a-Methyldopa sesquihydrate is used in the treatment of hypertension; over 20 million prescriptions are written annually for a -methyldopa or a-methyldopa sesquihydrate in the United States. a-Methyldopa sesquihydrate (USP grade, greater than 99% pure) was selected for study because of widespread human exposure and the lack of carcinogenicity studies on this compound.
Fourteen-day, 13-week, and 2-year studies were conducted in F344/N rats and B6C3F1 mice. The chemical was administered in feed because human exposure is primarily by the oral route. Short-term studies were performed in bacteria and mammalian cells to evaluate the potential for genetic damage.
Fourteen-day and thirteen-week studies
In the 14-day studies, the chemical was administered at dietary concentrations of 0 and 6,250-100,000 ppm. All rats receiving 100,000 ppm and 2/5 female rats receiving 50,000 ppm died. All mice lived until the end of the studies. Final mean body weights of dosed male rats were 14%-43% lower than that of controls, and those of dosed female rats were 9%-24% lower. Feed consumption by dosed male and female rats was reduced. Final mean body weights of dosed mice were generally within 10% of those of controls; feed consumption by dosed groups was lower than that by controls during the first week of the studies.
In the 13-week studies, the chemical was administered at dietary concentrations of 0 and 3,100-50,000 ppm. Deaths occurred in 4/10 male rats, 7/10 female rats, and 2/10 female mice at 50,000 ppm and in 1/10 female rats at 25,000 ppm. Final mean body weights of dosed rats were 6%-46% lower than those of controls. Feed consumption by dosed rat groups was lower than that by controls. Final mean body weights of male mice at 25,000 and 50,000 ppm and female mice at 50,000 ppm were reduced 12%-19%. Feed consumption by dosed and control mice was comparable.
Rats and mice receiving 25,000 and 50,000 ppm exhibited clinical signs of toxicity including lethargy, hyperexcitability, ocular discharge, and rough hair coats. Clinical signs of toxicity were judged to be more severe in dosed male mice than in female mice. Minimal to moderate kidney tubular cell regeneration was seen in male and female rats at 12,500, 25,000, and 50,000 ppm. Bone marrow hypoplasia occurred in male rats at 25,000 and 50,000 ppm and in female rats at 6,300 ppm and higher. Nuclear enlargement (karyomegaly) of the renal corticaltubular epithelium was observed in male and female mice administered 12,500-50,000 ppm; these kidney lesions were judged to be more severe and occurred more frequently at concentrations of 25,000 ppm and higher.
Because of kidney lesions, bone marrow responses, and body weight effects at 12,500 ppm and higher and increased deaths and clinical signs at 25,000 and 50,000 ppm, dietary concentrations selected for male and female rats in the 2-year studies were 0, 3,100, and 6,300 ppm. Based on clinical signs, kidney effects, and body weight decreases at 25,000 and 50,000 ppm, dietary concentrations selected for male and female mice in the 2-year studies were 0, 6,300, and 12,500 ppm. Diets containing the chemical at these concentrations were fed to groups of 50 male and 50 female rats and 50 male and 50 female mice for 103 weeks.
Two-year studies
Body weight and survival
Mean body weights of dosed rats were generally 8%-17% lower than those of controls, and mean body weights of dosed mice were generally 5%-22% lower than those of controls throughout the studies. The average amount of a-methyldopa sesquihydrate consumed per day was approximately 110-120 or 230-240 mg/kg per day by low and high dose rats and 830-890 or 1,760-1,800 mg/kg by low and high dose mice. Survival was comparable among dosed and control groups (male rats: control, 28/50; low dose, 26/50; high dose, 27/50; female rats: 35/50; 34/50; 29/50; male mice: 44/50; 42/50; 39/50; female mice: 42/50; 40/50; 38/50). Clinical signs considered to be dose-related included fighting in male rats, irritability in male mice, and rough hair coats in female mice.
