Brief Overview of Alternative Models
The TG.AC transgenic line was produced in FVB/N mice by pronuclear injection of a v-Ha-ras transgene linked to a fetal zeta-globin promoter and an SV40 polyadenylation/splice sequence. TG.AC mice behave like genetically initiated mice, rapidly developing epidermal papillomas in response to topical tumor promoter or carcinogen treatment. In some experiments, the time to first tumor appearance has been as short as 3 weeks, but all chemicals tested that produced papillomas did so in less than 20 weeks. A dose-response relationship has been observed with promoters and carcinogens studied to date and in some studies mice have developed papillomas too numerous to count. In addition, TG.AC mice appear to respond to genotoxic as well as nongentotoxic carcinogens. While the response in treated mice is dramatic, it is important to note that untreated, singly housed TG.AC mice usually develop few, if any, spontaneous tumors and the histology of the skin is normal. With respect to mechanism, it has been shown that the v-Ha-ras transgene is not significantly expressed in non-tumor bearing TG.AC tissues but is over-expressed in the proliferating component of benign and malignant tumors. Thus, it appears that expression of the transgene drives proliferation and subsequent tumor development in carcinogen treated TG.AC skin.
References:
Tennant, R.W., J.E. French and J.W. Spalding (1995) Identification of chemical carcinogens and assessing potential risk in short term bioassays using transgenic mouse models, Environ. Health Perspect., 103, 942-950.
Tennant, R.W., Spalding, J. and French,J.E. Evaluation of transgenic mouse bioassays for identifying carcinogens and noncarcinogens, Mutation Res. 365, 119-127 (1996).
p53 def
This mouse model has an alteration of the p53 tumor suppressor gene, a gene critical to cell cycle control and DNA repair and one often found to be mutated or lost in human and rodent tumors. Mice with a single copy of the wildtype p53 allele (p53+/- heterozygous) offer a single target for mutagens, a condition analogous to humans with some heritable forms of cancer. The heterozygous state should increase the probability for either loss of p53 tumor suppressor function or gain of transforming activity by requiring only a single mutation. P53 heterozygous mice are viable and show a low background tumor incidence up to almost 12 months of age. Nullizygous (p53-/-)mice have a higher rate of spontaneous tumors at sites apparently determined by the mouse strain's genetic background.
References:
Tennant, R.W., J.E. French and J.W. Spalding (1995) Identification of chemical carcinogens and assessing potential risk in short term bioassays using transgenic mouse models, Environ. Health Perspect., 103, 942-950.
Tennant, R.W., Spalding, J. and French,J.E. Evaluation of transgenic mouse bioassays for identifying carcinogens and noncarcinogens, Mutation Res. 365, 119-127 (1996).
CB6F1-TgHras2
These transgenic (Tg) mice carry the prototype human c-Hras gene with its own promoter region, which encodes the prototype c-Hras gene product (i.e., p21). Five or six copies of human c-Hras gene are integrated into the genome of each Tg mouse in a tandem array. Transgenes are expressed in the tumors and in normal tissues, and the total amount of p21 detected by immunoblot analysis is two to three times higher in Tg mice than in non-Tg mice (CB6F1 not carrying human c-Hras). No mutations of the transgenes are detected in the normal tissues of the Tg mice. The Tg mice develop spontaneous tumors within 18 months after birth; however, none or only a few spontaneous lung tumors are observed at 33 to 35 weeks of age, when a short term carcinogenicity testing is terminated. Early results indicate that a more rapid onset and a higher incidence of more malignant tumors can be expected with a higher probability after treatment with various genotoxic carcinogens in the CB6F1-Hras2 mice than in control non-Tg mice.
Yamamoto, S.; Hayashi, Y.; Mitsumori, K; Nomura, T. (1997) Rapid Carcinogenicity Testing System with Transgenic Mice Harboring Human Prototype c-Hras Gene. Laboratory Animal Science, 47 (2) 121-126.
