kg hours hours 2 250 mg every 4-6 1.0-1.5 g 5 mg/kg every 4-6 20-30 mg/kg hours hours 3 250 mg every 6-8 0.75-1.0 g 5 mg/kg every 6-8 15-20 mg/kg hours hours 4,5… 250 mg every 8- 0.50-0.75 g 5 mg/kg every 8-12 10-15 mg/kg 12 hours hours 2624 2625 13.3 Precautions and contraindications 2626 2627 Unithiol should not be administered in acute arsine poisoning (AsH3), because it is 2628 ineffective and can increase arsine toxicity. 2629 2630 The administration of unithiol in cases of asymptomatic or suspected poisoning (a 2631 challenge or mobilisation test) cannot be recommended, because the metal-binding 2632 agent mobilises the metal from tissue stores resulting in redistribution and potentially 2633 increasing the concentration in the target organ, despite increasing excretion. 2634 2635 The administration of more than one metal-binding agent at the same time cannot be 2636 recommended, as the risks and/or benefits of such therapy have not been evaluated. 2637 2638 The efficacy of a metal-binding agent may be difficult to determine. After 2639 discontinuation of metal exposure (and absorption) a decrease in the blood 2640 concentration will occur without any therapy. Clinical efficacy should not be judged 2641 only by the quantity of metal excretion or the decrease of blood concentrations. The 2642 reduction of the tissue content in the target organ and the restoration of pathological 2643 alterations also need to be considered. It is important to note that enhancement of the

54

2644 metal excretion by mobilisation may increase the metal burden of the target organ by 2645 redistribution, and conversely the body burden may be reduced without a striking 2646 decrease of the blood concentrations. 2647 2648 During long-term therapy the blood concentrations and excretion of trace elements 2649 should be monitored carefully, because depletion of trace metals may play a role in the 2650 toxicity of metal-binding agent agents. 2651 2652 13.4 Pharmaceutical incompatibilities and drug interactions 2653 2654 Unithiol must not be mixed with other infusion solutions, as this may reduce antidotal 2655 efficacy. 2656 2657 Unithiol solution should be administered immediately after opening of the vials and all 2658 remainder must be discarded, because the compound is oxidised rapidly in contact 2659 with air. 2660 2661 Unithiol should not be given orally with mineral preparations or activated charcoal 2662 because unithiol may be inactivated. For the same reason the unithiol capsules 2663 should be taken at least one hour before a meal. 2664 2665 13.5 Side effects 2666 2667 Unithiol is generally well tolerated and the incidence of adverse effects is low. 2668 2669 Administration of unithiol also increases elimination of some trace elements, particularly 2670 zinc and copper, but also selenium and magnesium. This effect is only likely to be of 2671 clinical significance in patients on chronic unithiol therapy. 2672 2673 Skin reactions including rashes, pruritis and blistering have been reported. Erytheme 2674 multiforme with buccal ulceration and depigmentation has been reported. Stevens- 2675 Johnson syndrome has been reported in a small number of cases. Anaphylactic shock 2676 has not been reported. In most cases allergic reactions have resolved within 3-5 days 2677 and generally no treatment is required. However, antihistamines and/or corticosteroids 2678 may be given if necessary. 2679 2680 Nausea may occur from oral administration, and body fluids usually have a sulphur 2681 odour for 6-8 hours after unithiol administration. Mild elevations of liver enzymes, 2682 diuresis, fever and leucocytosis have also been reported. 2683 2684 With the parenteral preparation cardiovascular reactions may occur, particularly if 2685 injected too rapidly. These effects are hypotension, nausea, dizziness and weakness. 2686 2687 Necrosis and ulceration may occur at the injection site, but this is associated with high 2688 doses. 2689 2690 13.6 Pregnancy and lactation 2691 2692 Teratogenic effects have not been demonstrated in animal studies, indeed studies 2693 have demonstrated that unithiol can protect against the developmental toxicity of

55

2694 arsenic and mercury. Even though safety in human has not been established, 2695 pregnancy is not regarded as a contraindication. If unithiol is administered to 2696 pregnant women, the essential minerals should be monitored carefully, because 2697 metal-binding may cause a depletion of trace elements and it has been shown that 2698 zinc deficiency can cause teratogenic effects. 2699 2700 In general lactation should be avoided in metal poisoning. 2701 2702 13.7 Storage 2703 2704 The capsules should be stored in a dry place. 2705 2706 The shelf-life for the commercial available pharmaceutical preparations Dimaval® is 2707 claimed to be 5 years for the capsules 4 years for the ampoules. The expiry date is 2708 stated on each package. 2709 2710 2711 14. References 2712 2713 Aaseth J (1983) Recent advances in the therapy of metal with chelating 2714 agents. Hum Toxicol, 2: 257-272. 2715 2716 Aaseth J, Alexander J & Raknerud N (1982) Treatment of mercuric chloride poisoning 2717 with dimercaptosuccinic acid and diuretics: preliminary studies. J Toxicol Clin Toxicol, 2718 19: 173-186. 2719 2720 Aaseth J, Skaug V & Alexander J (1984) Haemolytic activity of copper as influenced 2721 by chelating agents, albumine and chromium. Acta Pharmacol Toxicol, 54: 304-310. 2722 2723 Aaseth J, Halse J & Falch J (1986) of silver in argyria. Acta Pharmacol 2724 Toxicol (Copenh), 59 (Suppl 7): 471-474. 2725 2726 Adam B, Felgenhauer N, Zilker T (2003) DMPS: the new chelating agent of choice in 2727 the treatment of arsenic poisoning [abstract]. J Toxicol Clin Toxicol, 41: 440. 2728 2729 Anatovskaya VS (1962) [The use of unithiol in the treatment of chronic lead 2730 intoxication] (Russian). Gigieny Truda Profzabol, 29: 50-56. Cited in: Ruprecht J th 2731 (1997) Scientific monograph Dimaval® (DMPS), 6 edition. Berlin, Heyl Company. 2732 2733 Andersen O (1999) Principles and recent developments in chelation treatment of 2734 metal intoxication. Chem Rev, 99: 2683-2710. 2735 2736 Andersen O & Nielsen JB (1988) Oral cadmium chloride intoxication in mice: effects 2737 of penicillamine, dimercaptosuccinic acid and related compounds. Pharmacol 2738 Toxicol, 63: 386-9. 2739 2740 Anderson RA, McAllister WA & Taylor A (1996) Acute mercuric iodide poisoning. 2741 Ann Clin Biochem, 33: 468-470. 2742 2743 Angle CR (1993) Childhood lead poisoning and its treatment. Annu Rev Pharmacol

