title: NTP Toxicology and Carcinogenesis Studies of Hexachlorocyclopentadiene (CAS No. 77-47-4) in F344/N Rats and B6C3F1 Mice (Inhalation Studies).
abstract: Hexachlorocyclopentadiene is an intermediate used in the manufacture of flame retardants, resins, and chlorinated cyclodiene pesticides. Toxicology and carcinogenesis studies were conducted by exposing male and female F344/N rats and B6C3F1 mice to atmospheres containing hexachlorocyclopentadiene (approximately 98% pure) for 6 hours per day, 5 days per week, for 13 weeks or 2 years. A stop-exposure evaluation was conducted in male B6C3F1 mice to determine the influence of exposure level and exposure duration on the development of nonneoplastic lesions of the respiratory tract and on their regression or progression after exposure was stopped. Genetic toxicology studies were conducted in Salmonella typhimunum, cultured Chinese hamster ovary cells, Drosophila melanogaster, and mouse peripheral blood samples were analyzed for frequency of micronucleated normochromatic erythrocytes. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female rats were exposed to atmospheres containing 0, 0.04, 0.15, 0.4, 1, or 2 ppm (equivalent to 0, 0.45, 1.67, 4.46, 11.14, and 22.28 mg/m(3)) hexachlorocyclopentadiene. Additional rats were exposed to 0, 0.04, 0.4, or 2 ppm hexachlorocyclopentadiene and evaluated for differences in clinical pathology parameters. All rats in the 1 and 2 ppm groups died during the first 4 weeks of the study. The final mean body weight and mean body weight gain of males exposed to 0.4 ppm were significantly lower than those of the controls. Listlessness was observed in 2 ppm rats from week 1, in 1 ppm rats from week 2, and in 0.4 ppm rats during week 3. Rats exposed to 1 or 2 ppm also experienced respiratory distress. No chemical-related differences in hematology, clinical chemistry, or urinalysis parameters were observed in male or female rats. Absolute and relative lung weights of 0.4 ppm males were significantly greater than those of the controls. Inflammation (necrotizing, chronic, or suppurative) of the nose, larynx, trachea, and lung was observed in 0.4, 1, and 2 ppm males and females. Squamous metaplasia of the epithelial lining of the nose of 0.4 ppm males and 1 and 2 ppm males and females was also observed. 13-WEEK STUDY IN MICE: Groups of 10 male and 10 female mice were exposed to atmospheres containing 0, 0.04, 0.15, 0.4, 1, or 2 ppm (equivalent to 0, 0.45, 1.67, 4.46, 11.14, and 22.28 mg/m(3)) hexachlorocyclopentadiene. Additional mice were exposed to 0, 0.04, 0.4, or 2 ppm and evaluated for differences in clinical pathology parameters. All 2 ppm mice died during the first week of exposure. All 1 ppm mice died during the first 5 weeks of exposure. Five males and two females in the 0.4 ppm group died during the first 2 weeks of exposure. Deaths in the other groups were not related to hexachlorocyclopentadiene exposure. Final mean body weights of males exposed to 0.15 and 0.4 ppm and the body weight gain of 0.4 ppm males were significantly lower than those of the controls. Treatment-related clinical findings included listlessness in 0.4 and 1 ppm males and females. No chemical-related differences in hematology, clinical chemistry, or urinalysis parameters were observed in male or female mice. Necrosis or inflammation of the nose, larynx, trachea, or lung occurred in mice exposed to 0.4,1, and 2 ppm hexachlorocyclopentadiene. Squamous metaplasia of the larynx or trachea was observed in 0.15, 0.4, and 1 ppm males and in 0.4 and 1 ppm females. 2-YEAR STUDY IN RATS: Survival, Body Weights, Clinical Findings, and Urinalysis Groups of 60 male and 60 female rats were exposed to atmospheres containing 0, 0.01, 0.05, or 0.2 ppm (equivalent to 0, 0.11, 0.56, and 2.28 mg/m(3)) hexachlorocyclopentadiene. Survival rates and mean body weights of exposed rats were similar to those of the controls. No chemical-related clinical findings were observed in male or female rats during the 2-year study. No differences in urinalysis parameters at the 15-month interim evaluation could be attributed to exposure to hexachlorocyclopentadiene. Pathology Findings: No increases in neoplasm incidences could be attributed to hexachlorocyclorocyclopentadiene. Toxicity was limited to the respiratory tract and included an increase in the incidence of pigmentation of the respiratory epithelium of the nose, trachea, and the bronchi and bronchioles of the lung in both males and females. Exposure to hexachlorocyclopentadiene also caused an increase in the incidence of squamous metaplasia of the laryngeal epithelium of exposed females; the incidences in 0.01 and 0.2 ppm females were significantly greater than that of the controls. The severity of squamous metaplasia was minimal in all exposed and control females. 