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A targeted constitutive mutation in the APC tumor suppressor gene underlies mammary but not intestinal tumorigenesis.

Gaspar C, Franken P, Molenaar L, Breukel C, van der Valk M, Smits R, Fodde R - PLoS Genet. (2009)

Bottom Line: Moreover, somatic APC mutations play a rate-limiting and initiating role in the majority of sporadic colorectal cancers.This hypomorphic mutant allele results in intermediate levels of Wnt/beta-catenin signaling activation when compared with other Apc mutations associated with multifocal intestinal tumors.The histology of the Apc1572T primary mammary tumours is highly heterogeneous with luminal, myoepithelial, and squamous lineages and is reminiscent of metaplastic carcinoma of the breast in humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Josephine Nefkens Institute, Erasmus MC, Rotterdam, The Netherlands.

ABSTRACT
Germline mutations in the adenomatous polyposis coli (APC) gene are responsible for familial adenomatous polyposis (FAP), an autosomal dominant hereditary predisposition to the development of multiple colorectal adenomas and of a broad spectrum of extra-intestinal tumors. Moreover, somatic APC mutations play a rate-limiting and initiating role in the majority of sporadic colorectal cancers. Notwithstanding its multifunctional nature, the main tumor suppressing activity of the APC gene resides in its ability to regulate Wnt/beta-catenin signaling. Notably, genotype-phenotype correlations have been established at the APC gene between the length and stability of the truncated proteins encoded by different mutant alleles, the corresponding levels of Wnt/beta-catenin signaling activity they encode for, and the incidence and distribution of intestinal and extra-intestinal tumors. Here, we report a novel mouse model, Apc1572T, obtained by targeting a truncated mutation at codon 1572 in the endogenous Apc gene. This hypomorphic mutant allele results in intermediate levels of Wnt/beta-catenin signaling activation when compared with other Apc mutations associated with multifocal intestinal tumors. Notwithstanding the constitutive nature of the mutation, Apc(+/1572T) mice have no predisposition to intestinal cancer but develop multifocal mammary adenocarcinomas and subsequent pulmonary metastases in both genders. The histology of the Apc1572T primary mammary tumours is highly heterogeneous with luminal, myoepithelial, and squamous lineages and is reminiscent of metaplastic carcinoma of the breast in humans. The striking phenotype of Apc(+/1572T) mice suggests that specific dosages of Wnt/beta-catenin signaling activity differentially affect tissue homeostasis and initiate tumorigenesis in an organ-specific fashion.

