Colorectal Cancer and Genetic Alterations in the Wnt Pathway

Oncogene (2006) 25, 7531–7537 & 2006 Nature Publishing Group All rights reserved 0950-9232/06 $30.00 www.nature.com/onc REVIEW Colorectal cancer and genetic alterations in the Wnt pathway S Segditsas and I Tomlinson Molecular and Population Genetics Laboratory, London Research Institute, Cancer Research UK, London, UK In colorectal tumours, Wnt pathway genetics continues to other Wnt pathway components in benign and malig- be dominated by mutations in the adenomatous polyposis nant colorectal tumours. The functional consequences coli (APC) gene. Germline mutations cause familial of these changes are mentioned briefly, but many are adenomatous polyposis and at least two-thirds of sporadic covered more comprehensively by other papers in this colorectal tumours also acquire APC mutations, quite volume. In order to minimize repetition in this issue, we possibly as the initiating events in tumorigenesis. These have not included a figure outlining the Wnt pathway. mutations almost always cause loss of the C-terminal Please refer to other papers if needed. functions of the APC protein – probably involved in microtubule binding, cell polarity and chromosome segregation – and deletion of the SAMP repeats that are important for binding to axin and formation of the APC mutation spectrum beta-catenin phosphorylation complex. The truncated APC APC proteins are, in general, stable and almost certainly Although germline mutations only account for 1% retain some activity in beta-catenin binding. The ‘two hits’ of all colorectal cancer cases, they almost invariably at APC are coselected so as to produce an optimal cause a FAP phenotype. This condition is typified by activation of Wnt signalling (just-right hypothesis). In a hundreds or thousands of colorectal adenomas, one or minority of colorectal tumours, Wnt activation can occur more of which usually progresses to carcinoma by the through mutations that affect phosphorylation sites within fourth decade if the patient is untreated (Galiatsatos and exon 3 of beta-catenin, causing protein stabilization. In Foulkes, 2006). The great majority of FAP-associated APC other tumours, epigenetic transcriptional silencing or mutations lead to a truncated protein. Frameshift mutation of the secreted frizzled-related proteins may mutations outnumber nonsense changes by about two- modulate Wnt levels. Mutations in the Wnt components to-one, and the most common nonsense changes are AXIN1, AXIN2 and TCF4 have been found in micro- C>T mutations, probably resulting from spontaneous et al satellite-unstable colon cancers, but it is not clear in every deamination of 5-methylcytosine (Laurent-Puig ., case whether these changes are functional. Therapeutic 1998). In addition to protein-truncating mutations, modulation of the Wnt pathway remains an attractive deletions of the whole gene or exon(s) have been et al therapeutic possibility for colorectal carcinomas. reported in about 5% of FAP cases (Sieber ., 2002). Oncogene (2006) 25, 7531–7537. doi:10.1038/sj.onc.1210059 The protein-truncating mutations can occur anywhere in the APC gene, but the classical phenotype of more Keywords: APC; beta-catenin; ‘just right’ than 100 adenomas is associated with mutations between codons 178 and 309 and between codons 409 and 1580, corresponding to exons 5–8, 9–14 and the first half of the large final exon 15 (Fearnhead et al., 2001). Frameshift mutations at codons 1061 and 1309 are Introduction particularly common in western European populations; both of these regions contain short repeats, and a degree It is over 15 years since the adenomatous polyposis coli of hypermutation, plus genetic drift, probably accounts (APC) gene was found to be mutated in familial for their relatively high frequency. From the persepec- adenomatous polyposis (FAP) and, subsequently, in a tive of APC mutations, the most important functional very high proportion of sporadic colorectal tumours. Of domains of the APC gene appear to be the first serine course, the connection with the Wnt pathway was not alanine methionine proline (SAMP) (axin binding) known at that time, but it is arguable that without the repeat at codon 1580 (Smits et al., 1999) and the first, cloning of APC, this issue of Oncogene would not exist. second and third 20-amino acid repeats (20AARs) In this paper, we shall focus on the genetic and involved in beta-catenin binding and degradation. The epigenetic changes that occur in APC, beta-catenin and great majority of pathogenic APC mutations truncate the protein before the first SAMP repeat and leave a stable, truncated protein that encodes 0–3 20AARs Correspondence: Professor I Tomlinson, Molecular and Population Genetics Laboratory, London Research Institute, Cancer Research (Figure 1). UK, 44, Lincoln’s Inn Fields, London WC2A3PX, UK. In colorectal tumours from FAP patients, the E-mail: [email protected] germline wild-type allele either undergoes loss of Colorectal cancer and genetic alterations in the Wnt pathway S Segditsas and I Tomlinson 7532 Heptad repeats 15aa repeats