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2744 Toxicol, 33: 409-434. 2745 2746 Anon (1992/1993) [Unpublished studies on Dimaval® (DMPS) and DMPS-Heyl®] 2747 (German). Heyl company. Cited in: Ruprecht J (1997) Scientific monograph Dimaval® th 2748 (DMPS), 6 edition. Berlin, Heyl Company. 2749 2750 Aposhian HV, Tadlock CH & Moon TE (1981) Protection of mice against lethal 2751 effects of sodium arsenite - a quantitative comparison of a number of chelating 2752 agents. Toxicol Appl Pharmacol, 61: 385-392. 2753 2754 Aposhian HV (1982) Biological chelation: 2,3-dimercapto-propanesulfonic acid and 2755 meso-dimercaptosuccinic acid. Adv Enzyme Regul, 20: 301-319. 2756 2757 Aposhian HV, Mershon MM, Brinkley FB, Hsu CA & Hackley BE (1982) Anti-lewisite 2758 activity and stability of meso-dimercaptosuccinic acid and 2,3-dimercapto-1- 2759 propanesulfonic acid. Life Sci, 31: 2149-2156. 2760 2761 Aposhian HV (1983) DMSA and DMPS - water soluble antidotes for heavy metal 2762 poisoning. Ann Rev Pharmacol Toxicol, 23: 193-215. 2763 2764 Aposhian HV, Carter DE, Hoover TD, Hsu CA, Maiorino RM & Stine E (1984) DMSA, 2765 DMPS, and DMPA - as arsenic antidotes. Fundam Appl Toxicol, 4: S58-S70. 2766 2767 Aposhian HV, Dart RC, Aposhian MM & Dawson BV (1987) Tissue decorporation of 2768 polonium-210 in rats by DMPA. Res Commun Chem Pathol Pharmacol, 58: 157-171. 2769 2770 Aposhian HV, Maiorino RM, Gonzalez-Ramirez D, Zuniga-Charles M, Xu Z, Hurlbut 2771 KM, Junco-Munoz P, Dart RC & Aposhian MM (1995) Mobilization of heavy metals 2772 by newer, therapeutically useful chelating agents. Toxicology, 97: 23-38. 2773 2774 Aposhian MM, Maiorino RM, Xu Z & Aposhian HV (1996) Sodium 2,3-dimercapto-1- 2775 propanesulfonate (DMPS) treatment does not redistribute lead or mercury o the 2776 brain of rats. Toxicology, 109: 49-55. 2777 2778 Aposhian HV, Arroyo A, Cebrian ME, del Razo LM, Hurlbut KM, Dart RC, Gonzalez- 2779 Ramirez D, Kreppel H, Speisky H, Smith A, Gonsebatt ME, Ostrosky-Wegman P & 2780 Aposhian MM (1997) DMPS-arsenic challenge test. I: Increased urinary excretion of 2781 monomethylarsonic acid in humans given dimercaptopropane sulfonate. J 2782 Pharmacol Exp Ther, 282: 192-200. 2783 2784 Ashton CE & House I (1989) Two cases of severe inorganic mercury ingestion treated 2785 with dimercapto-1-propane sulphonate [abstract]. Annual Meeting of the European 2786 Association of Control Centres, Münster. 2787 2788 Ashton CE, Hla KK, Mant T & Volans G (1992a) 2,3-Dimercaptopropane-1-sulfonate 2789 (DMPS) in the treatment of heavy metal poisoning, an effective and potentially life 2790 saving treatment [abstract]. Annual Meeting of the European Association of Toxicology 2791 and Poison Control Centres, Istanbul. 2792 2793 Ashton CE, House I & Volans G (1992b) Severe intravenous metallic mercury

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2794 poisoning successfully treated with long-term 2,3-Dimercaptopropane-1-sulfonate 2795 (DMPS) [abstract]. Annual Meeting of the European Association of Toxicology and 2796 Poison Control Centres, Istanbul. 2797 2798 Autenrieth T, Schmidt T & Habscheid W (1998) [Lead poisoning caused by a Greek 2799 ceramic cup] (German). Dtsch Med Wochenschr, 123: 353-358. 2800 2801 Bakir F, Al-Kalidi A, Clarkson TW & Greenwood MR (1976) Clinical observations on 2802 treatment of alkylmercury poisoning in hospital patients. Bull WHO, 53: 87-92. 2803 2804 Bakka A, Aaseth J & Rugstad HE (1981) Influence of certain chelating agents on 2805 egress of cadmium from cultured epithelial cells containing high amounts of 2806 metallothionein: a screening of Cd-releasing and toxic effects. Acta Pharmacol 2807 Toxicol (Copenh), 49: 432-437. 2808 2809 Basinger MA, Jones MM & Tarka MP (1980) Relative efficacy of chelating agents as 2810 antidotes for acute nickel (II) acetate intoxication. Res Commun Chem Pathol 2811 Pharmacol, 30: 133-141. 2812 2813 Basinger MA & Jones MM (1981a) Structural requirements for chelate antidotal 2814 efficacy in acute antimony (III) intoxication. Res Comm Chem Pathol Pharmacol, 32: 2815 355-363. 2816 2817 Basinger MA & Jones MM (1981b) Chelate antidotal efficacy in acute zinc 2818 intoxication. Res Commun Chem Pathol Pharmacol, 33: 263-273. 2819 2820 Basinger MA, Jones MM & McCroskey SA (1983) Antidotes for acute bismuth 2821 intoxication. J Toxicol Clin Toxicol, 20: 159-165. 2822 2823 Basinger MA, Gibbs SJ, Forti RL, Mitchell WM & Jones MM (1985) Antidotes for 2824 gold (sodium bis[thiosulfato]gold[I]) intoxication in mice. J Rheumatol, 12: 274-278. 2825 2826 Basinger MA, Jones MM, Craft WD, Walker EM & Sanders MM (1987) Chelating-agent 2827 suppression of cadmium-induced hepatotoxicity. J Toxicol Environ Health, 22: 261- 2828 271. 2829 2830 Basinger MA, Jones MM, Holscher MS & Vaughn WK (1988) Antagonists for acute 2831 oral cadmium chloride intoxication. J Toxicol Environ Health, 23: 77-89. 2832 2833 Batora , Teply I, Ulicna O, Kostolanska K, Urbanova E, Plackova S & Kresanek J 2834 (2001) Intrabronchial aspiration of elemental mercury: a 14-month follow-up [abstract]. 2835 J Toxicol-Clin Toxicol, 39: 271. 2836 2837 Berlin M & Ullberg S (1963) Increased uptake of mercury in mouse brain caused by 2838 2,3-dimercaptopropanol (BAL). Nature, 197: 84-85. 2839 2840 Berman E (1980) Toxic metals and their analysis. London, Heyden. 2841 2842 Bertram HP (1977) Influence of chelating agents in chronic mercury poisoning. Naunyn 2843 Schmiedebergs Arch Pharmacol, 297 (S2): 19.

