Background:

Phenolphthalein was formerly used as a laxative. Phenolphthalein is known to cause cancer in rats and mice. We tested phenolphthalein in a genetically modified mouse strain that lacks two tumor suppressor genes as part of a study to determine if this mouse model could detect cancer-causing chemcals more rapidly than the standard 2-year rodent bioassay.

Methods:

We gave groups of 15 male or 15 female haploinsufficient p16^Ink4a/p19^Arf mice feed containing phenolphthalein for 27 weeks. The concentrations were 200, 375, 750, 3,000 or 12,000 parts per million (ppm) of phenolphthalein; other animals receiving untreated feed served as the control groups. Tissues from 22 organs were examined for every animal.

Results:

Exposure to phenolphthalein caused atrophy of the testes, reduction in spermatids, and kidney nephropathy and hypertrophy in male haploinsufficient p16^Ink4a/p19^Arf mice and hyperplasia of the thymus and hematopoietic cell proliferation of the spleen in male and female haploinsufficient p16^Ink4a/p19^Arf mice. However, there were no increased incidences of cancer in the exposed animals.

Conclusions:

We conclude that phenolphthalein did not cause cancer in male or female haploinsufficient p16^Ink4a/p19^Arf mice, although it has in some other strains of mice and in rats. Phenolphthalein did cause atrophy of the testes and nephropathy and hypertrophy of the kidney in male haploinsufficient p16^Ink4a/p19^Arf mice and hyperplasia of the thymus and hematopoietic cell proliferation of the spleen in male and female haploinsufficient p16^Ink4a/p19^Arf mice.

27-Week Study in Mice:

Groups of 15 male and 15 female mice were exposed to 0, 200, 375, 750, 3,000, or 12,000 ppm phenolphthalein (equivalent to average daily doses of approximately 35, 65, 135, 540, and 2,170 mg phenolphthalein/kg body weight to males and 50, 90, 170, 680, 2,770 mg/kg to females) in feed for 27 weeks. Survival of all exposed groups of male and female mice was similar to that of the control groups. Mean body weights of males in the 12,000 ppm group were less than those of the control group after week 11. No differences in feed consumption were noted between exposed and control groups.

Atypical hyperplasia of the thymus, a premalignant change of chemically induced thymic lymphoma, occurred in exposed males and females, and the incidence was significantly increased in 12,000 ppm females. Atrophy of the seminiferous tubules in the testis, hyperplasia of the testicular interstitial (Leydig) cells, and epididymal hypospermia occurred in most 3,000 and 12,000 ppm males. Additionally, the left and right testis weights, the left epididymis weights, sperm motility, the numbers of spermatid heads per testis, and sperm heads per cauda and per gram cauda epididymis were generally significantly less in 3,000 and 12,000 ppm males than in the control group. The incidences of nephropathy were significantly increased in 3,000 and 12,000 ppm males; incidences of hypertrophy of renal tubules were significantly increased in males receiving 750 ppm or greater. Hematopoietic cell proliferation of the spleen occurred in all 12,000 ppm males, and the incidences of this lesion were significantly increased in 375, 750, and 12,000 ppm females.

Genetic Toxicology:

The frequency of micronucleated erythrocytes was assessed at four time points during the 27-week study in male and female haploinsufficient p16^Ink4a/p19^Arf mice. Significant concentration-related increases in micronucleated cells were observed at all time points in male and female mice.

Conclusions:

Under the conditions of this 27-week feed study, there was no evidence of carcinogenic activity^* of phenolphthalein in male or female haploinsufficient p16^Ink4a/p19^Arf mice exposed to 200, 375, 750, 3,000, or 12,000 ppm. Because this is a new model, there is uncertainty whether the study possessed sufficient sensitivity to detect a carcinogenic effect.

Phenolphthalein induced atypical hyperplasia, a preneoplastic lesion of the thymus, in male and female mice, hematopoietic cell proliferation of the spleen in male and female mice, and toxicity to the kidney and reproductive system in male mice.