Nonneoplastic and neoplastic effects
Several lesions of the forestomach, including edema, chronic inflammation, epithelial hyperplasia, and ulcers, were seen at low incidences in high dose rats. No forestomach neoplasms occurred. No neoplastic lesions were observed in either male or female rats which were considered related to a-methyldopa sesquihydrate exposure.
Nephropathy (control, 3/50; low dose, 21/50; high dose, 32/50), karyomegaly (nuclear enlargement) of cells of the tubular epithelium (0/50; 46/50; 44/50, and cysts (2/50; 10/50; 10/50) were observed in the kidney of dosed female mice. Low incidences of tubular cell hyperplasia (0/50; 1/50; 1/50), tubular cell adenomas (0/50; 2/50; 0/50), and tubular cell adenocarcinomas (0/50; 0/50; 1/50) were observed in male mice. Tubular cell adenomas (3/2,029, 0.15%) and tubular cell adenocarcinomas (3/2,029, 0.15%)are uncommon in untreated control male B6C3F1 mice. No neoplastic lesions in female mice were considered related to a-methyldopa sesquihydrate exposure.
Decreased incidences of several site-specific neoplasms were observed in dosed rats and mice; these decreases might have been due in part to decreased weight gain in dosed groups. The decreases occurred in the adrenal medulla of male rats (pheochromocytomas or malignant pheochromocytomas, combined: 21/49; 3/49; 10/50), uterus of female rats (endometrial stromal polyps: 15/50; 5/49; 1/50), liver of male and female mice (hepatocellular adenomas or carcinomas, combined-- male: 15/50; 5/50; 6/50; female: 4/50; 1/50; 0/50), and anterior pituitary gland of female mice (adenoma: 9/49; 4/40; 2/50). The incidences of malignant tumors (male: 19/50; 9/50; 8/50; female: 21/50; 16/50; 12/50) and benign or malignant tumors (combined) (male: 32/50; 15/50; 17/50; female: 33/50; 22/50; 21/50) were reduced in dosed mice.
Reproductive studies
a-Methyldopa sesquihydrate was administered to male F344/N rats in corn oil by gavage 5 days per week for 65 days at doses of 0, 50, 100, 200, or 400 mg/kg. Decreased body weight was seen in dosed animals. Male rats were mated to untreated female F344/N rats on days 57-61, necropsies were performed on days 65-67, and reproductive toxicity was measured by sperm count, sperm motility, organ weights, hormone levels, and histologic evaluation of the testis. Decreased fertility was observed in males dosed with a-methyldopa sesquihydrate at 200 and 400 mg/kg. Decreases were also seen in sperm count, sperm motility, apparent number of late spermatids, and plasma testosterone levels in males in the 200 and 400 mg/kg groups. This alteration of reproductive function in male rats was found to be reversible after a 13-week recovery period (without dosing). The decreased fertility observed after a-methyldopa sesquihydrate administration was probably due in part to the decreases in plasma testosterone levels.
Genetic toxicity
a-Methyldopa sesquihydrate was not mutagenic when tested with or without exogenous metabolic activation with a preincubation protocol in four strains of Salmonella typhimurium (TA97, TA98, TA100, or TA1535). No increase in chromosomal aberrations or sister chromatid exchanges was observed in Chinese hamsterovary (CHO) cells exposed to a-methyldopa sesquihydrate with or without S9.
Data audit
The data, documents, and pathology materials from the 2-year studies of a-methyldopa sesquihydrate 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 a-methyldopa sesquihydrate for male or female F344/N rats fed diets containing 3,100 or 6,300 ppm. There was equivocal evidence of carcinogenic activityof a-methyldopa sesquihydrate for male B6C3F1 mice, as shown by three dosed mice having uncommon tubular cell tumors of the kidney. There was no evidence of carcinogenic activity of a -methyldopa sesquihydrate for female B6C3F1 mice fed diets containing 6,300 or 12,500 ppm. Nonneoplastic lesions of the kidney including karyomegaly were observed in dosed female mice. Decreased incidences of several tumor types (in the adrenal gland in male rats, uterus in female rats, liver in male and female mice, and anterior pituitary gland in female mice) were considered related to a-methyldopa sesquihydrate exposure.