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3294 Maiorino RM, Dart RC, Carter DE & Aposhian HV (1991) Determination and 3295 metabolism of dithiol chelating agents. XII. Metabolism and pharmacokinetics of 3296 sodium 2,3-dimercaptopropane-1-sulfonate in humans. J Pharmacol Exp Ther, 259: 3297 808-814. 3298 3299 Maiorino RM, Xu ZF & Aposhian HV (1996) Determination and metabolism of dithiol 3300 chelating agents. XVII. In humans, sodium 2,3-dimercaptopropanesulfonate is bound 3301 to plasma albukmin via mixed disulfide formation and is found in the urine as cyclic 3302 polymeric disulfides. J Pharmacol Exp Ther, 277: 375-384. 3303 3304 Mant TGK (1985) Clinical studies with dimercaptopropane sulphonate in mercury 3305 poisoning. Hum Toxicol, 4: 346. 3306 3307 Mathur S, Flora SJ, Mathur R, Kannan GM & Das Gupta S (1994) Beryllium- 3308 induced biochemical alterations and their prevention following co-administration of 3309 meso-2,3-dimercaptosuccinic acid or 2,3-dimercaptopropane sulphonate in rats. J 3310 Appl Toxicol, 14: 263-267. 3311 3312 Merkord J, Weber H, Kröning G & Hennighausen G (2000) Antidotal effects of 2,3- 3313 dimercaptopropane-1-sulfonic acid (DMPS) and meso-2,3-dimercaptosuccinic acid 3314 (DMSA) on the organotoxicity of dibutyltin dichloride (DBTC) in rats. Hum Exp 3315 Toxicol, 19: 132-137. 3316 3317 Mitchell WM, Basinger MA & Jones MM (1982) Antagonism of acute copper (II)- 3318 induced renal lesions by sodium 2,3 dimercaptopropanesulfonate. Johns Hopkins Med 3319 J, 151: 283-285. 3320 3321 Mizyukova IG & Petrunkin VG (1974) [Unithiol and mercaptid as antidotes in cases of 3322 poisoning by arsenic containing substances] (Russian). Vrach Delo, 2:126-129. Cited 3323 in Aposhian HV (1983) DMSA and DMPS - water soluble antidotes for heavy metal 3324 poisoning. Ann Rev Pharmacol Toxicol, 23: 193-215. 3325 3326 Moore DF, O`Callaghan CA, Berlyne G, Ogg CS, Davies HA, House IM & Henry JA 3327 (1994) Acute arsenic poisoning: absence of polyneuropathy after treatment with 2,3- 3328 dimercaptopropanesulfonate (DMPS). J Neurol Neurosurg Psychiatry, 57:1133-1135. 3329 3330 Mráz L, Sýkora J & Eybl V (1985) Palladium and chelating agents. Plzen Lek Sb, 49 3331 (suppl): 142-145. 3332 3333 Mückter H, Liebl B, Reichl FX, Hunder G, Walther U & Fichtl B (1997) Are we ready 3334 to replace dimercaprol (BAL) as an arsenic antidote? Hum Exp Toxicol, 16: 460-465. 3335 3336 Mulkey JP & Oehme FW (2000) Are 2,3-dimercapto-1-propanesulfonic acid or 3337 prussian blue beneficial in acute thallotoxicosis in rats? Vet Hum Toxicol, 42: 325- 3338 329. 3339 3340 Müller C, Bertram HP, Rau W & Morandini T (1989) Diagnosis and DMPS-treatment of 3341 accidental cobalt chloride ingestion-case report [abstract]. Annual Meeting of the 3342 European Association of Poison Control Centres, Münster. 3343

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3344 Nadig J, Knutti R & Hany A (1985) [DMPS treatment in acute sublimate (mercury 3345 chloride) poisoning] (German). Schweiz Med Wochenschr, 115: 507-511. Cited in: th 3346 Ruprecht J (1997) Scientific monograph Dimaval® (DMPS), 6 edition. Berlin, Heyl 3347 Company. 3348 3349 Nielsen JB & Andersen O (1991) Effect of four thiol-containing chelators on 3350 disposition of orally administered mercuric chloride. Hum Exp Toxicol, 10: 423-430. 3351 3352 Ogiński M & Kloczkowski K (1973) Use of unithiol for reducing radiation hazard of renal 203 203 3353 scintigraphy with chlormerodrin Hg. I. Estimation of the urinary excretion of Hg. 3354 Int Urol Nephrol, 5: 371-376. 3355 3356 Ovaska H, Wood DM, House I, Dargan PI, Jones AL & Murray S (2008) Severe 3357 iatrogenic bismuth poisoning with bismuth iodoform paraffin paste treated with DMPS 3358 chelation. Clin Toxicol, 46: 855-857. 3359 3360 Pagliuca A, Mufti GJ, Baldwin D, Lestas AN, Wallis RM & Bellingham AJ (1990) 3361 Lead poisoning: clinical, biochemical, and haematological aspects of a recent 3362 outbreak. J Clin Pathol, 43: 277-281. 3363 3364 Pai P, Thomas S, Hoenich N, Roberts R, House I & Brown A (2000) Treatment of a 3365 case of severe mercuric salt overdose with DMPS (dimercapo-1-propane 3366 sulphonate) and continuous haemofiltration. Nephrol Dial Transplant, 15: 1889-1890. 3367 3368 Paul M, Mason R & Edwards R (1989) Effect of potential antidotes on the acute 3369 toxicity, tissue disposition and elimination of selenium in rats. Res Commun Chem 3370 Pathol Pharmacol, 66: 441-450. 3371 3372 Pelclová D, Lukáš E, Urban P, Lebenhart P, Tesař V, Preiss J, Fenclová Z & 3373 Lebedová J (2001) Percutaneous mercury intoxication in young diabetic man 3374 [abstract]. J Toxicol Clin Toxicol, 39: 270-271. 3375 3376 Pethran A, Szinicz L & Forth W (1990) Effect of various dithiols on acute toxicity of 3377 different metals in mice. Plzen Lek Sb, 62 (suppl), 69-70. 3378 3379 Petrunkin VE (1956) [Synthesis and properties of dimercapto derivatives of 3380 alkylsulfonic acids. I: Synthesis of sodium 2,3-dimercaptopropylsulfonate (unithiol) and 3381 sodium 2-mercaptoethylsulfonate] (Russian). Ukr Khim Zh, 22: 603-607. 3382 3383 Pfab R, Muckter H, Roider G & Zilker T (1996) Clinical course of severe poisoning 3384 with thiomersal. J Toxicol Clin Toxicol, 34: 453-460. 3385 3386 Pingree SD, Simmonds PL & Woods JS (2001) Effects of 2,3-dimercapto-1- 3387 propanesulfonic acid (DMPS) on tissue and urine mercury levels following prolonged 3388 methylmercury exposure in rats. Toxicol Sci, 61: 224-233. 3389 3390 Planas-Bohne F (1977) The effect of mercuric chloride on the excretion of two urinary 3391 enzymes in the rat. Arch Toxicol, 37: 219-225. 3392 3393 Planas-Bohne F, Gabard B, & Schäffer EH (1980) Toxicological studies on sodium 2,3-

