Recurrent CDH1 mutations in families with hereditary diffuse gastric cancer are due to both independent mutational events and common ancestry. The presence of a founder mutation from Newfoundland is strongly supported.
Background: Mutations in the E-cadherin (CDH1) gene are a well documented cause of hereditary diffuse gastric cancer (HDGC). Development of evidence based guidelines for CDH1 screening for HDGC have been complicated by its rarity, variable penetrance, and lack of founder mutations. Methods: Forty three new gastric cancer (GC) families were ascertained from multiple sources. In 42 of these families at least one gastric cancer was pathologically confirmed to be a diffuse gastric cancer (DGC); the other family had intestinal type gastric cancers. Screening of the entire coding region of the CDH1 gene and all intron/exon boundaries was performed by bi-directional sequencing. Results: Novel mutations were found in 13 of the 42 DGC families (31% overall). Twelve of these mutations occur among the 25 families with multiple cases of gastric cancer and with pathologic confirmation of diffuse gastric cancer phenotype in at least one individual under the age of 50 years. The mutations found include small insertions and deletions, splice site mutations, and three non-conservative amino acid substitutions (A298T, W409R, and R732Q). All three missense mutations conferred loss of E-cadherin function in in vitro assays. Multiple cases of breast cancers including pathologically confirmed lobular breast cancers were observed both in mutation positive and negative families. Conclusion: Germline truncating CDH1 mutations are found in 48% of families with multiple cases of gastric cancer and at least one documented case of DGC in an individual under 50 years of age. We recommend that these criteria be used for selecting families for CDH1 mutational analysis.
E-cadherin is involved in the formation of cell-junctions and the maintenance of epithelial integrity. Direct evidence of E-cadherin mutations triggering tumorigenesis has come from the finding of inactivating germline mutations of the gene (CDH1) in hereditary diffuse gastric cancer (HDGC). We screened a series of 66 young gastric cancer probands for germline CDH1 mutations, and two novel missense alterations together with an intronic variant were identified. We then analysed the functional significance of the two exonic missense variants found here as well as a third germline missense variant that we previously identified in a HGDC family. cDNAs encoding either the wild-type protein or mutant forms of E-cadherin were stably transfected into CHO (Chinese hamster ovary) E-cadherin-negative cells. Transfected cell-lines were characterized in terms of aggregation, motility and invasion. We show that a proportion of apparently sporadic early-onset diffuse gastric carcinomas are associated with germline alterations of the E-cadherin gene. We also demonstrate that a proportion of missense variants are associated with significant functional consequences, suggesting that our cell model can be used as an adjunct in deciding on the potential pathogenic role of identified E-cadherin germline alterations.
Purpose: To identify germ line CDH1 mutations in hereditary diffuse gastric cancer (HDGC) families and develop guidelines for management of at risk individuals. Experimental Design: We ascertained 31 HDGC previously unreported families, including 10 isolated early-onset diffuse gastric cancer (DGC) cases. Screening for CDH1 germ line mutations was done by denaturing high-performance liquid chromatography and automated DNA sequencing. Results: We identified eight inactivating and one missense CDH1 germ line mutation. The missense mutation conferred in vitro loss of protein function. Two families had the previously described 1003C>T nonsense mutation. Haplotype analysis revealed this to be a recurrent and not a founder mutation. Thirty-six percent (5 of 14) of the families with a documented DGC diagnosed before the age of 50 and other cases of gastric cancer carried CDH1 germ line mutations. Two of10 isolated cases of DGC in individuals ages <35 years harbored CDH1germ line mutations. One mutation positive family was ascertained through a family history of lobular breast cancer (LBC) and another through an individual with both DGC and LBC. Occult DGC was identified in five of six prophylactic gastrectomies done on asymptomatic, endoscopically negative 1003C>T mutation carriers. Conclusions: In addition to families with a strong history of early-onset DGC, CDH1 mutation screening should be offered to isolated cases of DGC in individuals ages <35 years and for families with multiple cases of LBC, with any history of DGC or unspecified GI malignancies. Prophylactic gastrectomy is potentially a lifesaving procedure and clinical breast screening is recommended for asymptomatic mutation carriers.Gastric cancer is one of the three leading causes of cancer death worldwide (1). Although the incidence of gastric cancer in older patients is decreasing, in younger patients as well as in cases with familial clustering it remains stable, suggesting that genetic predisposition is an increasingly important risk factor for gastric cancer (2). In this respect, as few as 1% to 3% of all
Gastric cancer's (GC) incidence shows large geographic differences worldwide with the lowest rates occurring in most Western industrialized countries including the United States and the United Kingdom; in contrast, relatively high rates of GC occur in Japan, Korea, China, and South America, particularly Chile. The Laurén classification system classifies GC under two major histopathological variants: 1) an intestinal type and 2) a diffuse type. The intestinal type is more common in the general population, more likely to be sporadic and related to environmental factors such as diet, particularly salted fish and meat as well as smoked foods, cigarette smoking, and alcohol use. It exhibits components of glandular, solid, or intestinal architecture, as well as tubular structures. On the other hand, the diffuse type is more likely to have a primary genetic etiology, a subset of which, known as hereditary diffuse gastric cancer (HDGC), is due to the E-cadherin (CDH1) germline mutation. The diffuse type pathology is characterized by poorly cohesive clusters of cells which infiltrate the gastric wall, leading to its widespread thickening and rigidity of the gastric wall, known as linitis plastica. Helicobacter pylori infection is associated with risk for both the intestinal and diffuse varieties of gastric cancer. Germline truncating mutations of the CDH1 gene, which codes for the E-cadherin protein, were initially identified in three Maori families from New Zealand that were predisposed to diffuse GC. Since then, similar mutations have been described in more than 40 additional HDGC families of diverse ethnic backgrounds. It is noteworthy that two-thirds of HDGC families reported to date have proved negative for the CDH1 germline mutation. A number of candidate genes have been identified through analysis of the molecular biology of E-cadherin. Patients with evidence of the CDH1 germline mutation in the context of a family history of HDGC must be considered as candidates for prophylactic gastrectomy, given the extreme difficulty in its early diagnosis and its exceedingly poor prognosis when there is regional or distant spread. Specifically, the E-cadherin cytoplasmic tail interacts with catenins, assembling the cell-adhesion complex involved with E-cadherin mediated cell:cell adhesion. Beta-catenin and gamma-catenin compete for the same binding site on the E-cadherin cytoplasmic tail, directly linking the adhesion complex to the cytoskeleton through alpha-catenin. Beta-catenin gene (CTNNB1) mutations have been described predominantly in intestinal-type gastric cancers and CTNNB1 gene amplification and overexpression have recently been described in a mixed-type gastric cancer. This paper reviews the genetics of both intestinal and diffuse types of gastric carcinoma, their differential diagnosis, molecular genetics, pathology, and, when known, their mode of genetic transmission within families.
