Animal Models of Colorectal Cancer in Chemoprevention and Therapeutics Development

Considering the complexity of genetic and epigenetic events that occurs during colorectal carcinogenesis and the uncertainty in relying solely on extrapolation from cell culture models, it is essential to use animal models relevant to the molecular characteristics of colorectal cancer (CRC). Requirements for a mouse model of CRC are that the model has relevance to the molecular pathways involved in human CRC, that there are correlates with factors that affect the frequency of the disease in human populations, and that the chemopreventive/therapeutic agent-induced signal is sufficient to carry out the project with an affordable and statistically significant number of mice. In this chapter we will discuss how animal models have not only advanced our understanding of CRC initiation and progression but have also greatly facilitated the development of newer chemopreventive and therapeutic strategies to reduce mortality and incidence.


Introduction
Considering the complexity of genetic and epigenetic events that occurs during colorectal carcinogenesis and the uncertainty in relying solely on extrapolation from cell culture models, it is essential to use animal models relevant to the molecular characteristics of colorectal cancer (CRC).Requirements for a mouse model of CRC are that the model has relevance to the molecular pathways involved in human CRC, that there are correlates with factors that affect the frequency of the disease in human populations, and that the chemopreventive/therapeutic agent-induced signal is sufficient to carry out the project with an affordable and statistically significant number of mice.In this chapter we will discuss how animal models have not only advanced our understanding of CRC initiation and progression but have also greatly facilitated the development of newer chemopreventive and therapeutic strategies to reduce mortality and incidence.

Animal models of colorectal cancer
Use of animal models could significantly expedite not only the delineation of molecular pathogenesis of colorectal carcinogenesis but could also aid in the development of newer preventive and therapeutic strategies.Animal tumor models can be classified as spontaneous and artificially transplanted systems.Spontaneous tumor models are now being widely considered for studying the biology of carcinogenesis and development of chemopreventive or chemosuppressive drugs.Initial animal models of CRC involved use of chemical carcinogens in mouse, rat, as well as in guinea pig.In the last two decades genetically modified mice such as APC Min/+ (Min: multiple intestinal neoplasia) with germline APC mutations at different sites have been extensively used for the investigation of therapeutic, chemopreventive and dietary factors for management of colorectal cancer (Hu et al., 2006;Gerner, 2007).Furthermore, studies to identify genetic modifiers of CRC are undertaken by generating mouse models representing molecular events involved in colorectal carcinogenesis like mismatch repair deficiency (MSH2-/-) and crossing