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3394 dimercaptopropane-1-sulfonate in the rat. Arzneimittelforschung, 30: 1291-1294. 3395 3396 Planas-Bohne F (1981) The effect of 2,3-dimercaptopropane-1-sulphonate and 3397 dimercaptosuccinic acid on the distribution and excretion of mercuric chloride in rats. 3398 Toxicology, 19: 275-278. 3399 3400 Planas-Bohne F & Olinger H (1981) The interaction of chelating agents with 3401 methylmercuric chloride bound to erythrocytes. Biochem Pharmacol, 30: 667-669. 3402 3403 Planas-Bohne F, Shand E & Taylor DM (1982) The effects of dimercaptosuccinic 3404 acid and other chelating agents on the retention of platinum in the rat kidney after 3405 treatment with cisplatin. Cancer Chemother Pharmacol, 9: 120-121. 3406 3407 Planas-Bohne F & Lehman M (1983) Influence on chelating agents on the distribution 3408 and excretion of cadmium in rats. Toxicol Appl Pharmacol, 67: 408-416. 3409 3410 Playford RJ, Matthews CH, Campbell MJ, Delves HT, Hla KK, Hodgson HJ & Calam 3411 J (1990) Bismuth induced encephalopathy caused by tri potassium dicitrato 3412 bismuthate in a patient with chronic renal failure. Gut, 31: 359-360. 3413 3414 Poluboiarinova ZI & Streltsova VN (1964) [On the mechanism of functional and 3415 morphological renal changes in radiation sickness (Po 210) treated with unithiol] 3416 (Russian). Med Radiol (Mosk), 18: 22-27. Cited in: Aposhian HV, Dart RC, Aposhian 3417 MM & Dawson BV (1987) Tissue decorporation of polonium-210 in rats by DMPA. Res 3418 Commun Chem Pathol Pharmacol, 58: 157-171. 3419 3420 Pudill R, Siebeck HJ, Kobberling J & Schubert GE (1989) [The treatment and 3421 clinical-toxicological course of a death from potassium dichromate poisoning] 3422 (German). GIT Labor Medizin, 12: 469-473. 3423 3424 Rau W, Planas-Bohne F & Taylor DM (1987) Influence of several chelating agents 3425 on the distribution and binding of cadmium in rats. Hum Toxicol, 6: 451-458. 3426 3427 Reichl FX, Kreppel H, Szinicz L, Mückter H, Fichtl B, Schumann K & Forth W (1990) 3428 Effect of chelating agents on biliary excretion of arsenic in perfused of guinea 3429 pigs pretreated with As2O3. Vet Hum Toxicol, 32: 223-226. 3430 3431 Reichl FX, Hunder G, Liebl B, Fichtl B & Forth W (1995) Effect of DMPS and various 3432 adsorbents on the arsenic excretion in guinea-pigs after injection with As2O3. Arch 3433 Toxicol, 69:712-717. 3434 3435 Rencová J, Volf V, Jones MM & Singh PK (1993) Relative effectiveness of dithiol 3436 and dithiocarbamate chelating agents in reducing retention of polonium-210 in rats. 3437 Int J Radiat Biol, 63: 223-232. 3438 3439 Renner G & Kramer HJ (1983) Studies on the after administration of 3440 chelate-forming agents in mice. Int J Clin Pharmacol Ther Toxicol, 21: 115-117. 3441 3442 Romanov SS (1967) [Unithiol as an antidote in pulmonary oedema secondary to 3443 intravenous injection of silver nitrate] (Russian). Farmakol Toksikol, 30: 237-238. Cited

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3494 Shinobu LA, Jones MM, Basinger MA, Mitchell WM, Wendel D & Razzuk A (1983) In 3495 vivo screening of potential antidotes for chronic cadmium intoxication. J Toxicol 3496 Environ Health, 12: 757-765. 3497 3498 Sinković A, Strdin A & Svenšek F (2008) Severe acute copper sulphate poisoning: a 3499 case report. Arh Hig Rada Toksikol, 59: 31-35. 3500 3501 Slikkerveer A, Jong HB, Helmich RB & de Wolff FA (1992) Development of a 3502 therapeutic procedure for bismuth intoxication with chelating agents. J Lab Clin Med, 3503 119: 529-537. 3504 3505 Slikkerveer A, Noach LA, Tytgat GN, Van der Voet GB & De Wolff FA (1998) 3506 Comparison of enhanced elimination of bismuth in humans after treatment with 3507 meso-2,3-dimercaptosuccinic acid and D,L-2,3-dimercaptopropane-1-sulfonic acid. 3508 Analyst, 123: 91-92. 3509 3510 Srivastava RC, Gupta S, Ahmad N, Hasan SK, Farookh A & Husain MM (1996) 3511 Comparative evaluation of chelating agents on the mobilization of cadmium: a 3512 mechanistic approach. J Toxicol Environ Health, 47: 173-182. 3513 3514 Stevens PE, Moore DF, House IM, Volans GN & Rainford DJ (1995) Significant 3515 elimination of bismuth by haemodialysis with a new heavy-metal chelating agent. 3516 Nephrol Dial Transplant, 10: 696-698. 3517 3518 Stewart JR & Diamond GL (1987) Renal tubular secretion of alkanesulfonate 2,3- 3519 dimercapto-1-propane sulfonate in rat kidney. Am J Physiol, 252: F800-F810. 3520 3521 Stewart JR & Diamond GL (1988) In vivo renal tubular secretion and metabolism of 3522 the disulfide of alkanesulfonate 2,3-dimercapto-1-propane sulfonate in rat kidney. Am 3523 J Physiol, 16: 189-195. 3524 3525 Susa N, Ueno S & Furukawa Y (1994) Protective effects of thiol compounds on 3526 chromate-induced toxicity in vitro and in vivo. Environ Health Perspect, 102 (Suppl 3527 3): 247-250. 3528 3529 Szincicz L, Wiedemann P, Häring H & Weger N (1983) Effects of repeated treatment 3530 with sodium 2,3-dimercaptopropane-1-sulfonate in beagle dogs. Drug Res, 33: 818- 3531 821. 3532 3533 Tadlock CH & Aposhian HV (1980) Protection of mice against the lethal effects of 3534 sodium arsenite by 2,3 dimercapto-1-propane-sulfonic acid and dimercaptosuccinic 3535 acid. Biochem Biophys Res Commun, 94: 501-507. 3536 3537 Takahashi Y, Funakoshi T, Shimada H & Kojima S (1994) Comparative effects of 3538 chelating agents on distribution, excretion, and renal toxicity of gold sodium 3539 thiomalate in rats. Toxicology, 90: 39-51. 3540 3541 Tandon SK, Singh S & Jain VK (1994) Efficacy of combined chelation in lead 3542 intoxication. Chem Res Toxicol, 7: 585-589. 3543

