Nephrotoxicity of the flame retardant, tris(2,3-dibromopropyl) phosphate, and its metabolites

TOXICOLOGY

AND

APPLIED

PHARMACOLOGY

62, 179- 182 (1982)

SHORT COMMUNICATION Nephrotoxicity

of the Flame Retardant, Tris(2,3-dibromopropyl) Phosphate, and Its Metabolites

Nephrotoxicity of the Flame Retardant, Tris(2,3-dibromopropyl) Phosphate, and Its Metabolites. ELLIOT, W. C., LYNN, R. K., HOUGHTON, D. C., KENNISH, J. M., AND BENNEXT, W. M. (1982). Toxicol. Appl. Pharmacol. 62, 179-182. A single ip injection of the carcinogenic flame retardant, tris(2,3-dibromopropyl) phosphate (TRIS-BP), when administered to male Sprague-Dawley rats, caused polyuric acute renal failure with tubular necrosis involving the late proximal tubule. Glomerular filtration rate and in vitro transport of the organic acid, para-aminohippurate and the organic base, N-[ ‘%]methylnicotinamide, were depressed. An approximately equimolar dose of the TRIES-BP metabolite, bis(2,3-dibromopropyl) phosphate (BIS-BP), caused significantly more severe renal failure. In contrast, the metabolite 2,3dibromopropanol (BP) was nonnephrotoxic. These data suggest that TRIS-BP nephrotoxicity is mediated via its metabolite BIS-BP.

Tris(2,3-dibromopropyl) phosphate (TRISBP) is a flame retardant that was used extensively by the apparel industry in the early and mid-1970s (St. John et al., 1976). It is absorbed through the skin and excreted into the urine of children (Blum et al.., 1978). In 1977, its production was discontinued because of evidence that it was both mutagenic to bacteria (Prival et al., 1977) and carcinogenic to rodents fed chow containing TRIS-BP for 2 years (NCI, 1978). Recently, Soderlund et al. ( 1980) reported that a single high dose of TRIS-BP caused reversible acute renal failure with tubular necrosis and depression of glomerular filtration rate (GFR). Furthermore, pretreatment with CoC& (an inhibitor of the cytochrome P-450 mixed-function oxidase system) appeared to diminish the nephrotoxicity of TRIS-BP. They therefore speculated that a metabolite of TRIS-BP might mediate its nephrotoxic properties. To date, two metabolites of TRIS-BP have been identified: bis( 2,3-dibromopropyl) phosphate (BIS-BP) and 2,3-dibromopropanol (BP) (Lynn et al., 1980). The three moieties differ from one another in the number of 2,3-dibromopropanol groups: 3 (TRISBP), 2 (BIS-BP), and 1 (BP). Both BIS-BP and BP appear in the serum within minutes

of iv administration of TRIS-BP. To determine if the metabolites of TRIS-BP are nephrotoxic, we administered approximately equimolar doses of each to rats. METHODS TRIS-BP, 154 mg/kg, was administered as a single ip dose to male Sprague-Dawley rats (Simonsen Labs, Gilroy, Calif.) weighing 275-325 g. Approximately equimolar doses of BIS-BP (120 mg/kg) and BP (61 mg/kg) were administered to littermate controls. Fortyeight hours later, animals were sacrificed via ether anesthesia and exsanguination. Serum creatinine (Scr) was determined as an index of glomerular filtration rate (GFR). As an index of tubular function, we determined the activity of the organic acid and organic base transport systems. Renal cortical slices were incubated for 90 min in Cross and Taggart medium in the presence of the organic acid, para-aminohippurate (PAH), and the organic base, N-[ “C]methylnicotinamide (NMN). Slice uptake was expressed as slice-to-medium ratio (S/ M). Details of the technique are published elsewhere (Bennett et al., 1978). The unsliced kidney was processed by standard histologic methods (Houghton et al., 1976).

RESULTS TRIS-BP caused polyuric acute renal failure with elevation of Scr (Table 1). PAH uptake was moderately depressed. NMN 179

0041-008X/82/010179-04$02.00/0 Copyright 0 1982 by Academic Press, Inc. All rights of reproduction in any form reserved.