APC related models
The first heritable mouse model of colon cancer, APC Min/+ , was reported in 1990 as a result of ethylnitrosourea (ENU)-induced germline truncating mutation at the codon 850 of APC (Moser et al., 1990(Moser et al., , 1992;;Su et al., 1992).In the C57BL/6J mouse background APC Min/+ mice develop about 30 small intestinal polyps with occasional adenocarcinoma and essentially no tumor in the colon (McCart et al., 2008).Although this contrasts human FAP (familial adenomatous polyposis) where most of the adenomas are in the colon and these adenomas certainly progresses to invasive adenocarcinoma, the APC Min/+ models due to their phenotypic and histopathological similarities to human intestinal neoplasm are used not only to test therapeutic and chemopreventive interventions but also to understand the role of APC gene in CRC (Fodde & Smits, 2001).The APC Min/+ models also proved to be important for the study of genetic modifiers-of-Min (Mom) locus.When APC Min/+ C57BL/6J mice were crossed to AKR and MA mice, only 6 to 7 intestinal adenomas were observed and backcrossing F1 hybrids to the C57BL/6J helped identify a number of loci modifying number and distribution of adenomas (Moser et al., 1992;Gould et al., 1996;Fodde & Smits, 2001).The adenomas in the small intestine of the APC Min/+ mouse have dysplastic and hyperplastic crypts and villi but the colonic tumors are spherical and peduncular with dysplastic cells.Importantly, these adenomas display higher mitotic index than surrounding normal crypts (Kwong & Dove, 2009).The Min mice due to their many advantages have been extensively used by scientists to study molecular pathogenesis of CRC -i] Min mice contains a single genetic change that produces a organ-specific, consistent, and discrete tumor phenotype, ii] Adenomas in Min mice develop rapidly, with lesions visible as early as 60 days, iii] high tumor multiplicities (>100 /intestinal tract) providing strong statistical power, and iv] multiple pathways impacting tumorigenesis enable many entry points for basic or applied study (Kwong & Dove, 2009).Importantly, other mouse models with targeted genetic manipulations at different locations on APC have also been generated (summarized in Table 2).When heterozygous, the Δ474, Δ14, Δ716, lacZ, and Δ1309 mouse models show phenotypes similar to that of Min (Sasai et al., 2000;Oshima et al., 2001;Niho et al., 2003;Colnot et al., 2004).In contrast, heterozygosity for the 1638N allele results in 0-2 tumors (none in the colon) while the 1638T model is tumor-free and unlike any other truncating allele, 1638T homozygous is viable.The 1638N has only approximately 1-2% of the truncated protein and is referred to as leaky allele (Fodde & Smits, 2001).In contrast, 1638T has the full expression level of the truncated protein and is known as truncated allele.Furthermore, Li Q et al., (2005) inserted a neomycin cassette in either orientation (reverse (neoR) or forward (neoF)) into the 13 th intron of APC to generate full-length hypomorphic (expression reduced to 10-20%) alleles and showed that these heterozygous mice developed fewer than two adenomas per mouse (Li et al., 2005).The Cre/loxP conditional gene targeting system is developed to generate additional APC models to induce tumors specifically in the colon (Shibata et al., 1997;Colnot et al., 2004;Gounari et al., 2005;Hinoi et al., 2007).Because APC Min/+ mice rarely develop invasive cancer, efforts were made to develop compound mouse models by introducing alterations of genes known to be involved in CRC signaling pathways and produce invasive tumors mimicking human disease.The oncogene Kras, which is mutated in 40-50% of human CRC, is not altered in APC Min/+ polyps and the APC Min/+ mice with the introduction of Kras develop aggressive adenocarcinoma (Fodde & Smits, 2001).Loss of EphB receptor is an important event in the progression of CRC and APCMin/+ carrying a dominant negative EphB2 transgene showed 10 fold more tumor formation with greater number of invasive adenocarcinomas (Fodde & Smits, 2001;Batlle et al., 2005).The APC Min/+ mouse model has been quite useful as an experimental system for studying colorectal tumorigenesis and CRC chemoprevention strategies (Moser et al., 1990) because Min mice have two distinct advantages i] numerous adenomas with the same inherited APC mutation are available for analysis and ii] these adenomas develop in animals of uniform genetic background (Luongo et al., 1994).

Mismatch repair (MMR) deficient models
The HNPCC (hereditary nonpolyposis colorectal cancer) is an inherited condition with inactivated DNA mismatch repair (MMR) genes, like MLH1, MSH2, MSH6, and PMS2 (Fishel et al., 1993;Lynch & de la Chapelle, 2003) and leads to the development of a variety of cancers including that of the colon (Lynch & Smyrk, 1996).Mouse models with loss of function of MMR genes have been generated and mice lacking Mlh1, Msh2 and Msh6 develop tumors in stomach, small intestine, and colon.However, these mice also develop cancers of the lymphatic system, skin and lung (Reitmair et al., 1996;Edelmann et al., 1997Edelmann et al., , 1999Edelmann et al., , 2000)).Enhanced development of adenomas was observed in the APC Min/+ mice lacking Msh2 with increase in colonic adenoma numbers.Interestingly, these mice show normal growth and can reproduce but have reduced life span (Reitmair et al., 1996).
Although loss of Msh3 is not associated with increased tumors, loss of both the Msh3 and Msh6 leads to an increase in GI tumors at a younger age, similar to Mlh1 or Msh2-deficient mice (Edelmann et al., 2000).Mice bearing mutations in the Msh6 gene have a life span of 18 months and develop GI tumors within one year.The MMR mouse models carrying one functional copy of APC showed increasing mutation of APC and an enhanced frequency of intestinal neoplasia (Reitmair et al., 1996;Kuraguchi et al., 2001).Furthermore, mice lacking Mlh1 in APC1638N model have a 40-fold increase in adenomas compared to APC1638N mice alone (Edelmann et al., 1999).Interestingly, the PMS2 -/-mice are vulnerable to lymphomas but they do not develop GI tumors.However, the PMS2 -/-mice in APCMin background showed an increased number of adenomas in the GI-tract compared to Min alone (Prolla et al., 1998;Prolla, 1998;Baker et al., 1995).In contrast, the Mlh1/APC1638N mice showed a greater percentage of tumors progressed to invasive carcinomas (Edelmann et al., 1999).The MMR models are useful for screening of agents known to interfere with DNA mismatch repair processes for their therapeutic or carcinogenic effects.