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3544 Tandon SK, Singh S, Jain VK & Prasad S (1996) Chelation in metal intoxication. 3545 XXXVIII: Effect of structurally different chelating agents in treatment of nickel 3546 intoxication in rat. Fundam Appl Toxicol, 31: 141-148. 3547 3548 Teepker M, Hamer HM, Knake S, Bandmann O, Oertel WH & Rosenow F (2002) 3549 Myoclonic encephalopathy caused by chronic bismuth abuse. Epileptic Disord, 4: 3550 229-233. 3551 3552 Toet AE, van Dijk A, Savelkoul TJ & Meulenbelt J (1994) Mercury kinetics in a case 3553 of severe mercuric chloride poisoning treated with dimercapto-1-propane sulphonate 3554 (DMPS). Hum Exp Toxicol, 13: 11-16. 3555 3556 Torres-Alanís O, Garza-Ocañas L & Piñeyro-López A (1995) Evaluation of urinary 3557 mercury excretion after administration of 2,3-dimercapto-1-propane sulfonic acid to 3558 occupationally exposed men. J Toxcol Clin Toxicol, 33: 717-720. 3559 3560 Torres-Alanís O, Garza-Ocañas L & Piñeyro-López A (1997) Intravenous self- 3561 administration of metallic mercury: report of a case with 5-year follow-up. J Toxicol Clin 3562 Toxicol, 35: 83-87. 3563 3564 Torres-Alanís O, Garza-Ocañas L, Bernal MA & Piñeyro-López A (2000) Urinary 3565 excretion of trace elements in humans after sodium 2,3-dimercaptopropane-1- 3566 sulfonate challenge test. J Toxcol Clin Toxicol, 38: 697-700. 3567 3568 Twarog T & Cherian MG (1983) Chelation of lead with DMPS and BAL in rats injected 3569 with lead. Bull Environ Contaim Toxicol, 30: 165-169. 3570 3571 Twarog T & Cherian MG (1984) Chelation of lead by dimercaptopropane sulfonate and 3572 a possible diagnostic use. Toxicol Appl Pharmacol, 72: 550-556. 3573 3574 Van der Linde AAA, Pillen S, Gerrits GPJM & Bouwes Bavinck JN (2008) Stevens- 3575 Johnson syndrome in a child with chronic mercury exposure and 2,3- 3576 dimercaptopropane-1-sulfonate (DMPS) therapy. Clin Toxicol, 46: 479-481. 3577 3578 Vantroyen B, Heilier JF, Meulemans A, Michels A, Buchet JP, Vanderschueren S, 3579 Haufroid V & Sabbe M (2004) Survival after a lethal dose of arsenic trioxide. J 3580 Toxicol Clin Toxicol, 42: 889-895. 3581 3582 Volf V (1973) The effect of chelating agents on the distribution of 210 Po in rats. 3583 Experientia, 29: 307-308. 3584 3585 Volf V, Rencová J, Jones MM & Singh PK (1995) Combined chelation treatment for 3586 polonium after simulated wound contamination in rat. Int J Radiat Biol, 68: 395-404. 3587 3588 von Mühlendahl KE (1990) Intoxication from mercury spilled on carpets. Lancet, 3589 336:1578. 3590 3591 Walker AW (1998) SAS Trace Element Laboratories. Clinical and analytical rd 3592 handbook, 3 edition. Guildford, Royal Surrey Hospital. 3593

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3594 Walshe JM (1985) Unithiol in Wilson's disease. Br Med J, 290: 673-674. 3595 3596 Wang XP, Yang RM, Ren MS & Sun BM (2003) Anticopper efficacy of captopril and 3597 sodium dimercaptosulphonate in patients with Wilson's disease. Funct Neurol, 18: 3598 149-53. 3599 3600 Wannag A & Aaseth J (1980) The effect of immediate and delayed treatment with 2,3- 3601 dimercapto-propane-1-sulphonate on the distribution and toxicity of inorganic mercury 3602 in mice and in foetal and adult rats. Acta Pharmacol Toxicol, 46: 81-88. 3603 3604 Wax PM & Thornton CA (2000) Recovery from severe arsenic-induced peripheral 3605 neuropathy with 2,3-dimercapto-1-propanesulphonic acid. J Toxicol Clin Toxicol, 38: 3606 777-780. 3607 14 14 3608 Wiedemann P, Fichtl B & Szinicz L (1982) Pharmacokinetics of C-DMPS ( C-2,3- 3609 dimercaptopropane-1-sulphonate) in beagle dogs. Biopharm Drug Dispos, 3: 267-74. 3610 3611 Wildenauer DB, Reuther H & Weger N (1982) Interactions of the chelating agent 2,3- 3612 dimercaptopropane-1-sulfonate with red blood cells in vitro. I. Evidence for carrier 3613 mediated transport. Chem Biol Interactions, 42: 165-177. 3614 3615 Williams PAM & Baran EJ (2008) Vanadium detoxification: On the interaction of 3616 oxovanadium(IV) and other vanadium species with 2,3-dimercapto-1- 3617 propanesulfonate. J Inorg Biochem, 102: 1195-1198. 3618 3619 Willig RP, Drohn W & Stegner H (1984) [Mercury poisoning: successful treatment with 3620 DMPS] (German). Monatschr Kinderheilkd, 132: 701. Cited in: Ruprecht J (1997) th 3621 Scientific monograph Dimaval® (DMPS), 6 edition. Berlin, Heyl Company. 3622 3623 Xu ZF & Jones MM (1988) Comparative mobilization of lead by chelating agents. 3624 Toxicology, 53: 277-288. 3625 3626 Zalups RK, Parks LD, Cannon VT & Barfuss DW (1998) Mechanisms of action of 3627 2,3-dimercaptopropane-1-sulfonate and the transport, disposition, and toxicity of 3628 inorganic mercury in isolated perfused segments of rabbit proximal tubules. Mol 3629 Pharmacol, 54: 353-363. 3630 3631 Zalups RK & Bridges CC (2009) MRP2 involvement in renal proximal tubular 3632 elimination of methylmercury mediated by DMPS or DMSA. Toxicol Appl Pharmacol, 3633 235: 10-17. 3634 3635 Zhang J (1984) Clinical observations in ethyl mercury chloride poisoning. Am J Ind 3636 Med, 5: 251-258. 3637 3638 Zheng W, Maiorino RM, Brendel K & Aposhian HV (1990) Determination and 3639 metabolism of dithiol chelating agents. VII. Biliary excretion of dithiols and their 3640 interactions with cadmium and metallothionein. Fundam Appl Toxicol, 14: 598-607. 3641 3642 3643 15. Author names, address

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3644 3645 Initial draft by FW Jekat & FH Kemper 3646 3647 Updated by Nicola Bates MSc, MA: Guy's & St Thomas' Unit, London, UK 3648 3649 3650 16. Additional information 3651 3652 16.1 Description of search strategy 3653 3654 Table 2: Results of EMBASE search, 5 April 2009. 3655 Number Keywords Results 1 Unithiol [any field] 569 2 DMPS [any field] 446 3 (dimercaptopropanol OR dimercaptopropane AND sulfonate) 100 [any field] 4 poisoning OR toxicity OR overdose [any field] 186824 5 1 and 4 277 6 2 and 4 145 7 3 and 4 51 8 5 and 6 and 7 remove duplicates 295 3656 3657 Table 3: Results of Pubmed search, using the EMBASE interface, 5 April 2009. 3658 Number Keywords Results 1 Unithiol [any field] 541 2 DMPS [any field] 437 3 dimercaptopropanol OR dimercaptopropane AND sulfonate 103 [any field] 4 poisoning OR toxicity OR overdose [any field] 265512 5 1 and 4 208 6 2 and 4 158 7 3 and 4 62 8 5 and 6 and 7 remove duplicates 276 3659 3660 Table 4: Results of Cochrane Library, 5 April 2009 3661 Number Keywords Results 1 Unithiol [MeSH term] in Central Register of Controlled Trials 11 2 Unithiol [MeSH term] in Cochrane reviews 0 3 Unithiol [MeSH term] in other reviews 0 4 Unithiol [MeSH term] in economic evaluations 1 3662 3663 16.2 Description of published evidence 3664 3665 Table 5: Summary of evidence of use of unithiol in metal and metalloid poisoning.

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3666 Metal data Case reports Animal studies Antimony No data. Used with apparent benefit Unithiol has been shown to in a small number of increase survival (Basinger paediatric trivalent antimony & Jones, 1981a) and compounds (Iffland & reduce the LD50 (Chih- Bösche, 1987; Kemper et Chang, 1958) in antimony- al., 1989; Jekat & Kemper, poisoned experimental 1990). There is no animals. information on its use in pentavalent antimony compounds. Arsenic One randomized, 5 reports of acute arsenic Unithiol increases survival single blinded, poisoning involving 7 adult (Tadlock & Aposhian, 1980; placebo-controlled patients treated with unithiol Aposhian et al., 1982; Inns study of 21 patients as the only antidote (Moore et al., 1990), increases the with chronic et al., 1994; Kruszewska et LD50 (Aposhian et al., 1981), arsenicosis. Unithiol al., 1996; Horn et al., 2002; increases urinary (Maiorino resulted in significant Adam et al., 2003; Heinrich- & Aposhian, 1985; Flora et clinical improvement. Ramm et al. 2003). All al., 1995a) and faecal Cessation of patients showed increased (Maehashi & Murata, 1986; exposure and urinary excretion and Reichl et al., 1995) placebo also reduced decreased plasma elimination, reduces tissue clinical scores but concentrations of arsenic. concentrations (Kreppel et these were All patients recovered. al., 1989; Kreppel et al., significantly lower for 1990; Schäfer et al., 1991) unithiol-treated 1 report where dimercaprol and reduces the severity of subjects. In the was used followed by toxicity (Kreppel et al., 1989, placebo group succimer and unithiol Inns & Rice, 1993; Flora et improvement was together. Patient showed al., 1995a; Flora et al., attributed to clinical improvement but 2005) in arsenic poisoned cessation of contribution of unithiol experimental animals. exposure and difficult to determine hospitalisation. (Vantroyen et al., 2004) There were significant increases 1 report of chronic arsenic in urinary arsenic poisoning. Administration of excretion with unithiol unithiol resulted in increased treatment, compared urinary excretion of arsenic to no increase in the and clinical improvement placebo group. No (Wax & Thornton, 2000). adverse effects were reported (Guha Mazumder et al., 2001).

Beryllium No data. No data. Unithiol increases beryllium excretion and reduces beryllium-induced toxic effects in experimental animals (Mathur et al., 1994; Flora et al., 1995b; Johri et al., 2002; Johri et al., 2004). Bismuth No data. 3 cases of acute bismuth Unithiol increases survival poisoning, involving a 13- (Basinger et al., 1983) and year-old (Bogle et al., 2000) reduces tissue and 2 adults (Stevens et al., concentrations (Slikkerveer 1995; Dargan et al., 2001; et al., 1992; Jones et al.,

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Metal Clinical trial data Case reports Animal studies Ovaska et al., 2008). All 1996) in bismuth-poisoned showed improved clearance experimental animals. of bismuth, though one patient was also haemodialysed (Stevens et al., 1995).