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1

SUMMARY OF NEPHROTOXIC EFFECTS OF T~rs(2,3-DIBROMOPROPYL) PHOSPHATE BROMOPROPYL)PHOSPHATE(BIS-BP),AND~,~-DIBROMOPROPANOL(BP)INJECTEDINAPPROXIMATELYEQUIMOLARAMOUNTS' 48 hr BEFORESACRIFICE BUN’ bg/dl)

Serb bg/dl)

Group Control TRIS-BP BIS-BP BP

0.79 3.32 8.40 0.77

f 0.1 + 1.6’ + l.Og + 0.1

16+ 2 99 + 56/ 241 f 22g 15* 2

’ Tris(2,3-dibromopropyl) phosphate: 154 mg/kg; bis(2,3-dibromopropyl) opropanol: 6 1.6 mg/kg. ’ Serum creatinine. ’ Blood urea nitrogen. d In vitro slice to medium ratio of para-aminohippurate. ’ In vitro slice to medium ratio of N-1“‘Clmethylnicotinamide. _ ‘p < 0.05 compared to control and his groups. pp < 0.05 compared to control and tris groups.

uptake was minimally (but significantly) depressed. Histologic examination revealed acute tubular necrosis of the inner cortex affecting the convoluted and straight portions of the proximal tubule. BIS-BP, in equimolar dose, caused significantly higher Scr and greater depression of PAH and NMN transport than TRIS-BP. Histologic examination revealed more widespread involvement with necrosis extending into descending loops of Henle (Fig. 1). In contrast, BP caused neither functional nor histologic abnormalities. In addition, the vehicle caused no evidence of toxicity (data not shown). DISCUSSION These data show that BIS-BP, a metabolite of TRIS-BP in plasma and urine, caused acute renal failure. This result was not unexpected since Soderlund et al. ( 1980) reported that inhibition of the cytochrome P-450 drug metabolizing system reduced TRIS-BP nephrotoxicity and speculated that a TRIS-BP metabolite was the cause of renal failure. The characteristics of TRIS-BP metabolism also suggest that a metabolite

SM/PAHd 11.7 4.1 1.5 11.0

+ iz + +

(TRIS-BP),

Bls(2,3-DI-

SM/NMN’ 2.0 1.4’ 0.6$ 1.8

phosphate:

4.6 4.0 2.1 4.5 120 mg/kg;

f f f f

0.4 0.4’ 1.1s 0.4 2,3-dibrom-

might be responsible for its biologic actions: the serum half-life of TRIS-BP is less than 2 min, while both BIS-BP and BP appear rapidly and both have long serum half-lives (Kennish et al., 1979). Furthermore, 5 days after injection of 14C-labeled TRIS-BP, high concentrations of a non-TRIS-BP moiety were found in the kidney (Kennish et al., 1979). Our data thus suggest that BIS-BP may mediate TRIS-BP-associated acute renal failure. However, since BIS-BP accounted for only 7.8% of urinary label after administration of TRIS-BP (Lynn et al., 1980), an as yet unidentified metabolite(s) of BIS-BP could also be responsible for nephrotoxicity. TRIS-BP is better known for its mutagenic and carcinogenic properties than as an acute nephrotoxin (Prival et al., 1977; NCI, 1978). An intriguing possibility is that BISBP is the cause of all three effects. In support of this hypothesis, Soderlund et al. (1979) reported that TRIS-BP bacterial mutagenicity was increased in the presence of cytochrome P-450-stimulated microsomes, suggesting the conversion of TRIS-BP to a more toxic metabolite. Furthermore, BISBP itself has been reported to be a bacterial

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FIG. 1. Renal medulla of a BIS-BP-treated animal. The epithelial cells of many loops of Henle are necrotic. In other areas, continuity of some of these zones of destruction with similarly affected straight segments of proximal tubules indicate that most, if not all, are descending limbs of the loops. The bar in the lower lefthand corner represents 40 pM (H&E, 255X).

mutagen (Nakamura et al., 1979). Thus, the possibility that BIS-BP is the cause of nephrotoxicity, mutagenicity, and carcinogenicity is an attractive, but unproven, hypothesis. Several other halogenated hydrocarbons are nephrotoxic (Kluwe and Hook, 1980). Like TRIS-BP, both methoxyflurane (2,2dichloro- 1,l -difluoroethylmetbyl ether) and chloroform appear to produce injury via toxic metabolites. Furthermore, methoxyflurane nephrotoxicity was increased by pretreatment with the cytochrome P-450 inducer, phenobarbital, and reduced by SKF 525-A pretreatment (an inhibitor of drug metabolism) (Cousins et al., 1974). Chloroform-mediated nephrotoxicity also has

been altered by preexposure enzyme induction (Kluwe and Hook, 1980). TRIS-BP may be another example of a nephrotoxic agent activated by conversion to a toxic metabolite. Whether BIS-BP represents the toxic agent awaits further identification of metabolites and administration of inhibitors and inducers of drug metabolism before treatment with BIS-BP. ACKNOWLEDGMENTS The authors gratefully acknowledge the technical assistance of J. DeFehr, K. Kuettner, and J. Kimsey and the secretarial expertise of J. Paquet. This work was supported, in part, by NIEHS Contract NOl-ES-‘I2126.

