Germ line BRCA1 & BRCA2 mutations in Greek breast/ovarian cancer families: 5382insC is the most frequent mutation observed.

BRCA1 and BRCA2 genes were screened for loss-of-function mutations in a series of 85 patients having at least one first- or second-degree relative affected by breast and/or ovarian cancer. All BRCA1 exons and BRCA2 exons 10 and 11 were screened with a combination of methods including SSCP, PTT and direct sequencing. We have found disease-associated mutations in 14 families (16.5%), eleven in BRCA1 and three in BRCA2. The known founder mutation 5382insC of BRCA1 was identified in seven unrelated families. The other mutations identified include the non-sense R1751X, the splice junction variant 5586G>A of BRCA1 and three frameshifts, 2024del5, 3034del4, and 6631del5, of BRCA2. Nine out of these 14 families had a family history of three or more breast/ovarian cancer cases. A large number of polymorphic or unclassified variants is also reported. Combined with our previously published data 5382insC was found in nine out of 20 families (45%), suggesting that this mutation may represent a common founder mutation in the Greek population.

Novel mutations of the APC gene in familial adenomatous polyposis in Greek patients.

Familial adenomatous polyposis (FAP), a premalignant clinical entity inherited as an autosomal dominant trait, is characterized by the development thousands of adenomatous polyps of the colorectum during the 2nd and 3rd decade of life. Approximately 80% of patients with FAP harbor truncating germline mutations in the adenomatous polyposis coli (APC) tumor suppressor gene. We tested 24 members of six Greek families. All patients had the FAP phenotype, and one patient had an extracolonic tumor (medulloblastoma). Our method for testing was the polymerase chain reaction (PCR) amplification from genomic DNA extracted from whole blood, followed by automated DNA sequencing. Two novel truncating mutations (2601delGA and R923X) and three already-known mutations (R876X, Q1045X, and D1822V) were found. Other polymorphisms were also found. We identified the inactivating APC mutation in 12 of 13 of our FAP patients. Our results suggest that PCR sequencing is a reliable method for screening the APC gene for germline mutations.

Genetic alterations of the APC gene in familial adenomatous polyposis patients of the hellenic group for the study of colorectal cancer.

Familial Adenomatous Polyposis (FAP)-a premalignant clinical entity inherited as an autosomal dominant trait-is characterized by the development of hundreds to thousands of adenomatous polyps of the colorectum during the second and third decade of life. Approximately 80% of the FAP patients harbour truncating germ-line mutations in the APC tumor suppressor gene (Adenomatous Polyposis Coli). We tested 48 members from 9 families. Two novel truncating mutations were identified-2601delGA, R923X--and five already known mutations R564X, R876X, Q1045X, 3927-3931delAAAGA and D1822V were found. Our method for testing was PCR amplification from genomic DNA extracted from whole blood, followed by automated DNA sequencing.

Documentation of Legionella pneumophila and Mycobacterium tuberculosis co-existence in a patient with acute respiratory distress syndrome.

BACKGROUND: Bacterial lung infections are common causes of ARDS and, despite intensive research for decades, the mortality rate remains very high. Only two reports suggest the co-existence of Legionnaires' disease and pulmonary tuberculosis based mainly on clinical presentation and serologic results for Legionella and positive cultures for Mycobacterium tuberculosis (M. tuberculosis). MATERIALS AND METHODS: A variety of specimens from a 61-year-old man was used for detection of Legionella pneumophila (L. pneumophila) and M. tuberculosis by PCR. Further identification of the pathogens was carried out by sequence analysis. RESULTS: L. pneumophila region mip was detected in bronchial washings, bronchoalveolar lavage and urine specimens of the patient. M. tuberculosis regions IS6110 and mtp40 were detected in endo-bronchial secretions and bronchoalveolar lavage. CONCLUSION: By using polymerase chain reaction (PCR) and DNA sequencing we documented L. pneumophila and M. tuberculosis co-existence, in multiple specimens of a patient presenting with acute respiratory distress syndrome (ARDS). Furthermore, the efficacy of the specific antibiotic treatment, based on the PCR results, suggest the co-existence of these two pathogens.

hMSH2 is the most commonly mutated MMR gene in a cohort of Greek HNPCC patients.

Germline mutations in genes encoding proteins involved in DNA mismatch repair are responsible for the autosomal dominantly inherited cancer predisposition syndrome hereditary nonpolyposis colorectal cancer (HNPCC). We describe here analysis of hMLH1 and hMSH2 in nine Greek families referred to our centre for HNPCC. A unique disease-causing mutation has been identified in seven out of nine (78%) families. The types of mutations identified are nonsense (five out of seven) (hMLH1: E557X, R226X; hMSH2: Q158X, R359X and R711X), a 2 bp deletion (hMSH2 1704_1705delAG) and a 2.2 kb Alu-mediated deletion encompassing exon 3 of the hMSH2 gene. The majority of mutations identified in this cohort are found in hMSH2 (77.7%). Furthermore, four of the mutations identified are novel. