title: Toxicology and Carcinogenesis Studies of Hydrochlorothiazide (CAS No. 58-93-5) in F344/N Rats and B6C3F1 Mice (Feed Studies).
abstract: Hydrochlorothiazide is a diuretic active at the distal convoluted tubule and collecting duct. Toxicology and carcinogenesis studies were conducted by feeding diets containing hydrochlorothiazide (USP grade, greater than 98% pure) to groups of F344/N rats and B6C3F1 mice of each sex for 15 days, 13 weeks, 1 year, or 2 years. Additional studies were performed to evaluate teratologic effects in CD(R). rats and CD(R).-1 mice. Genetic toxicology studies were performed with Salmonella, Chinese hamster ovary (CHO) cells, mouse lymphoma cells, and Drosophila. Fifteen-Day and Thirteen-Week Studies: All rats and mice lived to the end of the 15-day studies (dietary concentrations of 0 and 3,125-50,000 ppm). The final mean body weights of all dosed rat groups were 5%-11% lower than those of controls. The final mean body weights of the groups of male mice that received 6,250-50,000 ppm were 10%-14% lower than that of controls. The final mean body weights of dosed and control female mice were similar. Calculi were seen in the urinary bladder of 2/5 male and 2/5 female mice at 50,000 ppm and in 1/5 male and 1/5 female mice at 25,000 ppm. All rats lived to the end of the first 13-week studies (dietary concentrations of 0 and 3,125-50,000 ppm). Final body weights of dosed rats were 7%-16% lower than those of controls. Mineralization in the kidney was observed in all dosed rats and because of this, additional 13-week studies in rats were conducted at lower dietary concentrations. All rats lived to the end of the second 13-week studies (dietary concentrations of 0 and 250-4,000 ppm). The final mean body weights of all dosed rat groups were 5%-10% lower than those of controls. Renal mineralization was dose related and judged to be minimal to mild at the lowest dose. In the 13-week studies in mice, 7/10 males and 1/10 females that received 50,000 ppm hydrochlorothiazide died. The final mean body weights of mice that received 50,000 ppm were 11% lower than those of controls for males and females. Calculi were seen in the urinary bladder of mice that received hydrochlorothiazide at 12,500 ppm and above. Nephrosis occurred with dose-related incidences in mice receiving 12,500 ppm and above. Based on these results, 2-year studies were conducted by feeding diets containing 0, 250, 500, or 2,000 ppm hydrochlorothiazide to groups of 50 male and 50 female rats for 105-106 weeks. Diets containing 0, 2,500, or 5,000 ppm hydrochlorothiazide were fed to groups of 50 male and 50 female mice for 103-104 weeks. Ten additional rats per sex and dose group were placed on study and killed at 1 year for blood-clotting studies and histopathologic examination. Effects in the One-Year Studies: One of 10 female rats in the 1-year study group that received 2,000 ppm died with internal hemorrhage. In addition, evidence of hemorrhage was found in 11 of the 16 dosed female rats that died during the first year of the 2-year study. Hematologic analyses revealed no compound-related effects; however, activated partial thromboplastin times (APTTs) were highly variable and were lengthened in some dosed male rats. No effects on APTTs were seen for females, and no effects on prothrombin times or on the fibrinogen content of plasma were observed for dosed male or female rats. Nephropathy occurred in dosed and control rats, and the severity was judged to be greater in dosed male and high dose female rats. Increased incidences of mild focal renal mineralization were also seen in mid and high dose male rats and dosed female rats. Body Weight and Survival in the Two-Year Studies: Mean body weights of dosed rats were 8%-25% lower than those of controls. Mean body weights of dosed and control mice were similar throughout the studies. No significant differences in survival were observed between rats or mice of either sex (rats-- male: control, 18/50; low dose, 16/50; mid dose, 9/50; high dose, 11/50; female: 31/50; 26/50; 30/50; 27/50; mice--male: control, 43/50; low dose, 42/50; high dose, 43/50; female: 38/50; 40/50; 35/50). Survival of all groups of male rats was low because a lar female: 38/50; 40/50; 35/50). Survival of all groups of male rats was low because a large number of animals were killed in a moribund condition late in the study. The average daily feed consumption by dosed rats was 89&percnt;-94&percnt; that by controls. The average amount of hydrochlorothiazide consumed per day was approximately 11, 23, or 89 mg/kg for low, mid, or high dose rats. The average daily feed consumption by dosed mice was 100&percnt;-105&percnt; that by controls. The average amount of hydrochlorothiazide consumed per day was approximately 280 or 575 mg/kg for low dose or high dose mice. Nonneoplastic and Neoplastic Effects in the Two-Year Studies: Nephropathy occurred in nearly all male and female rats, but the severity of this disease was greater in dosed rats, as evidenced by increases in renal cysts and epithelial hyperplasia of the renal pelvis in dosed rats shown in the following table (see page 4 of the Technical Report). Mineralization was observed at increased incidences in dosed male and dosed female rats. Changes associated with or secondary to renal injury were increased in dosed rats. These lesions included parathyroid hyperplasia, fibrous osteodystrophy of bone, and mineralization of multiple organs. Adenomas or carcinomas (combined) of the Zymbal gland in male rats occurred in 1/50 control, 1/49 low dose, 2/50 mid dose, and 4/50 high dose animals. The historical incidence of Zymbal gland neoplasms in untreated F344/N rats is 19/1,936 (1.0&percnt;), and the highest observed control group incidence is 4/50. This marginal increase was not considered to be chemically related. The incidences of fibroadenomas of the mammary gland were decreased in dosed female rats (30/50; 12/50; 11/49; 5/50). The incidence of hepatocellular neoplasms was increased in high dose male mice (adenomas or carcinomas, combined: control, 7/48; low dose, 10/49; high dose, 21/50). The historical incidence of hepatocellular adenomas or carcinomas (combined) is 609/2,032 (30&percnt;) in untreated controls. Teratology: Hydrochlorothiazide produced no teratologic effects in the offspring of CD&reg;. rats or CD&reg;.-1 mice after gavage administration to pregnant females on day 6 through day 15 of gestation. Genetic Toxicology: In the absence of exogenous metabolic activation, hydrochlorothiazide produced an equivocal increase in revertant colonies in Salmonella typhimurium strain TA98; no increase was observed in strains TA100, TA1535, or TA1537 with or without activation. Hydrochlorothiazide induced an increase in trifluorothymidine (Tft)-resistant cells in a mouse lymphoma L5178Y/TK+/- assay without exogenous metabolic activation; this assay was not performed with activation. In cultured CHO cells, hydrochlorothiazide induced sister chromatid exchanges (SCEs) in the presence and absence of exogenous metabolic activation but did not induce chromosomal aberrations. Hydrochlorothiazide did not increase the frequency of sex-linked recessive lethal mutations when administered by feeding or injection to adult male Drosophila melanogaster. Audit: The data, documents, and pathology materials from the 2-year studies of hydrochlorothiazide have been audited. The audit findings show that the conduct of the studies is documented adequately and support the data and results given in this Technical Report. Conclusions: Under the conditions of these 2-year feed studies, there was no evidence of carcinogenic activity of hydrochlorothiazide for male or female F344/N rats given feed containing 250, 500, or 2,000 ppm hydrochlorothiazide. There was equivocal evidence of carcinogenic activity of hydrochlorothiazide for male B6C3F1 mice, based on increased incidences of hepatocellular neoplasms. There was no evidence of carcinogenic activity for female B6C3F1 mice given diets containing 2,500 or 5,000 ppm hydrochlorothiazide. Chronic renal disease was more severe in rats administered hydrochlorothiazide, and increased incidences of secondary lesions (parathyroid hyperplasia, fibrous osteodystrophy, and mineralization in multiple organs) occurred in dosed rats. Synonym: 6-chloro-3,4-dihydro-2H-1,2,4-benzothia-diazine-7-sulfonamide 1,1-dioxide Trade Names: Aquarius; Bremil; Chlorzide; Cidrex; Dichlorosal; Dichlotride; Diclotride; Direma; Disalunil; Esidrix; Fluvin; Hidroronol; Hydril; Hydro-Aquil; Hydro-Diuril; Hydrosaluric; Hydrothide; Hypothiazide; Ivaugan; Jen-Diril; Maschitt; Nefrix; Neo-Codema; Neoflumen; Oretic; Panurin; Ro-Hydrazide; Thiaretic; Thiuretic; Urodiazin; Vetidrex