Multiplex ligation-dependent probe amplification (MLPA) is a recently described method for detecting gross deletions or duplications of DNA sequences, aberrations which are commonly overlooked by standard diagnostic analysis. To determine the incidence of copy number variants in cancer predisposition genes from families in the Wessex region, we have analysed the hMLH1 and hMSH2 genes in patients with hereditary nonpolyposis colorectal cancer (HNPCC), BRCA1 and BRCA2 in families with hereditary breast/ovarian cancer (BRCA) and APC in patients with familial adenomatous polyposis coli (FAP). Hereditary nonpolyposis colorectal cancer (n ¼ 162) and FAP (n ¼ 74) probands were fully screened for small mutations, and cases for which no causative abnormality were found (HNPCC, n ¼ 122; FAP, n ¼ 24) were screened by MLPA. Complete or partial gene deletions were identified in seven cases for hMSH2 (5.7% of mutation-negative HNPCC; 4.3% of all HNPCC), no cases for hMLH1 and six cases for APC (25% of mutation negative FAP; 8% of all FAP). For BRCA1 and BRCA2, a partial mutation screen was performed and 136 mutation-negative cases were selected for MLPA. Five deletions and one duplication were found for BRCA1 (4.4% of mutation-negative BRCA cases) and one deletion for BRCA2 (0.7% of mutation-negative BRCA cases). Cost analysis indicates it is marginally more cost effective to perform MLPA prior to point mutation screening, but the main advantage gained by prescreening is a greatly reduced reporting time for the patients who are positive. These data demonstrate that dosage analysis is an essential component of genetic screening for cancer predisposition genes.

Familial adenomatous polyposis (FAP) is a dominantly inherited predisposition to the development of many hundreds to thousands of adenomatous polyps of the colon. The mean age of onset is around 15 years, symptoms may arise in the third decade, and the median age for the development of colonic cancer is 35-40 years. Prophylactic colectomy reduces the risk of death from colorectal cancer to such an extent that late sequelae such as upper gastrointestinal tumours have become the main cause of mortality in appropriately managed patients. The age at which colonic surveillance begins reflects the natural history of the disease. Onset of polyp formation and cancer in childhood is very unusual, but has recently been associated with a specific mutation at codon 1309 in exon 15 where a more severe phenotype is sometimes observed. The case histories of two families are reported in which there is childhood onset of polyps in the youngest generation and in one case a carcinoma, in whom mutations have been identified in exon 11 of the APC gene. Several other aVected relatives were diagnosed at ages ranging from 5-48 years, some already with a cancer at the time of first screening. Since the aim of screening for colonic polyps is prevention of colonic cancer, family members at risk should be oVered genetic assessment and direct mutation testing where this is possible, usually in the early teens. In the absence of a genetic test (the situation in about one third of families) or in a known gene carrier, annual colonoscopy examination is advised from the same age. Clinicians should take note of the family history and be prepared to consider much earlier intervention if symptoms occur in a child with a family history of FAP. Where childhood onset of polyps has occurred, other children at risk in the family must be oVered earlier genetic testing and endoscopic surveillance.

Nine new causative mutations and seven previously characterised mutations of the APC gene of patients with familial adenomatous polyposis (FAP) were analysed for any genotype-phenotype correlations. The only clear genotype-phenotype correlation found was between the position of the mutation site and the presence or absence of congenital hypertrophy of the retinal pigment epithelium (CHRPE). A more distal mutation site was associated with an earlier age of onset of symptoms and a larger number of colonic polyps, but a notable amount ofintrafamilial variation was observed. In 1993, an FAP register was established in Wessex as a two year pilot project with regional funding. Following a systematic screen of the APC gene of 30 unrelated people with FAP from this register, we report nine new causative mutations plus seven previously characterised mutations. In order to study genotype-phenotype effects and any intrafamilial variation in expression, the position of the mutation site in the proband of each pedigree, and in any other available affected family members, was correlated to three phenotypic features: CHRPE status, age at diagnosis of polyps, and number of polyps. Materials and methods PATIENTSAll patients tested were from the Wessex Polyposis Register. All available patients were examined by the same ophthalmologist using indirect ophthalmoscopy. Data concerning polyp number and age at diagnosis were retrieved from hospital notes. Molecular analysis was carried out on DNA extracted from circulating peripheral blood leucocytes. HETERODUPLEX ANALYSISGenomic DNA (100 ng) was amplified using the relevant primers and conditions listed by Groden et al.' The PCR products were heated to 95°C for three minutes then cooled to 37°C over a period of 30 minutes to form heteroduplexes before running 10 gl of sample on a 24 cm Hydrolink gel (AT Biochem). Heteroduplex bands were visualised by ethidium bromide staining.

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