In Hereditary Diffuse Gastric Cancer syndrome, E-cadherin germline mutations of the missense type harbour significant functional consequences. In this study, we have characterised the effect of T340A, A617T, A634V and V832M E-cadherin germline missense mutations on cell morphology, motility and proliferation. Wild-type E-cadherin and A617T expressing cells have an epithelial-like morphology, with polarised cells migrating unidirectionally. T340A and A634V expressing cells, fibroblast-like, have a high motile phenotype. We show that this phenotype is dependent on an increased level of active RhoA. V832M expressing cells grow in piled-up structure of round cells, as an effect of the disturbance of the binding between alpha-catenin and beta-catenin. The destabilisation of the adhesion complex is shown to hamper the motile capabilities of these cells. We did not observe any effect of the E-cadherin mutations on cell proliferation. We show the existence of a genotype-phenotype correlation between different E-cadherin mutations and cell behaviour. However, we demonstrate that the ability of cells expressing the different E-cadherin mutations to invade is independent on their motile capabilities, providing evidence that motility is neither necessary nor sufficient for cells to invade. Our data give new insights into the understanding of the mechanisms linking invasion and E-cadherin mutations in diffuse gastric cancer.
In hereditary diffuse gastric cancer (HDGC), CDH1 germline gene alterations are causative events in 30% of the cases. In 20% of HDGC families, CDH1 germline mutations are of the missense type and the mutation carriers constitute a problem in terms of genetic counseling and surveillance. To access the pathogenic relevance of missense mutations, we have previously developed an in vitro method to functionally characterize them. Pathogenic E-cadherin missense mutants fail to aggregate and become more invasive, in comparison with cells expressing the wild-type (WT) protein. Herein, our aim was to develop a complementary method to unravel the pathogenic significance of E-cadherin missense mutations. We used cells stably expressing WT E-cadherin and seven HDGC-associated mutations (five intracellular and two extracellular) and studied by proximity ligation assays (PLA) how these mutants bind to fundamental regulators of E-cadherin function and trafficking. We focused our attention on the interaction with: p120, b-catenin, PIPKIc and Hakai. We showed that cytoplasmic E-cadherin mutations affect the interaction of one or more binding partners, compromising the E-cadherin stability at the plasma membrane and likely affecting the adhesion complex competence. In the present work, we demonstrated that the study of the interplay between E-cadherin and its binding partners, using PLA, is an easy, rapid, quantitative and highly reproducible technique that can be applied in routine labs to verify the pathogenicity of E-cadherin missense mutants for HDGC diagnosis, especially those located in the intracellular domain of the protein. Keywords: HDGC; E-cadherin; CDH1 mutations; E-cadherin trafficking; E-cadherin binding partners; diagnostic method INTRODUCTIONHereditary diffuse gastric cancer (HDGC) is an autosomal dominant cancer syndrome characterized by a high risk of developing diffuse gastric cancer 1-3 and lobular breast cancer 4-6 during life-time. CDH1 germline gene alterations (mutations or deletions), resulting in E-cadherin inactivation, are the only causative events described till now and were identified in approximately 30% of HDGC cases. 2,3,7 To date, 122 different germline mutations have been described in these families, 8 being the majority of them of the nonsense type, leading to alternative premature termination codons. 3 This type of CDH1 mutant transcripts is commonly downregulated by nonsensemediated decay leading to E-cadherin loss of function 9 and these patients are considered high-risk carriers and are counseled to perform prophylactic total gastrectomy. 10 In about 20% of HDGC families, carriers show CDH1 germline missense mutations 11 and, in contrast to truncating mutations, their pathogenic significance is not straightforward, therefore constituting a problem in terms of genetic counseling and surveillance.In 2004, Fitzgerald and Caldas 10 suggested that the significance of CDH1 missense mutations should be assessed in at least four affected members within a HDGC family in combination with functional and
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