TGF- models
The TGF- (transforming growth factor-) signaling pathway regulates a number of cellular processes including cellular differentiation, growth suppression, deposition of extracellular matrix and apoptosis (Figure 1).The TGF ligands through a heteromeric receptor mediate their effects on cells and dysregulation of the TGF- receptor 2 (TGF-R2) is the most common occurrence in the CRC (Bellam & Pasche, 2010;Grady et al., 1999).Although TGFR2 has been suggested to have a tumor suppressor function in CRC, recent reports indicate that it could act as a tumor suppressor as well as a tumor promoter (Tang et al., 1998;

Figure 1
Normal epithelium Dysplas c ACF Early adenoma adenoma Carcinoma Yang et al., 2002a, b;Blobe et al., 2000).Mouse models related to the TGF- pathway have been used to delineate the multifaceted role of this pathway during colorectal carcinogenesis.It has been reported that the TGF-1 deficient mice due to widespread inflammation die around three weeks of age (Shull et al., 1992;Kulkarni & Karlsson, 1993).Importantly, in the absence of Rag2 the TGF-1-deficient mice survive until adulthood (Diebold et al., 1995).Due to the significant incidence of carcinoma of the cecum and colon in these mice, the Rag2/TGF-1deficient mice remains a useful model to study the role of inflammation in CRC in relation to the TGF1 signaling (Engle et al., 1999;Maggio-Price et al., 2006).Mouse lacking TGF-2 in the colonic epithelium showed increased adenoma and adenocarcinoma formation after carcinogen treatment.Apart from TGF-, mouse models representing alterations in the other factors in this important pathway like SMAD2, SMAD4 and SMAD3 have been reported.(Eppert et al., 1996).Although mice lacking SMAD2 and SMAD4 are embryonically lethal, the SMAD3 deficient mice are viable and are a useful model of CRC (Zhu et al., 1998).Importance of the TGF- related models lies in the fact that the APC remains intact in the adenomas of these mice and these models could serve as a valuable tool to investigate non-WNT/APC/catenin-mediated colorectal carcinogenesis (Kaiser et al., 2007).Interestingly, the TGF- related models in the APC Min/+ background showed increased incidence of invasive carcinoma specifically in the distal colon (Takaku et al., 1998;Sodir et al., 2006).

Inflammation mediated models
Inflammatory bowel diseases (IBD) like ulcerative colitis (UC) and Crohn's Disease are predisposing conditions of CRC (Itzkowitz & Harpaz, 2004;Itzkowitz & Yio, 2004).Prolonged administration of dextransulfate sodium (DSS) in mice resulting in chronic colitis and formation of high-grade dysplasia confirmed the involvement of chronic inflammation in colorectal carcinogenesis (Okayasu et al., 1990).Interestingly intestinal tumorigenesis was augmented by combined administration of AOM and DSS.(Tanaka et al., 2003).To demonstrate that deficiency of Sigirr (single immunoglobulin and tollinterleukin 1 receptor domain) along with bacteria-induced inflammation increases susceptibility to CRC investigators effectively utilized the combined AOM/DSS model (Wald et al., 2003;Xiao et al., 2007;Uronis & Threadgill, 2009).Furthermore, the combined AOM/DSS model was also an important instrument in defining the role of the JAK/STAT (Janus kinase/signal transducers and activators of transcription) and NFkB (nuclear factor of kappa light chain gene enhancer in B-cells) pathways in inflammation-induced CRC (Wirtz & Neurath, 2007).The role of the JAK/STAT pathway in colorectal carcinogenesis was further confirmed by the fact that dysfunctional Socs1 and Socs3 (suppressors of cytokine signaling) leads to enhanced activation of STAT1, STAT3 and NFkB and subsequent growth of colorectal tumors (Hanada et al., 2006).Although lack of only Socs3 in the intestinal epithelial cells is not associated with chronic inflammation or tumor formation, these mice when treated with AOM/DSS showed distinct inflammatory response followed by colonic tumors (Hanada et al., 2006;Rigby et al., 2007).In contrast to increased nuclear -catenin in the Socs3 deficient mice, the colorectal tumors in Socs1 deficient mice display enhanced expression of Myc (Sutherland et al., 2006).In addition, the Muc2 -/-is an important animal model to study the role of inflammation in the colorectal carcinogenesis.This model specifically targets mucin-forming Muc2 and unlike other models tumor formation is also observed in the rectum (Mack & Hollingsworth, 1994;Yang et al., 2005;Femia et al., 2009).

Immune system related models
Models have been developed to investigate the role of immune system in colorectal carcinogenesis.Immune cells are involved in inflammatory response and inflammation is intimately related to CRC.Therefore, the IL-2 (interleukin-2), IL-10, and TCR knockout models were developed and used in studying role of diet and inflammation in colorectal cancer initiation and progression.(Rudolph et al., 1995;Kullberg et al., 1998;Mizoguchi et al., 2000;Seril et al., 2005).

Carcinogen-induced models
Although important information on familial and sporadic colorectal carcinogenesis was obtained from genetic animal models, it is the carcinogen-induced animal models that were instrumental in delineating molecular events specifically in the sporadic CRC.It is worth mentioning that the colon-specific carcinogen dimethylhyrdrazine (DMH) along with AOM has been useful in developing our current understanding of the molecular mechanisms underlying sporadic colorectal carcinogenesis (Druckrey et al., 1966).To its advantage the carcinogen-induced mouse model develops tumor which show much similarities to the pathophysiology of the human CRC (Kaiser et al., 2007;Uronis et al., 2007).The carcinogenmediated tumors show alterations in the WNT/-catenin pathway (Takahashi et al., 2000).Interestingly, the AOM-induced colorectal carcinogenesis, unlike in the APC Min/+ , is mainly due to the mutations in the Ctnnb1 gene, which encodes -catenin protein.Mutations in the Ctnnb1 results in ubiquitination-resistant stabilization of the -catenin leading to growth of colorectal adenomas associated with upregulation of proliferation markers like cyclin D1 and cMyc (Wang et al., 1998;Kaiser et al., 2007).The carcinogen-induced model was also a key player in the identification of the modifier loci like the Ptprj (a receptor-type protein tyrosine phosphatase), which has been shown to modify susceptibility to DMH and has shown frequent loss of heterozygosity in human colon cancer (Ruivenkamp et al., 2002).Additionally, the carcinogen-induced models led to the recognition of Pref1 as a modifier of CRC and the promoter of the Pref1 is suggested to contain a -catenin/TCF response element (Dong et al., 2004;Ruivenkamp et al., 2002;Uronis et al., 2007).

Mouse models in chemoprevention and therapeutics development
The genetic mouse models as well as the carcinogen-induced models of CRC (APC, TGF- and mismatch repair based models) are used to evaluate the effect of diets and chemopreventive agents.The chemopreventive and dietary interventions are usually started between 3 and 6 weeks of age and invariably the principal biological endpoint is the number and grade of the tumors.It is important to note that in contrast to human CRC where no small intestinal tumors are observed, the tumors in the mutant models are mostly in the small intestine.However, the concept that the non-steroidal anti-inflammatory drugs (NSAID) have a chemopreventive role in the CRC was first established in these models showing reduction in tumor number in the small intestine.The chemopreventive properties of NSAIDs like celecoxib and piroxicam were also validated in the carcinogen-induced models showing decrease in polyp number and size.Due to commercial availability, the APC Min/+ is the model of choice in many studies and a list of agents screened in APC Min/+ mouse model is shown in Table 3.Most of the other CRC mouse models are used for specific biological end points.But a few of them such as APC1638N model because of its longer life span and good signal to noise ratio is used for carcinogen testing and is suggested to be more useful than APC Min/+ mice (Trani et al., 2010).

Xenograft models of colon cancer
Colon cancer xenograft models are created by implantation of cells subcutaneously, intrasplenically, or into the renal capsule.It is important to implant the xenograft into the immunocompromised mice and commonly the T-cell deficient ''nude'' mice or NOD-SCID (non-obese diabetic/severe combined immunodeficiency) mice are used (Rygaard & Povlsen, 1969).The xenograft models of CRC are commonly used to assess newer therapeutics and understand the pathogenesis of human disease.Indeed, subcutaneous xenografts have found an important place in CRC research due to the fact that anesthetics are not required and the tumors are accessible for external measurement.Some of the disadvantages of the subcutaneous model are i] lack of tumor microenvironment representative of the CRC, and ii] in contrast to the >50% hepatic metastatic incidence of CRC, no metastasis is observed in the subcutaneous xenograft models.However, xenograft models involving intrasplenic or intra-renal-capsule, although have shown metastasis similar to human CRC, does not represent tumor microenvironment of CRC and signaling pathway could be different than the human disease (Furukawa et al., 1993;Fidler, 1991a, b, c).Consequently, implantation of CRC xenografts into mouse colon, the orthotopic model, is much preferred by the investigators due to their similar characteristics of the human ailment.