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Phenotypic and molecular analysis of the compound in cis Apc+/1572T/Smad4+/Sad mouse model.(A) Schematic illustration of the chr. 18 LOH event in intestinal tumors from in cis Apc+/1572T/Smad4+/Sad mice leading to loss of both Smad4 and Apc wild-type alleles. (B) Comparative phenotypic analysis of the intestinal and mammary tumor predisposition among Apc+/1572T, Smad4+/Sad, and Apc+/1572T/Smad4+/Sad mice. Notes: (1) The incidence of GI tumors was calculated after exclusion of the pyloric lesions as these present in clusters often difficult to count. (2) The multiplicity of GI tumors was calculated based on all animals with the exception of those where the high tumor burden made the count not feasible. The asterisks indicate that the apparent absence of intestinal tumor in Smad4+/Sad control animals is not in contradiction with what previously published. These mice were sacrificed at time points matched with the ages at which compound Apc+/1572T/Smad4+/Sad mice had to be sacrificed due to the high GI and mammary tumor burden (♀: 90.4 days +/−28.4; ♂: 118.5 days +/−26.2). However, in Smad4+/Sad animals the majority of the tumors appear at 9 months of age [15]. (C) H&E staining of intestinal tumor sections from Apc+/1572T (top), Apc+/1572T/Smad4+/Sad (middle), and Smad4+/Sad (bottom) mice. (D) Smad4 IHC analysis of two intestinal adenomas from Apc+/1572T/Smad4+/Sad mice showing loss of Smad4 expression. LOH was observed in 100% of the polyps (n = 15) analyzed. PCR–based LOH analysis of the same cohort of Apc+/1572T/Smad4+/Sad polyps revealed loss of wild-type Apc allele in 87% of the cases (13/15; data not shown).
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pgen-1000547-g006: Phenotypic and molecular analysis of the compound in cis Apc+/1572T/Smad4+/Sad mouse model.(A) Schematic illustration of the chr. 18 LOH event in intestinal tumors from in cis Apc+/1572T/Smad4+/Sad mice leading to loss of both Smad4 and Apc wild-type alleles. (B) Comparative phenotypic analysis of the intestinal and mammary tumor predisposition among Apc+/1572T, Smad4+/Sad, and Apc+/1572T/Smad4+/Sad mice. Notes: (1) The incidence of GI tumors was calculated after exclusion of the pyloric lesions as these present in clusters often difficult to count. (2) The multiplicity of GI tumors was calculated based on all animals with the exception of those where the high tumor burden made the count not feasible. The asterisks indicate that the apparent absence of intestinal tumor in Smad4+/Sad control animals is not in contradiction with what previously published. These mice were sacrificed at time points matched with the ages at which compound Apc+/1572T/Smad4+/Sad mice had to be sacrificed due to the high GI and mammary tumor burden (♀: 90.4 days +/−28.4; ♂: 118.5 days +/−26.2). However, in Smad4+/Sad animals the majority of the tumors appear at 9 months of age [15]. (C) H&E staining of intestinal tumor sections from Apc+/1572T (top), Apc+/1572T/Smad4+/Sad (middle), and Smad4+/Sad (bottom) mice. (D) Smad4 IHC analysis of two intestinal adenomas from Apc+/1572T/Smad4+/Sad mice showing loss of Smad4 expression. LOH was observed in 100% of the polyps (n = 15) analyzed. PCR–based LOH analysis of the same cohort of Apc+/1572T/Smad4+/Sad polyps revealed loss of wild-type Apc allele in 87% of the cases (13/15; data not shown).

Mentions: To provide additional experimental support for the “just right” signaling model for Apc-driven mammary tumorigenesis, we have taken advantage of a recent study according to which Tgf-β signaling antagonizes canonical Wnt signaling thus negatively regulating stem cell self-renewal [13]. Hence, Tgf-β alterations such as those resulting from Smad4 loss of function mutations, are expected to lead to a further increase of Wnt/β-catenin signaling in the Apc-mutant cellular background. Therefore, we have bred Apc+/1572T animals with Smad4+/Sad, a mouse model for juvenile polyposis previously developed in our laboratory [14]. Smad4+/Sad animals are characterized by a late-onset predisposition to hyperplastic intestinal polyps which develop in the absence of a 2nd hit at either the Smad4 or the Apc locus [15]. As both these tumor suppressor genes map to chromosome 18 in the mouse, we have generated Apc+/1572T/Smad4+/Sad compound heterozygous mice where both targeted alleles are in the in cis phase on chr. 18 as previously described for the Apc1638N model [15]. As shown in Figure 6A, Apc+/1572T/Smad4+/Sad mice show a similar incidence of mammary adenocarcinomas similarly to Apc+/1572T, but are characterized by multiple GI-tract tumors. These polyps are of the adenomatous type and become apparent at a much earlier age than the hyperplastic lesions with a pronounced stromal component characteristic of the Smad4+/Sad model (Figure 6B). Moreover, the vast majority of the Apc+/1572T/Smad4+/Sad intestinal polyps show loss of the entire chr. 18 carrying the wild type alleles of both tumor suppressor genes [15] (Figure 6C), whereas the intestinal lesions characteristic of the Smad4+/Sad mice retain the wild type Smad4 allele at first, and show Smad4 LOH (but not at the Apc locus) only at more advanced progression stages [15]. Although it cannot be excluded that loss of Smad4 function underlies intestinal tumour formation in these animals through Tgf-β/BMP downstream effectors independent of Wnt signaling, the histology and molecular features of the Apc+/1572T/Smad4+/Sad GI polyps strongly suggest that the further increase of Wnt/β-catenin signaling conferred by the Smad4 mutation in the Apc-mutant background results in intestinal tumours in the compound mice without apparently affecting the mammary cancer phenotype.