SHE Assay
In vitro cell-transformation systems have been developed so that at least part of the carcinogenesis process can be simulated with cultured cells. These systems allow tumor production to be used as an end point. In addition, it appears that cell-transformation systems are suited to identify genotoxic and non-genotoxic carcinogens. Syrian hamster embryo (SHE) cells have been used as recipients of DNA transfection and is one of these systems used for identification of potential oncogenes. In general, short comings of cell-transformation systems preventing them from gaining widespread use have been an ambiguity in judging transformed cells and the absence of inter-laboratory reproducibility. The SHE cell transformation suffered from the fact that the number of transformed colonies was too low to provide credible results. A more sensitive SHE cell-transformation assay has been developed which shows a dose-response relationship for carcinogens. Data so far obtained from this model provide support for the use of the modified SHE cell-transformation protocol for the detection of carcinogens which operate through genotoxic or non-genotoxic mechanisms.
References:
LeBoeuf RA, Kerckaert GA, Aardema MJ, Gibson DP, Brauninger R, Isfort RJ. (1996) The pH 6.7 Syrian hamster embryo cell transformation assay for assessing the carcinogenic potential of chemicals. Mutat. Res. 356(1):85-127.
Neonatal Mouse
The neonatal B6C3F1 mouse tumorigenicity bioassay is highly sensitive to direct-acting genotoxic carcinogens, with the liver being the principal target organ. Mechanistic studies on metabolic activation, exogenous DNA adduct formation, and analysis of ras-oncogene activation are required to confirm that a direct-acting mechanism is responsible for tumor formation. The major genetic determinant for the sensitivity of the mouse liver to hepatocarcinogenesis has been attributed to the presence of the Hcs (hepatocarcinogen sensitivity) locus, and current data suggests a biological origin for the decreased latency of tumorigenesis in the B6C3F1 neonatal mouse model. The tumorigenic response once initiated will be expressed more quickly in the rapidly growing liver of the newborn compared with that of the adult. Data from several neonatal mouse studies in which chemicals had been tested in common with the the 2-year bioassay indicate good concordance in results. However, most of the chemicals tested in the neonatal assay were known direct-acting carcinogens. It is unlikely that the neonatal mouse bioassay is sensitive to indirect-acting genotoxic carcinogens since, in most cases, indirect-acting carcinogens act as cell proliferators (tumor promoters) and in neonates, liver cells are replicating at such a high rate that any additional increase caused by exogenous proliferators would be negligible. The neonatal mouse bioassay appears to be sensitive to a wide range of genotoxic chemicals that are believed to intiate the tumor response via the formation of a covalently bound carcinogen-DNA adduct. Thus, the neonatal mouse bioassay appears to distinguish between direct- and indirect-acting carcinogens. It is believed that convenient and economical mechanistic studies can be developed to distinguish between an indirect-acting genotoxic carcinogen and a non-carcinogen.
References:
Flammang, T.J., L.S. Von Tungeln, F.F. Kadlubar, and P.P. Fu (1997) Neonatal mouse assay for tumorigenicity: Alternative to the chronic rodent bioassay. Regul. Toxicol. and Pharmacol. (in press)
Fu, P.P., L.S. Von Tungeln, G.J. Hammons, G. McMahon, G.N. Wogan, T.J. Flammang, and F.F. Kadlubar. Carcinogenicity in the B6C3F1 neonatal mouse: a potential alternative bioassay. In: Progress in Clinical and Biological Research, R.A. LeBoeuf, T.J. Slaga, and R. Tennant (eds), Wiley-Liss series, (in press)
XPA-/-
Nucleotide excision repair (NER) is involved in the removal of a broad range of DNA lesions induced by ultraviolet light and various chemical compounds. Defects in the NER pathway are associated with several human inheritable diseases including xeroderma pigmentosum (XP). XP patients have a high frequency of UVB-induced skin tumors. In XP there are seven complementation groups of which XP complementation group A (XP-A) is one of the most severe and frequently found groups in humans. In XP-A patients, mutations and small deletions have been found in the XP-A correcting gene. The XPA protein is presumably involved in the recognition of DNA damage, i.e., the pre-incision step of NER, because XPA binds to damaged DNA and has also been found to interact with other NER proteins. XPA-deficient mice (XPA -/- ) appear to be normal up to 1 yr in age. However, when their skins were exposed to UVB, the XPA -/- mice developed skin tumors at a high frequency comparable to that of human XP-A. In addition, it has been shown that XPA -/- mice are sensitive toward various genotoxic carcinogens irrespective of their application routes. Further studies will explore the suitability of the XPA -/- mouse model as a rapid and sensitive short-term test of genotoxic carcinogens.
