PRINCIPLES OF TOXICOLOGY - Biology East Borneo

13.5 TESTING CHEMICALS FOR CARCINOGENIC ACTIVITY 291number of animals required to detect a small percent increase in tumor incidence above the animal’sbackground rate. This means that it may be difficult to detect small increases for those tumor typesthat have large spontaneous background rates in an animal model when the test group contains only50–100 animals.To increase the safety of the animal-to-human extrapolations, the number of animals tested in acancer bioassay may need to be increased if (1) the number of humans to be exposed to the chemicalis either expected to be large or (2) a small margin of safety exists between the animal dose tested andthe expected human exposure. In general, however, resource limitations are such that only 50 animalsof each sex are tested at each dose for both species; this limits the total number of animals tested toabout 400 animals, plus 200 animals to serve as controls.Short-Term Cancer Bioassays and Other Measures of Carcinogenic PotentialBecause of the large number of animals, lengthy timeframe, and expense associated with the chroniccarcinogenesis bioassay, there has long been a need for reliable shorter-term tests of carcinogenicpotential that could be used to complement standard carcinogenicity testing protocols. In the past, suchshort-term tests were limited to abbreviated initiation–promotion experiments with defined endpointsand the induction of tumors in susceptible animal models (e.g., lung tumors in strain A mice). Recentlyhowever, the tools of molecular biology have made it possible to construct genetically altered(transgenic) animals that may prove to be useful models for predicting the carcinogenic potential ofchemicals. The U.S. National Toxicology Program is currently in the process of validating two of thesetransgenic models, Tg.AC mice and p53 +/– mice, with chemicals previously tested in the standard2-year chronic bioassay. These transgenic models are described briefly below.1. The Tg.AC line was produced in FVB/N mice by the incorporation of a v-H-ras transgene intothe cellular DNA. Mutations in ras oncogenes, which encode a family of GTP binding proteins criticalto many growth factor signaling pathways, have been detected in a large proportion of human tumors.Tg.AC mice behave like genetically initiated mice, and rapidly develop epidermal papillomas inresponse to topical treatment with carcinogens. Researchers have shown that the mutant transgene isoverexpressed in the proliferating cells in benign and malignant tumors but is not expressed in normalcells. Interestingly, treatment with initiators or tumor promoters induces the development of skintumors. While treated mice have a dramatic increase in tumor yield with abbreviated time-to-tumorresponse, untreated mice have a normal skin histology and do not usually develop spontaneous tumorswithin the testing period. Treatment with carcinogens results in the production of papillomas in lessthan 6 months, substantially reducing the period of time for a typical initiation-promotion experimentin mouse skin.2. Heterozygous p53 +/– mice possess only a single functional copy of the p53 gene. As discussedpreviously, p53 function is lost through mutation or deletion in over 50 percent of all human cancers.With only a single functional allele, p53 mice develop normally but are at an increased susceptibilityto the induction of tumors. This situation is analogous to an individual who has inherited a defectivecopy of a tumor suppressor gene. Upon dosing with mutagenic carcinogens, p53 +/– mice rapidlydevelop tumors compared to normal mice, usually within 6 months. Untreated p53 +/– mice do notusually develop tumors within the test period.In the chemicals tested thus far, there has been a high degree of concordance with results fromtraditional chronic bioassays. Examination of the discordant cases indicates that a number of these aredue to the absence of hepatocellular carcinoma in the p53 +/– mice. As will be discussed, the B6C3F 1mice typically used in chronic bioassays have a high spontaneous background rate of these tumors,making interpretation of positive carcinogenic responses in this tissue problematic. These and othertransgenic animal models hold promise as less expensive and time-consuming adjuncts or replacementsfor conventional chronic bioassays. In addition, some scientists believe that transgenic animal models