A targeted constitutive mutation in the APC tumor suppressor gene underlies mammary but not intestinal tumorigenesis.

Gaspar C, Franken P, Molenaar L, Breukel C, van der Valk M, Smits R, Fodde R - PLoS Genet. (2009)

Phenotypic and molecular analysis of the compound in cis Apc+/1572T/Smad4+/Sad mouse model.(A) Schematic illustration of the chr. 18 LOH event in intestinal tumors from in cis Apc+/1572T/Smad4+/Sad mice leading to loss of both Smad4 and Apc wild-type alleles. (B) Comparative phenotypic analysis of the intestinal and mammary tumor predisposition among Apc+/1572T, Smad4+/Sad, and Apc+/1572T/Smad4+/Sad mice. Notes: (1) The incidence of GI tumors was calculated after exclusion of the pyloric lesions as these present in clusters often difficult to count. (2) The multiplicity of GI tumors was calculated based on all animals with the exception of those where the high tumor burden made the count not feasible. The asterisks indicate that the apparent absence of intestinal tumor in Smad4+/Sad control animals is not in contradiction with what previously published. These mice were sacrificed at time points matched with the ages at which compound Apc+/1572T/Smad4+/Sad mice had to be sacrificed due to the high GI and mammary tumor burden (♀: 90.4 days +/−28.4; ♂: 118.5 days +/−26.2). However, in Smad4+/Sad animals the majority of the tumors appear at 9 months of age [15]. (C) H&E staining of intestinal tumor sections from Apc+/1572T (top), Apc+/1572T/Smad4+/Sad (middle), and Smad4+/Sad (bottom) mice. (D) Smad4 IHC analysis of two intestinal adenomas from Apc+/1572T/Smad4+/Sad mice showing loss of Smad4 expression. LOH was observed in 100% of the polyps (n = 15) analyzed. PCR–based LOH analysis of the same cohort of Apc+/1572T/Smad4+/Sad polyps revealed loss of wild-type Apc allele in 87% of the cases (13/15; data not shown).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2697381&req=5

pgen-1000547-g006: Phenotypic and molecular analysis of the compound in cis Apc+/1572T/Smad4+/Sad mouse model.(A) Schematic illustration of the chr. 18 LOH event in intestinal tumors from in cis Apc+/1572T/Smad4+/Sad mice leading to loss of both Smad4 and Apc wild-type alleles. (B) Comparative phenotypic analysis of the intestinal and mammary tumor predisposition among Apc+/1572T, Smad4+/Sad, and Apc+/1572T/Smad4+/Sad mice. Notes: (1) The incidence of GI tumors was calculated after exclusion of the pyloric lesions as these present in clusters often difficult to count. (2) The multiplicity of GI tumors was calculated based on all animals with the exception of those where the high tumor burden made the count not feasible. The asterisks indicate that the apparent absence of intestinal tumor in Smad4+/Sad control animals is not in contradiction with what previously published. These mice were sacrificed at time points matched with the ages at which compound Apc+/1572T/Smad4+/Sad mice had to be sacrificed due to the high GI and mammary tumor burden (♀: 90.4 days +/−28.4; ♂: 118.5 days +/−26.2). However, in Smad4+/Sad animals the majority of the tumors appear at 9 months of age [15]. (C) H&E staining of intestinal tumor sections from Apc+/1572T (top), Apc+/1572T/Smad4+/Sad (middle), and Smad4+/Sad (bottom) mice. (D) Smad4 IHC analysis of two intestinal adenomas from Apc+/1572T/Smad4+/Sad mice showing loss of Smad4 expression. LOH was observed in 100% of the polyps (n = 15) analyzed. PCR–based LOH analysis of the same cohort of Apc+/1572T/Smad4+/Sad polyps revealed loss of wild-type Apc allele in 87% of the cases (13/15; data not shown).
Mentions: To provide additional experimental support for the “just right” signaling model for Apc-driven mammary tumorigenesis, we have taken advantage of a recent study according to which Tgf-β signaling antagonizes canonical Wnt signaling thus negatively regulating stem cell self-renewal [13]. Hence, Tgf-β alterations such as those resulting from Smad4 loss of function mutations, are expected to lead to a further increase of Wnt/β-catenin signaling in the Apc-mutant cellular background. Therefore, we have bred Apc+/1572T animals with Smad4+/Sad, a mouse model for juvenile polyposis previously developed in our laboratory [14]. Smad4+/Sad animals are characterized by a late-onset predisposition to hyperplastic intestinal polyps which develop in the absence of a 2nd hit at either the Smad4 or the Apc locus [15]. As both these tumor suppressor genes map to chromosome 18 in the mouse, we have generated Apc+/1572T/Smad4+/Sad compound heterozygous mice where both targeted alleles are in the in cis phase on chr. 18 as previously described for the Apc1638N model [15]. As shown in Figure 6A, Apc+/1572T/Smad4+/Sad mice show a similar incidence of mammary adenocarcinomas similarly to Apc+/1572T, but are characterized by multiple GI-tract tumors. These polyps are of the adenomatous type and become apparent at a much earlier age than the hyperplastic lesions with a pronounced stromal component characteristic of the Smad4+/Sad model (Figure 6B). Moreover, the vast majority of the Apc+/1572T/Smad4+/Sad intestinal polyps show loss of the entire chr. 18 carrying the wild type alleles of both tumor suppressor genes [15] (Figure 6C), whereas the intestinal lesions characteristic of the Smad4+/Sad mice retain the wild type Smad4 allele at first, and show Smad4 LOH (but not at the Apc locus) only at more advanced progression stages [15]. Although it cannot be excluded that loss of Smad4 function underlies intestinal tumour formation in these animals through Tgf-β/BMP downstream effectors independent of Wnt signaling, the histology and molecular features of the Apc+/1572T/Smad4+/Sad GI polyps strongly suggest that the further increase of Wnt/β-catenin signaling conferred by the Smad4 mutation in the Apc-mutant background results in intestinal tumours in the compound mice without apparently affecting the mammary cancer phenotype.

