The Cdx2 homeobox gene has a tumour suppressor function in the distal colon in addition to a homeotic role during gut development.

BACKGROUND: During development, the homeobox gene Cdx2 exerts a homeotic function, providing the positional information necessary for correct specification of the midgut endoderm. This is illustrated by the non-neoplastic gastric-type heteroplasias present at birth in the pericaecal region of Cdx2(+/-) mice. Furthermore, intestinal expression of Cdx2 continues throughout life but diminishes in colorectal cancers compared with adjacent normal tissue, suggesting a role in tumorigenesis. AIM: To investigate the consequence of altered Cdx2 expression on colon tumour initiation and/or progression. METHODS: Heterozygous Cdx2(+/-) mice were analysed for spontaneous malignant tumours and for tumour development after treatment with a DNA mutagen, azoxymethane. RESULTS: Cdx2(+/-) mice did not spontaneously develop malignant tumours. After azoxymethane treatment, the gastric-like heteroplasias in the pericaecal region did not evolve into cancer indicating that they are not precancerous lesions. However, azoxymethane treated Cdx2(+/-) mice developed tumours specifically in the distal colon 12 weeks after azoxymethane treatment whereas no tumours were found in wild-type littermates at this stage. Histopathological and molecular analyses indicated that these tumours were invasive adenocarcinomas that recapitulated the malignant sequence observed in the majority of sporadic colorectal cancers in human. In addition, we found that the colonic epithelium was less sensitive to radiation induced apoptosis in Cdx2(+/-) than in wild-type mice. CONCLUSION: This study provides the first experimental evidence that Cdx2 is a tumour suppressor gene involved in cancer progression in the distal colon. This action in adults is functionally and geographically distinct from its homeotic role during gut development.

Homeobox genes and gut development.

The gut of vertebrates exhibits a common anteroposterior regional differentiation. The role of homeobox genes in establishing this pattern is inferred by their sites of expression. It is suggested that the primary source of positional information is in the endoderm, which subsequently establishes a 'dialogue' with the surrounding visceral layer of the lateral plate mesoderm. This results in the anatomical and physiological specialization of the adult gut.

Reprogramming of intestinal differentiation and intercalary regeneration in Cdx2 mutant mice.

The homeobox gene Cdx2, a homologue of the Drosophila gene caudal, has been implicated in the control of cell differentiation in the intestinal epithelium. Recently, we showed that mice in which one allele of the Cdx2 gene had been inactivated by homologous recombination developed multiple intestinal polyp-like lesions that did not express Cdx2 and that contained areas of squamous metaplasia in the form of keratinizing stratified squamous epithelium, similar to that occurring in the mouse esophagus and forestomach. We have now examined colonic lesions from 98 Cdx2+/- mice and report that the lesions are composed of heterotopic stomach and small intestinal mucosa. We conclude that Cdx2 directs endodermal differentiation toward a caudal phenotype and that haploinsufficient levels of expression in the developing distal intestine lead to homeotic transformation to a more rostral endodermal phenotype, such as forestomach epithelium that does not express Cdx2 during normal development. Intercalary growth (epimorphic regeneration), which previously has never been described in mammals, then occurs, resulting in the ordered "filling in" of tissue types at the discontinuity between the gastric and colonic epithelia. This intercalary growth in a restricted space results in the formation of the polypoid lesions observed.

CDX2, a human homologue of Drosophila caudal, is mutated in both alleles in a replication error positive colorectal cancer.

The Cdx2 gene is one of three murine homologues of the Drosophila homeobox gene caudal. Mice heterozygous for a null mutation in Cdx2 exhibit a variable phenotype including tail abnormalities, stunted growth and a homeotic shift of vertebrae. Most strikingly, however, 90% of heterozygous mice were reported to develop multiple intestinal adenomatous polyps, most notably in the proximal colon (Chawengsaksophak et al., 1997). These observations led us to propose that mutation of CDX2 may be involved in the genesis of some human colorectal tumours. A survey of DNA from 85 colorectal tumours revealed that one with extensive microsatellite instability (RER+ phenotype) has mutations in both alleles of CDX2. Both mutations occur in coding regions which contain repetitive elements and are consistent with those found in RER + tumours.

Homeosis and intestinal tumours in Cdx2 mutant mice.

In Drosophila, disturbing the expression of the homeobox gene caudal causes a severe disruption in body segmentation and global body patterning. There are three mouse homologues of Drosophila caudal: Cdx1 (ref. 2), Cdx2 (ref. 3) and Cdx4 (ref. 4). We have generated a null mutation of murine Cdx2 by homologous recombination. Cdx2 homozygote null mutants die between 3.5 and 5.5 days post coitum (d.p.c.). Cdx2 heterozygote mutants exhibit a variable phenotype, with many showing tail abnormalities or stunted growth. Skeletal analysis demonstrates a homeotic shift of vertebrae and compatible malformations of the ribs. Within the first three months of life, 90% of Cdx2 heterozygotes develop multiple intestinal adenomatous polyps, particularly in the proximal colon. These polyps occasionally contain areas of true metaplasia. In contrast to the surrounding intestinal epithelium, the neoplastic cells do not express Cdx2 from the remaining allele. These results suggest that Cdx2 mutation is the primary event in the genesis of some intestinal tumours.