2 cases of chronic bismuth poisoning. In 1 case unithiol resulted in improvement of neurological toxicity (Playford et al., 1990) and in the other the patient deteriorated and unithiol was stopped (Teepker et al., 2002). Cadmium No data. 1 case report. Unithiol in a Unithiol increases the LD50 patient with chronic (Pethran et al., 1990), occupational exposure increases survival (Aposhian increased urinary cadmium (1982; Andersen & Nielsen, concentrations. No further 1988; Basinger et al., 1988; information available Srivastava et al., 1996), (Daunderer, 1995). reduces tissue concentrations (Planas- Bohne & Lehman, 1983; Eybl et al., 1984; Srivastava et al., 1996) in cadmium poisoned experimental animals. However, in some studies there was no effect on survival (Eybl et al., 1984), excretion (Eybl et al., 1984, Rau et al., 1987; Zheng et al., 1990) or tissue distribution of cadmium (Cherian, 1980) and an increase in tissue concentrations was reported in some studies (Shinobu et al., 1983; Basinger et al., 1988). Although unithiol may be effective, other metal- binding agents appear to be more efficacious (Eybl et al., 1984; Eybl et al., 1985; Andersen & Nielsen, 1988; Srivastava et al., 1996). Chromium No data. 2 case reports in adults; the Apparent benefit effect of unithiol is unclear demonstrated, but data are in both. 1 patient recovered limited. Unithiol reduces (Donner et al., 1986); the chromate-induced other also received cytotoxicity (in some haemodialysis and circumstances) (Susa et al, haemofiltration but died 48 1994), reduces lethality and hours after admission increases renal excretion of (Pudill et al., 1989). (Susa et al, 1994) in chromium-poisoned experimental animals.

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Metal Clinical trial data Case reports Animal studies Cobalt No data. 2 paediatric cases of acute Apparent benefit exposure. Unithiol was demonstrated, but data are used after penicillamine and limited. Unithiol has been was associated within shown to reduce the increased urinary cobalt lethality of cobalt (Cherkes concentrations (Müller et al., & Braver-Chernobulskaya, 1989). 1958; Eybl et al., 1985) but to increase cobalt concentrations in some tissues (Eybl et al., 1985). Copper No data. One paediatric case of Apparent benefit acute ingestion; unithiol was demonstrated, but data are associated with increased limited. Unithiol increased urinary copper excretion the LD50 (Pethran et al., (Donner et al., 1986). 1990) and reduced toxicity (Mitchell et al., 1982) in In an adult case there was copper-poisoned no measurement of copper experimental animals. excretion; the patient survived (Sinković et al., 2008). Gold Wilson’s disease Wilson’s disease Unithiol increases survival In a randomised trial In an unspecified number of (Basinger et al., 1985), of 28 patients patients copper excretion reduces renal comparing unithiol with unithiol was concentrations (Gabard, and captopril, unithiol comparable to that of 1980; Kojima et al., 1991; had a more potent penillicamine. Unithiol was Takahashi et al., 1994), anticopper effect. 1 discontinued in 2 patients increases urinary excretion patient on unithiol due to adverse effects (Kojima et al., 1991) and developed a (Walshe, 1985) reduces renal toxicity transient, adverse (Kojima et al., 1991; effect (Wang et al., Copper poisoning Takahashi et al., 1994) in 2003). 1 case of iatrogenic gold-poisoned in poisoning where unithiol experimental animals was thought to be effective in removing gold although no details are given (Ashton et al., 1992a). Lead One controlled study In 2 adult patients with Unithiol increases survival of 60 males with chronic lead exposure (Llobet et al., 1990), chronic lead toxicity. unithiol decreased blood increases excretion Unithiol-treated lead concentrations and (Hofmann & Segewitz, patients had increased urinary lead 1975; Llobet et al., 1990), increased elimination excretion (Donner et al., reduces tissue and biochemical and 1987; Hruby & Donner, concentrations (Twarog & clinical improvement, 1987; Autenrieth et al., Cherian, 1983; Twarog & and were discharged 1998). Cherian, 1984; Xu & Jones, 6 weeks earlier than 1988; Llobet et al., 1990), controls Succimer is more commonly except in the brain (Sharma (Anatovskaya, 1962). used in the management of et al., 1987; Aposhian et al., lead poisoning. 1996) and reduces toxicity 12 children (aged 31 (Twarog & Cherian, 1983; to 69 months) with Sharma et al., 1987; chronic lead toxicity Tandon et al., 1994) in lead- received one of two poisoned experimental dose regimens of animals. unithiol. All had reduced blood lead concentrations and

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Metal Clinical trial data Case reports Animal studies increased urinary excretion (Chisolm & Thomas, 1985).

Mercury In patients with Inorganic mercury Unithiol increases excretion chronic exposure 6 acute cases reported; 3 (Gabard, 1976a; Gabard, penicillamine (12 patients initially received 1976b; Wannag & Aaseth, patients), N-acetyl-DL- dimercaprol (Nadig et al., 1980; Planas-Bohne, 1981; penicillamine (17), 1985; Ashton & House, Aaseth et al., 1982; Buchet unithiol (10) and a 1989; Toet et al., 1994) and & Lauwerys, 1989), thiolated resin (8) 4 developed renal failure decreases tissue were compared. The (Nadig et al., 1985; Ashton concentrations (Gabard, study was not & House, 1989; Toet et al., 1976a; Gabard, 1976b Cikrt clinically controlled. 1994; Dargan et al., 2003a). & Lenger, 1980; Wannag & Although all agents Unithiol was shown to Aaseth, 1980; Aaseth et al., reduced blood reduce the mercury 1982; Aaseth, 1983; Kachru mercury elimination half-life in two & Tandon, 1986) and concentrations and cases (Toet et al., 1994; reduces toxicity (Planas- unithiol was the most Dargan et al., 2003a). All Bohne, 1977; Jones et al., effective there was no patients recovered. 1980; Nielson & Andersen, immediate clinical 1991) in mercury-poisoned improvement, In 2 chronic exposure cases experimental animals. presumably because unithiol decreased the the duration of therapy elimination half-life of was too short and mercury (Campbell et al., therapy was started 1986). months after exposure (Clarkson et al., Organic mercury 1981). 3 acute cases in adults. In 1 case unithiol was 27 patients were determined to be relatively treated with unithiol ineffective possibly due to and/or succimer. All coadministration of a had some relief (19 copper and zinc became supplement (Lund et al., asymptomatic). Most 1984). In another case had increased where unithiol was mercury excretion. alternated with Unithiol was found to penicillamine, the unithiol be more effective reduced mercury protein (data not provided binding (Köppel et al., and the two antidotes 1982). The 3rd patient was were used also treated with succimer; interchangeably in neither significantly some patients) increased mercury (Zhang, 1984). clearance (Pfab et al., 1996). In a study of 95 patients with chronic Metallic mercury exposure (vapour, 2 cases of aspiration; both inorganic mercury and patients remained well but in methyl mercury) 1 the mercury blood and unithiol increased urine concentrations urinary mercury remained high (Batora et al., excretion in some but 2001) and in the other the others (number not blood concentration specified) showed no decreased rapidly (Kummer increase in mercury & Michot, 1984). excretion. In more