SAMP repeats Basic domain (dimerisation) (β-catenin binding) (axin binding) (microtubule binding) 6 57 453 767 1020 1169 1266 1495 2033 2200 2400 2560 2843 1379 1581 Armadillo repeats 20aa repeats EB1 and HDLG (β-catening binding/degradation) binding sites a Gl mut allele Gl wt allele b Gl mut allele Gl wt allele c Gl mut allele Gl wt allele Figure 1 The ‘just-right’ model. The figure shows the multiple domains of the APC protein and the correlation between the position of the germline (gl) mutation and that of the somatic mutation. (a) Germline mutations between the first and the second 20AAR are associated with LOH. (b) Germline mutations before the first 20AAR are associated with somatic mutations between the second and third 20AAR. (c) Germline mutations after the second 20AAR are associated with somatic mutations before the first 20AAR. heterozygosity (LOH) or acquires a protein-truncating one-third to one-half of tumours, a frequency lower mutation. In general, these are thought to be sponta- than that at many other bona fide tumour suppressor neous events and APC is often cited as a classical loci. tumour-suppressor gene, both alleles of which must be The reason for the MCR and relatively low frequency inactivated for tumorigenesis to begin. It has been of LOH at APC came from studies of FAP (Lamlum argued, however, that the number of adenomas in the et al., 1999). It was found that LOH is strongly FAP colon is too great for each one to have acquired a associated with germline mutations between the first ‘second hit’, especially if FAP adenomas are frequently and second 20AAR (codons 1285–1379). Germline polyclonal. The validity of such calculations is inhe- mutations before codon 1280 are associated with rently difficult to confirm or refute, since they depend on somatic mutations between the second and third the one hand on estimations of the susceptible cell 20AAR (codons 1400 and 1495); and germline muta- population and, on the other, on finding ‘second hits’ in tions after codon 1400 are associated with somatic almost all early tumours, a technically very demanding mutations before codon 1280 (Lamlum et al., 1999; task. For the purposes of this paper, it will be assumed Albuquerque et al., 2002; Crabtree et al., 2003) that FAP adenomas do have (at least) two hits, although (Figure 1). Most tumours end up with APC alleles that the possibility that very early lesions can grow without a encode a total of two 20AARs. Similar associations exist somatic APC mutation must be borne in mind. for sporadic colorectal cancers. This association has The somatic mutations that occur in sporadic colo- been proposed to cause an optimal level of Wnt rectal tumours are similar in spectrum and location to signalling/beta-catenin activation (Lamlum et al., 1999; the somatic mutations in FAP polyps. Albuquerque et al., 2002). Whether this level is the maximum level is unclear, since APC protein truncated at codon 1309 has been shown to have increased Selective constraints on APC mutations in inherited and stability compared with other mutants (Dihlmann sporadic tumours et al., 1999) and many truncated APC proteins retain some ability to bind and, perhaps, degrade beta-catenin In recent years, it has become established that the (Rubinfeld et al., 1997). Absence of APC appears to be somatic APC mutation in FAP tumours – and sporadic selectively disadvantageous (Schneikert and Behrens, colorectal carcinomas – is not simply an event that 2006). Whatever the case, it is clear that selective removes function of the protein encoded by the germline constraints act on colorectal tumours such that some wild-type allele. It had been known for some years that combinations of APC mutations provide a superior somatic APC mutations tend to occur in a restricted growth advantage for the tumour cell. region of the gene (mutation cluster region (MCR)) that The ‘1st hit-2nd hit’ association at APC has been lay approximately between codons 1250 and 1450 dubbed the ‘just right’ hypothesis (Albuquerque et al., (Miyoshi et al., 1992). LOH occurred in about 2002). This is a convenient term that encapsulates the Oncogene Colorectal cancer and genetic alterations in the Wnt pathway S Segditsas and I Tomlinson 7533 notion well – and it will be used below – although we did (i) Loss of germline wt + truncating mutation not use such a term originally, because the selective constraints are evidently not so rigid that deviation from the total of two 20AARs cannot be tolerated in a Phenotype substantial minority of lesions (Lamlum et al., 1999; Crabtree et al., 2003). Further studies have shown that a LOH as first somatic event the ‘just right’ model applies to extracolonic lesions in FAP, but that the combination of mutations is different X (Lamlum et al., 1999; Groves et al., 2002). In desmoids LOH Allelotype + + or and uppergastrointestinal tumours, LOH is associated with germline APC mutations between the second and third 20AARs, and the optimum mutant proteins X appear to encode a total of three or four 20AARs.

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