Orthotopic mouse model
An orthotopic mouse model involves placing of colorectal cancer cell or tumor tissue into the intestinal sub-mucosa (Tseng et al., 2007).The orthotopic model, unlike the subcutaneous model, is associated with all of the components of the tumor microenvironment as well as all of the angiogenic and growth factors, and cytokines.In addition to mimicking the human CRC in terms of metastasis and microenvironments, the orthotopic model also allows assessment of the alterations in the microenvironment on tumor initiation and progression.From a technical point of view, generation of orthotopic models demands specific expertise and more time than subcutaneous models.Because of technical difficulties in the physical measurement of the tumors, the orthotopic model also requires that an appropriate reporter like luciferase be in place for measuring tumor growth to determine the efficacy of a drug treatment.As with any animal model of human diseases there are inherent shortcoming and the orthotopic animal model is no exception.Because the tumors are in the colon, the orthotopic model requires sacrifice of the animals at a predetermined time for quantitative and qualitative analysis of the tumor.However, the orthotopic model has the advantage of mimicking human CRC including tumor microenvironment.

Zebrafish -A non-murine model of colorectal cancer
Signaling pathways involved in colorectal carcinogenesis are conserved across species and zebrafish, a well-characterized simple model system for human disease, are widely used to understand the molecular basis of cancer including CRC.Water borne carcinogens induce a wide variety of benign and malignant tumors in many organs of zebrafish.Zebrafish due to its easy maintainence and breeding along with conservation of human cancer-relevant oncogenes, and tumor suppressor and cell cycle genes makes it a useful model to study carcinogenesis.Interestingly, the zebrafish mutants display phenotypes similar to many human disorders, including cancer, cardiovascular disease, and neurodegeneration.Zebrafish carrying a mutation in the region representing most of the observed human APC www.intechopen.commutations were identified recently and like murine models the heterozygous fish develop intestinal adenomas (12%), which resembles its murine counterpart (Goessling et al., 2007).
The APC-heterozygous fish when exposed to dimethylbenzanthracene, showed significant increase in the tumor number with 44% developing liver tumors and 35% developing intestinal tumors and could serve as an important model system to screen carcinogens (Goessling et al., 2007).

Zebrafish model in chemoprevention and therapy
Zebrafish model is in use for the target selection, bioactive compound screening as well as in the drug toxicity and efficacy studies (Lieschke & Currie, 2007).Angiogenesis supports cancer progression including CRC and anti-angiogenic therapy inhibits cancer growth.On the contrary anti-angiogenic therapy has been implicated in inflammation a known risk factor of colorectal carcinogenesis.Our understanding of the correlation between tumor angiogenesis, inflammation, and metastasis was much enhanced by studies in the zebrafish model (Moshal et al., 2010).Furthermore, the zebrafish model has been used to study potential therapeutic agents like SKLB610 (inhibitor of angiogenesis related tyrosine kinase), which was reported to inhibit angiogenesis in zebrafish sub-intestinal veins (Cao et al., 2011).The zebrafish model was useful in determining the role of DNA demthylase in maintaining intestinal epithelial cells lacking APC in an undifferentiated state (Rai et al., 2010).Additionally, it has been reported that the zebrafish expressing a truncated form of APC with either retinoic acid or a selective COX-2 inhibitor decreased -catenin in the cell.Curcumin-loaded biodegradable polymeric micelles (Cur/MPEG-PCL) has been shown to efficiently block angiogenesis in transgenic zebrafish model (Gou et al., 2011).Similarly, the incorporation of doxorubicin in MPEG-PCL micelles enhanced the anticancer activity and decreased the systemic toxicity of doxorubicin in Zebrafish and has implications for CRC treatment (Lee et al., 2006).