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Metal Clinical trial data Case reports Animal studies than two-thirds there In 3 children (<7 years) was improvement in unithiol was associated with subjective complaints improved clinical signs and and objective enhanced mercury excretion neurological (von Mühlendahl, 1990). In parameters. The 14 a 14-year-old with acrodynia day regimen was too there was slow recovery short to have a with unithiol (Böckers et al., permanent effect on 1983). mercury concentrations (Böse- Dermal mercury O’Reilly et al., 2003). In 2 cases of chronic Study limitations: exposure, unithiol absence of a control increased urinary mercury group, lack of details concentrations (Böckers et of patients (age, al., 1985l Pelclová et al., weight) and continued 2001). exposure to mercury during therapy. Parenteral mercury 4 cases of mercury injection In 8 patients with (Ashton et al., 1992b; chronic exposure from Torres-Alanís et al., 1997; facial mercurous Batora et al., 2000; Eyer et chloride cream there al., 2006). Although unithiol was a significant increases urinary mercury increase in urinary concentrations, the effect mercury may be small (Eyer et al., concentrations after 2006) and there may be no 24 hours of unithiol. change in radiographic One symptomatic deposits of mercury patient recovered and (Torres-Alanís et al., 1997) the other had or clinical signs. persistent tremor (Garza-Ocañas et al., 1997). Nickel No data. No data. Unithiol increases survival (Basinger et al., 1980), increases excretion (Sharma et al., 1987) and reduces nickel-induced toxic effects (Sharma et al., 1987; Tandon et al., 1996). Palladium No data. No data. No benefit demonstrated, but data are limited. Unithiol did not influence toxicity or reduce lethality in palladium-poisoned animals (Mráz et al., 1985). Platinum No data. No data. No benefit demonstrated, but data are limited. A single dose of unithiol had no significant effect on renal platinum concentrations. After 4 treatments there was significant increase in urinary excretion of platinum, but this was low (Planas-Bohne et al., 1982). Polonium No data. Children (number not Not recommended.

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Metal Clinical trial data Case reports Animal studies known, but <10) treated Although unithiol can with unithiol after remove polonium-210 from contamination with most tissues (Aposhian et poloniumn-210 remained al., 1987) it results in well over the 46 month concentration of polonium in period of monitoring with the kidneys (Volf, 1973; only impairment of protein Rencová et al., 1993; Volf formation in the liver et al., 1995). (Shantyr et al., 1969). Selenium No data. No data. No benefit demonstrated, but data are limited. Unithiol had no effect in selenium- poisoned animals; the concentration of selenium in urine and faeces was unchanged (Paul et al., 1989). Silver No data. In 2 cases of chronic silver Unithiol increased the LD50 toxicity unithiol increased of silver chloride in mice urinary silver excretion but (Pethran et al., 1990), the quantity excreted was prevented the development low (Aaseth et al., 1986; of toxic pulmonary oedema Kemper et al., 1989; Jekat and death in dogs and Kemper, 1990). (Romanov, 1967) and in vitro completely reversed silver inhibition of Na,K- ATPase (Hussain et al., 1994). Strontium No data. No data No benefit demonstrated, but data are limited. Unithiol did not affect survival rate in strontium-poisoned experimental animals (Domingo et al., 1990; Pethran et al., 1990). Thallium No data. No data. Unithiol is ineffective in thallium-poisoned experimental animals (Pethran et al., 1990; Mulkey & Oehme, 2000). Tin No data. In 1 case unithiol increased Apparent benefit urinary tin concentrations demonstrated with reduced and clinical signs improved tin-induced lesions in rats (Hruschka, 1990). (Merkord et al., 2000), but data are limited. Vanadium No data. No data. Unithiol had no effect on lethality in vanadium- poisoned mice (Jones & Basinger, 1983) and no significant effect on the death rate, body weight reductions, or reduction in weights of legs and toes in chick eggs incubated with vanadium (Hamada, 1994). An in vitro study has shown that oxovanadium forms a complex with unithiol (Williams & Baran, 2008).

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Metal Clinical trial data Case reports Animal studies The significance of this in vivo remains to be elucidated. Zinc No data. No data. Unithiol increases excretion (Domingo et al., 1988) and reduced lethality in zinc- poisoned experimental animals (Basinger & Jones, 1981b), but more effective antidotes are available (Basinger & Jones, 1981b; Domingo et al., 1988; Llobet et al., 1988). 3667 3668 Abbreviations 3669 3670 AAS Atomic Absorption Spectroscopy 3671 ABCC2 ATP-binding cassette, sub-family C 3672 AES Atomic Emission Spectroscopy 3673 ALAD δ-aminolevulinate dehydratase 3674 BAL 2,3-dimercaptopropanol; British Anti-Lewisite; dimercaprol (rINN) 3675 BAPSA 2,3-bis-(acetylthio)-propanesulphonamide 3676 CAS Chemical Abstracts Service 3677 CDTA cyclohexanediamintetraacetic acid 3678 DDC sodium diethyldithiocarbamate 3679 DMPA N-(2,3-dimercaptopropyl) phthalamidic acid 3680 DMSA Dimercaptosuccinic acid; succimer (rINN) 3681 DTPA Diethylenetriaminepentaacetic acid; pentetic acid (rINN) 3682 EDTA Ethylenediaminetetraacetic acid; sodium calcium edetate (rINN) 3683 g gram 3684 HOEtTTC N,N’-di-(2-hydroxyethyl)-ethylenediamine-N’N’-biscarbodithioate 3685 HPLC High Performance Liquid Chromatography 3686 ICP Inductively Coupled Plasma 3687 i.m. intramuscular 3688 i.p. intraperitoneal 3689 IPCS International Programme on Chemical Safety 3690 IR infra-red 3691 i.v. intravenous 3 3692 k kilo (10 ) 3693 L litre 3694 LD Lethal Dose (subscript indicates percent mortality) -3 3695 m milli (10 ) -6 3696 µ micro (10 ) 3697 µCi microCurie 3698 min minute 3699 MRP2 multidrug resistance protein 2 3700 NOEL no observed effect level 3701 OAT1 organic anion transporter 1 3702 OAT3 organic anion transporter 3 3703 PAH p-aminohippurate -6 3704 ppm parts per million (10 )

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3705 rINN recognised international non-proprietary name 3706 3707

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