References:
de Vries A, van Oostrom CT, Dortant PM, Beems RB, van Kreijl CF, Capel PJ, van and Steeg H. (1997) Spontaneous liver tumors and benzo[a]pyrene-induced lymphomas in XPA-deficient mice. Mol. Carcinog. 19(1):46-53
Berg RJ, de Vries A, van Steeg H, de Gruijl FR (1997) Relative susceptibilities of XPA knockout mice and their heterozygous and wild-type littermates to UVB-induced skin cancer.Cancer Res. 57(4):581-584
de Vries A, van Steeg H (1996) Xpa knockout mice.Semin Cancer Biol 7(5):229-240
de Vries, A., Van Oostrom C.Th.M., Hofhuis, F.M.A., Dortant, P.M., Berg, R.J.W., De Gruijl, F.R., Wester, P.W., Van Kreijl, C.F, Capel, P.J.A., Van Steeg, H. and Verbeek, S.J. (1995) Increased susceptibility to ultraviolet-B and carcinogens of mice lacking the DNA excision repair gene XPA. Nature, 377, 169-173.
MMTV/RAS (Tg.SH) has the MMTV/vHa-ras transgene
MMTV/MYC (Tg.M) has the MMTV/cmyc transgene
MMTV/NEU (Tg.NK) has the MMTV/c-neu transgene
Breeding stock of Tg.M, Tg.SH and Tg.NK obtained from the Howard Hughes Medical Institute, Harvard Medical School were mated to cesarian derived FVB/N inbred mice. Transgene carriers from litters produced by mating heterozygote transgene carrier males of FVB/N origin with FVB/N females and were identified by Southern hybridization analysis on DNA extracted from tail samples.
PIM = pim-1.
Developed by van Lohuizen, et al., at the Netherlands Cancer Institute. This model exhibits a low level of spontaneous tumor formation however is predisposed to rapid lymphoma formation by chemical induction. Founder line pp64a received at GenPharm International in from the Berns lab at two backcrosses onto C57BL/LiA(N2) from the founder line (CBA/BrA X C57BL/LiA). Transferred to Taconic and cesarean derived. Backcrossed once on the C57BL/6Ntac at GenPharm International. Homozygous breeders are produced through intercrossing. Heterozygous mice are produced through mating of homozygous male and C57BL/6Ntac females.
TG.LAC1/C57BL/6 (Big Blue)
Stratagene has developed a line of inbred C57BL/6 transgenic mice for mutation analysis in vivo. Lambda phage shuttle vectors containing a LacIq target gene have been integrated into the genome of each mouse. After delivery of a test compound to the mouse, the lambda phage vectors containing the target gene can be quickly and easily recovered from isolated genomic DNA using Transpack&TM; in vitro packaging extract. Mutations in the target gene are detected by infecting host E. coli with the rescued phage and plating in the presence of the chromogenic substrate X-gal. Blue plaques are formed by mutations in the target LacIq genes, while intact LacIq genes result in clear plaques. The mutant frequency of the text compound is determined by counting the number of easy to ready blue plaques versus colorless plaques.
C57BL/6-APC+/APC-1638N (TG.APC)
A mouse lineage carrying a targeted 3¿ mutation in the endogenous Apc gene. Mice homozygous for the targeted frameshift mutation at a position corresponding to amino acid 1638 (Apc 1638N) are embryonic lethal. Heterozygous Apc 1638N mice develop 5 ¿ 6 intestinal adenomas per animal during their lifetime.
P16(INK4A)/(+/-) (C57BL/6)
Is heterozygous for the tumor suppressor gene INK4a which codes for p16INK4a and p14ARF. p16INK4a, a cell cycle inhibitor, is inactivated in many human tumors and in families with hereditary melanoma and pancreatic cancer. Heterozygous INK4a-/- mice are viable and fertile and are derived from parental strains C57BL/6 and WW6 that is ~ 75% 129/Sv, ~ 20% C57BL/6, and ~ 5% SJL. Mice carrying a targeted INK4a deletion develop tumors at an early age.