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REFERENCES BENNETT, W. M., PLAMP, C. E., GILBERT, D. N., AND PORTER, G. A. (1978). Kinetics of gentamicin uptake in rat cortical slices and the effect of aminoglycoside pretreatment on the transport of para-aminohippurate (PAH) and N-methyl-nicotinamide (NMN). In Nephrotoxicity: Interactions of Drugs with Membranes Systems Mitochondria Lysosomes (J. P. Fillastre, ed.), pp. 143-156. Masson, New York. BLUM, A., GOLD, M. D., AMES, B. N., KENYON, C., JONES, F. R., HETT, E. A., DOUGHERTY, R. C., HORNING, E. C., DZIDIC, I., CARROLL, D. I., STILLWELL, R. N., AND THENOT, J-P. (1978). Children absorb Tris-BP flame retardant from sleepware: Urine contains the mutagenic metabolite, 2,3-dibromopropanol. Science 201, 1020-1023. COUSINS, M. J., MAZZE, R. I., KOSEK, J. C., HITT, B. A., AND LOVE, F. V. (1974). The etiology of methoxytlurane nephrotoxicity. J. Pharmacol. Exp. Ther. 190, 530-541.

HOUGHTON, D. C., HARTNETT, M. N., CAMPBELLBOSWELL, M., PORTER, G. A., AND BENNE-I-~, W. M. (1976). A light and microscopic analysis of gentamicin nephrotoxicity in rats. Amer. J. Pathol. 82, 589-599.

KENNISH, J. M., WONG, K., GERBER, N., AND LYNN, R. K. (1979). Pharmacokinetics of the flame retardant tris (2,3-dibromopropyl) phosphate in the rat: Plasma concentrations, excretion and tissue distribution. Pharmacologist 21, 216. KLUWE, W. M., AND HOOK, J. B. (1980). Metabolic activation of nephrotoxic haloalkanes. Fed. Proc. 39, 3129-3133. LYNN, R. K., WONG, K., DICKINSON, R. G., GERBER, N., AND KENNISH, J. M. (1980). Diester metabolites of the flame retardant chemicals, tris (1,3-dichloro-2propyl) and tris (2,3-dibromopropyl) phosphate in the rat: Identification and quantification. Res. Comm. Chem. Path. Pharmacol. 28, 351-360. NAKAMURA, A., TATENO, N., KOJIMA, S., KANIWA, M-A., AND KAWAMURA, T. (1979). The mutagenicity of halogenated alkanols and their phosphoric acid esters for Salmonella typhimurium. Muta. Res. 66, 373-380.

National Cancer Institute, Carcinogenesis Testing Program. (1978). Bioassay of Tris(2,3-dibromopropyl) Phosphate for Possible Carcinogenicity. National Cancer Institute Carcinogenesis Technical Report Series No. 76. National Cancer Institute, Washington D.C. PRIVAL, M. J., MCCOY, E. C., GUTTER, B., AND ROSENKRANZ, H. S. (1977). Tris (2,3-dibromopropyl) phosphate: Mutagenicity of a widely used flame retardant. Science 195,16-78. MDERLUND, E. J., DYBING, E., AND NELSON, S. D. (1980). Nephrotoxicity and hepatotoxicity of Tris (2,3-dibromopropyl) phosphate in the rat. Toxicol. Appl. Pharmacol. 56, 17 1 - 18 1. S~DERLUND, E. J., NELSON, S. D., AND DYBING, E. (1979). Mutagenic activation of tris(2,3-dibromopropyl) phosphate: The role of microsomal oxidative metabolism. Acta Pharmacol. Toxicol. 45, 112- 12 1. ST. JOHN, JR., L. E., ELDEFRAWI, M. E., AND LISK, D. J. (1976). Studies of possible absorption of a flame retardant from treated fabrics worn by rats and humans. Bull. Environ. Contam. Toxicol. 15, 1?2-197.

W. CLAYTON

ELLIOTT

Department of Medicine Division of Nephrology ROBERT

K.

LYNN

Department of Pharmacology DONALD

C. HOUGHTON

Department of Pathology JOHN M. KENNISH Department of Pharmacology WILLIAM

M.

Department of Medicine, Division of Nephrology Oregon Health Sciences University 3181 Southwest Sam Jackson Park Road Portland, Oregon 97201 Received June 15. 1981

BENNETT