NTP Comparative Initiation/Promotion Skin Paint Studies of B6C3F1 Mice, Swiss (CD-1(R)) Mice, and SENCAR Mice.
In 1983, an ad hoc panel on chemical carcinogenesis testing and evaluation recommended additional methods that should be used by the National Toxicology Program (NTP) for the detection and evaluation of chemical carcinogens. One recommendation was that there should be an increased emphasis on short-term tests to detect agents that do not exert genetic effects such as some promoting agents. Initiation/promotion models have been used routinely to identify chemicals with promoting potential and to study tumorigenesis. In one model, a topical subcarcinogenic dose of a chemical is first applied to the back of the skin (initiation) followed by repeated topical applications of one or more chemicals (promotion) and the skin is monitored for tumor development. Mouse skin has been shown to be more responsive (i.e., develops tumors using this protocol) than other commonly used laboratory rodent models. However, not all mouse strains are equally sensitive. The skin tumor response of the B6C3F1 mouse using the initiation/promotion protocol was not known. Since the B6C3F1 mouse is commonly used in NTP carcinogenesis studies and much is known of its biology and response to chemical carcinogens, known initiators and promoters were used to compare the tumor response sensitivity of B6C3F1 mouse skin to that of two often-used responsive strains, Swiss (CD-1(R)) and SENCAR mice. The combination of 7,12- dimethylbenz(a)anthracene (DMBA) initiation and 12-O-tetradecanoylphorbol-13- acetate (TPA) promotion was selected because this pair is routinely used to study tumorigenesis. However, DMBA requires metabolic activation to achieve initiation and it was possible that the B6C3F1 mouse metabolism might not make this conversion (DiGiovanni and Juchau, 1980). Therefore, a second study was conducted using N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), a direct acting carcinogen, as the initiator. MNNG is not used as frequently for mouse skin studies as is DMBA. In addition to the promoter TPA, benzoyl peroxide (BPO), a non-phorbol ester and known promoter after DMBA initiation, was also used (Slaga et al., 1981). Each initiating chemical was used in combination with each promoting chemical as described on the study design table below. Additional groups of male and female mice of each strain were treated with repeated applications of acetone (vehicle control), repeated applications of promoter (TPA or BPO) without prior initiation treatment (promoter reference controls), or a single application of the initiator (DMBA or MNNG) followed by repeated applications of acetone (initiator controls). All three strains of mice demonstrated sensitivity by developing skin tumors after topical application of the chemicals under study (DMBA, MNNG, TPA, and BPO). The most sensitive of the three strains appeared to be SENCAR mice, in the sense that lower doses of the test chemical were generally required to produce effects equivalent to those in the other two strains. Skin tumors also tended to develop earlier and with greater multiplicity in SENCAR mice than in the other two strains. By these criteria, the overall sensitivity of Swiss (CD-1(R)) mice was intermediate, and B6C3F1 mice showed the least overall sensitivity to dermal carcinogenicity. In response to recommendations regarding specific short-term tests and also on the skin tumor response sensitivity of various initiators and promoters, SENCAR mice would be the most acceptable strain to use for such studies. Though the B6C3F1 mice were less responsive in the skin initiation/promotion protocol, promotion data from this strain may, at times, be of more use in explaining mechanisms of tumor development (e.g. when there is a strain specific response observed in 2-year carcinogenicity studies or effects on melanocytes are suspected). See the full report or study abstract for table of Study Design for the 1-Year Comparative Initiation/Promotion Skin Paint Studies. The 1-year complete carcinogen studies used repeated applications of low concentrations of the carcinogens DMBA and MNNG. The skin tumor response in aln all three strains under these conditions was more similar than in the initiation and promotion studies. There was a high incidence of skin tumors in all three strains with both carcinogens. More B6C3F1, and SENCAR mice developed skin tumors and averaged more tumors per mouse than did Swiss (CD-1&reg;) mice. Skin tumors developed earlier in SENCAR mice than in B6C3F1 and Swiss (CD-1&reg;) mice. Although B6C3F1 mice exhibited the lowest overall sensitivity to the initiation/promotion protocol when compared to Swiss (CD-1&reg;) and SENCAR mice, the response of B6C3F1 mice was similar to Swiss (CD-1&reg;) and SENCAR mice for complete carcinogen studies.