Animal models in chemoprevention and chemotherapeutics development
Rodent models have been used for CRC research providing insight into the complex oncogenic events contributing to the loss of cell growth and differentiation control.These models also offer prospects to identify and study both therapeutic and chemopreventive agents.In general, almost all popular human colorectal cancer prevention strategies, from dietary manipulations (such as folate or calcium supplementation), to drug testing (such as (NSAIDs) have been evaluated in both carcinogen-induced and genetically modified animal models.In most cases, suppression of polyps has demonstrated the preventive effects of these strategies, and in some cases, investigators have been able to dissect pathways where these agents block the development of aberrant crypt foci (ACF).Moreover, approaches that are effective in preventing the early stages of colorectal tumorigenesis have been shown to actually promote tumor growth in later stages of the adenoma-carcinoma sequence.This type of observation in mice is important in polyp prevention studies in humans where folate supplementation may actually be harmful in subjects already predisposed to colorectal neoplasia.Although it is important to note that caution has to be exercised in extrapolating animal model data to human, at least studies with NSAID have shown similar protective effects both in human and animal.However, some differential response of chemopreventive drugs has been observed between animal studies and human response.This could be because of higher genetic homogeneity of mice compared to humans, physiologic differences in gut motility, hormones and immune surveillance, and differences in genetic events in somatic cells during adenoma to carcinoma transition in mouse compared to human colorectal neoplasia.

Comparison of human data with animal model data
With few exceptions, a significant correlation was observed between animal and human studies.Both the AOM rat model and the mutant mouse models supported the chemopreventive effects of the NSAID.This is supportive of the epidemiological studies proposing that, NSAIDs might reduce the colorectal cancer incidence by at least 45% in humans.Also supportive is the effects of celecoxib and sulindac shown to decrease the number of polyps in FAP patient trials.Similar to human data, rats and mice fed a high-fat diet showed increased adenomas than those fed a low-fat diet thus establishing the relationship between the colon cancer incidence and the intake of fat.Fatty diets with high linoleic acid content and n-6-polyunsaturated fatty acids seem to increase the number of tumors in rodents.Moreover, caloric reduction is a strategy that seems very efficient in animals.A reasonable agreement is observed between the results of these animal studies and the more limited clinical studies with few differences (Corpet & Pierre, 2003).

Limitations of animal models
Currently a number of animal models are available to dissect various facets of CRC and to undertake risk estimation studies.Mutant mouse models provide a unique opportunity in studying numerous adenomas under defined experimental conditions and uniform genetic background.However, use of animal models in studying human disease has its own limitations.For example, in carcinogen-induced models of CRC, the tumor incidence and latency period could be modulated by amount of carcinogen used -higher amount of carcinogen leading to higher incidence of tumors.However, high ethanol consumption reduced carcinogen (DMH)-induced tumorigenesis suggesting that DMH model is not useful in determining the role of alcohol in CRC.This discrepancy was resolved using the APC Min/+ mice model where ethanol consumption was observed as a risk factor for CRC (Roy et al., 2002).Careful consideration is essential for the selection of animal model to study a particular agent and requires validation is two or more models for the unequivocal demonstration.

Conclusions
Future advances in animal model development will require combinations of dietary and genetic manipulation of rodents or other inexpensive animals to more accurately mimic the various factors that contribute to colorectal neoplasia in humans.As epidemiologic and molecular studies demonstrate the heterogeneity of colorectal tumor development in diverse populations (e.g. the microsatellite instability or CIMP pathways), it is expected that any one model will not answer all the questions about the CRC chemoprevention or therapeutic intervention strategies under investigation.

Acknowledgment
This work was supported by funding from NASA (Grant#NNX09AU95G).

Fig. 1 .
Fig. 1.Integrated molecular pathway implicated in development of colorectal cancer (Courtesy: KEGG pathway, www.genome.jp/kegg).Alterations in Wnt signaling including Wnt, APC, Axin and TCFs are associated increased -catenin level and increased cell proliferation.The MAPK signaling is associated with the oncogenic activation of RAS and ERK signaling leading to increase cell proliferation).The TGF- pathway is mainly a growth inhibitory pathway and any perturbations leads to suppressed apoptosis and increased cell proliferation.The mismatch repair (MMR) pathway maintains DNA homeostasis by facilitating post-replication repair and dysfunction results in accumulation of potential mutations and genetic instability implicated in the development of CRC.Important candidate proteins altered in CRC are highlighted in red color.

Allele/trunc ation location Average life- span Genetic back- ground Homozyg ous pheno- type Heterozygous phenotype Average no. of spontaneo us tumors References
*Conditional expression; N/A=Not applicable; N/R=Not reported Table 2. Adematous polyposis coli (APC) based intestinal tumor models www.intechopen.comColorectal Cancer -From Prevention to Patient Care 282