2-YEAR STUDY IN MICE: Survival, Body Weights, Clinical Findings, and Urinalysis Groups of 60 male and 60 female mice were exposed to atmospheres containing 0, 0.01, 0.05, or 0.2 ppm (equivalent to 0, 0.11, 0.56, and 2.28 mg/m(3)) hexachlorocyclopentadiene. The 2-year survival rate of female mice in the 0.2 ppm group was marginally lower than that of the controls due to a higher incidence of ovarian inflammation in 0.2 ppm females. Mean body weights of 0.2 ppm males (weeks 62 to 103) and females (throughout the study) were lower than those of the controls. No clinical findings in male or female mice were attributed to chemical exposure during the 2-year study. There were no chemical-related differences in urinalysis parameters at the 15-month interim evaluation. Pathology Findings: The site of toxicity of hexachlorocyclopentadiene exposure in mice in the 2-year study was the respiratory tract. Chemical-related pigmentation of the respiratory epithelium of the nose, trachea, and lung and suppurative inflammation of the nose were observed. No increased neoplasm incidences in males or females could be attributed to hexachlorocyclopentadiene exposure. STOP-EXPOSURE EVALUATION: Survival, Body Weights, and Clinical Findings Groups of male mice were exposed to atmospheres containing 0.2 ppm hexachlorocyclopentadiene for 33 or 66 weeks or 0.5 ppm for 26 or 42 weeks followed by exposure to air until the end of the study. Fifty male mice from each stop-exposure group were evaluated at 2 years. Two-year survival rates of stop exposure groups were similar to that of the controls. Final mean body weights of stop-exposure groups were similar to that of the controls. No chemical related clinical findings were observed. Pathology Findings: Nonneoplastic respiratory tract lesions similar to those observed in the core study were observed in males in the stop-exposure groups. Chemical-related pigmentation and inflammation of the respiratory epithelium were persistent as indicated by their presence in many male mice after recovery periods of 62 to 78 weeks, and the incidence and severity of the lesions were related to exposure concentration and duration. GENETIC TOXICOLOGY: Hexachlorocyclopentadiene was not mutagenic in Salmonella typhimurium strains TA98, TA100, TA1535, and TA1537 when tested with and without S9. Hexachlorocyclopentadiene did induce sister chromatid exchanges and chromosomal aberrations in cultured Chinese hamster ovary cells, with and without S9. No induction of sex-linked recessive lethal mutations was observed in male Drosophila melanogaster treated with hexachlorocyclopentadiene by feeding or injection, and no increase in the frequency of micronucleated erythrocytes was seen in male or female B6C3F1 mice exposed to hexachlorocyclopentadiene by inhalation for 13 weeks. CONCLUSIONS: Under the conditions of these 2-year studies, there was no evidence of carcinogenic activity of hexachlorocyclopentadiene in male or female F344/N rats or B6C3F1 mice exposed to 0.01, 0.05, or 0.2 ppm. Exposure of rats to hexachlorocyclopentadiene produced pigmentation of the respiratory epithelium of the nose, trachea (males), and bronchi and bronchioles of the lung. Squamous metaplasia of the laryngeal epithelium occurred in female rats exposed to hexachlorocyclopentadiene. Suppurative inflammation of the nose as well as pigmentation of the respiratory mucosal epithelium occurred in mice exposed to hexachlorocyclopentadiene. Synonyms: Perchlorocyclopentadiene, hexachloro-1,3-cyclopentadiene, HEX, HCPD, HCCP, HCCPD Trade Name: C-56-Graphlox