TRAMP
(Description from Dr. Norm Greenberg, Associate Professor, Department of Cell Biology, Michael DeBakey Center for Biomedical Education and Research at Baylor College of Medicine - http://www-tramp-model.cellb.bcm.tmc.edu/features.htm)
A transgene carrying the 426/+28 fragment of the rat probasin (PB) gene fused to the SV40 T antigen (Tag) has been used to generate an independent transgenic autochthonous model for prostate cancer in the C57Bl/6 inbred strain of mice. Expression of the PB-Tag transgene is initially regulated by androgens and restricted to the prostate epithelial cells of the dorsolateral and ventral lobes. By the time the mice are 12 weeks of age, TRAMP mice histologically display mild to severe hyperplasia with cribriform structures. Severe hyperplasia and adenocarcinoma is observed by 18 weeks of age. By 24-30 weeks of age, all TRAMP males display primary tumors and metastasis are commonly detected in the lymph nodes and lungs and less frequently in the bone, kidney and adrenal glands. The epithelial origin of the tumors and metastatic deposits has been demonstrated. When castrated at 12 weeks of age, 80% of TRAMP mice ultimately develop prostate cancer, demonstrating that the tumors are at least initially androgen dependent. Interestingly, the castrated mice develop more poorly differentiated primary tumors and twice the incidence of metastatic disease compared to non-castrate TRAMP controls. The TRAMP mice have also been used as a source for the development of new research materials and at least three novel prostate cancerepithelial cell lines (TRAMP-C1, -C2 and -C3) have been established from the primary prostate tumors. These cell lines all express the androgen receptor but no longer express the SV40 early genes. TRAMP-C1 and -C2 form tumors when grafted subcutaneously into syngeneic C57Bl/6 hosts.
Cyclooxygenase (COX-1- and COX-2-)
Deficient mice have unique physiological differences that have allowed investigation into the individual biological roles of the COX isoforms. In the following, the phenotypes of the two COX knockout mice are summarized, and recent studies to investigate the effects of COX deficiency on inflammatory responses and cancer susceptibility are discussed. The data suggest that both isoforms have important roles in the maintenance of physiological homeostasis and that such designations as house-keeping and/or response gene may not be entirely accurate. Furthermore, data from COX-deficient mice indicate that both isoforms can contribute to the inflammatory response and that both isoforms have significant roles in carcinogenesis.
Reference:
Langenbach R, Loftin CD, Lee C, Tiano H.Cyclooxygenase-deficient mice. A summary of their characteristics and susceptibilities to inflammation and carcinogenesis. Ann N Y Acad Sci 1999;889:52-61
The estrogen receptor knockout (ERKO) model was created by gene targeting in embryonic stem cells of C57BL/6J mice to disrupt the estrogen receptor gene. The estragen receptor gene encodes a protein that binds and mediates the effects of 17b-estradiol. Homozygous ERKO mice survive to adulthood but develop syndromes that result in infertility in both sexes. These alterations in physiology and behavior of the ERKO mouse provide broad and multiple advantages to the research efforts toward understanding the function and mechanism of estrogen action.
Lubahn DB, Moyer JS, Golding TS, Couse JF, Korach KS, Smithies O. Alteration of reproductive function but not prenatal sexual development after insertional disruption of the mouse estrogen receptor gene. Proc Natl Acad Sci U S A 1993 Dec 1;90(23):11162-6
Couse JF, Korach KS. Reproductive phenotypes in the estrogen receptor-alpha knockout mouse. Ann Endocrinol (Paris) 1999 Jul;60(2):143-8
F344/NTac
The HLA-B27 transgenic rat, expressing HLA-B27 and human ß2m, exhibits a phenotype similar to humans suffering B27 related rheumatic disorders such as reactive arthritis, ankylosing spondylitis, psoriasis and inflammatory bowel disease. This rat model is useful for research and pharmacological studies of spontaneous systemic inflammation, arthritis, inflammatory bowel disease and skin diseases including psoriasis and alopecia.
References: Hammer, R.E., Maika, S.D., Richardson, J.A., Tang, J.P., Taurog, J.D. (1990) Spontaneous Inflammatory Disease in Transgenic Rats Expressing HLA-B27 and Human b2M: An Animal Model of HLA-B27-Associated Human Disorders, Cell, Vol. 63, pp. 1099-1112.