Bottom Line: Moreover, somatic APC mutations play a rate-limiting and initiating role in the majority of sporadic colorectal cancers.This hypomorphic mutant allele results in intermediate levels of Wnt/beta-catenin signaling activation when compared with other Apc mutations associated with multifocal intestinal tumors.The histology of the Apc1572T primary mammary tumours is highly heterogeneous with luminal, myoepithelial, and squamous lineages and is reminiscent of metaplastic carcinoma of the breast in humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Josephine Nefkens Institute, Erasmus MC, Rotterdam, The Netherlands.

ABSTRACT
Germline mutations in the adenomatous polyposis coli (APC) gene are responsible for familial adenomatous polyposis (FAP), an autosomal dominant hereditary predisposition to the development of multiple colorectal adenomas and of a broad spectrum of extra-intestinal tumors. Moreover, somatic APC mutations play a rate-limiting and initiating role in the majority of sporadic colorectal cancers. Notwithstanding its multifunctional nature, the main tumor suppressing activity of the APC gene resides in its ability to regulate Wnt/beta-catenin signaling. Notably, genotype-phenotype correlations have been established at the APC gene between the length and stability of the truncated proteins encoded by different mutant alleles, the corresponding levels of Wnt/beta-catenin signaling activity they encode for, and the incidence and distribution of intestinal and extra-intestinal tumors. Here, we report a novel mouse model, Apc1572T, obtained by targeting a truncated mutation at codon 1572 in the endogenous Apc gene. This hypomorphic mutant allele results in intermediate levels of Wnt/beta-catenin signaling activation when compared with other Apc mutations associated with multifocal intestinal tumors. Notwithstanding the constitutive nature of the mutation, Apc(+/1572T) mice have no predisposition to intestinal cancer but develop multifocal mammary adenocarcinomas and subsequent pulmonary metastases in both genders. The histology of the Apc1572T primary mammary tumours is highly heterogeneous with luminal, myoepithelial, and squamous lineages and is reminiscent of metaplastic carcinoma of the breast in humans. The striking phenotype of Apc(+/1572T) mice suggests that specific dosages of Wnt/beta-catenin signaling activity differentially affect tissue homeostasis and initiate tumorigenesis in an organ-specific fashion.

Show MeSH
Related in: MedlinePlus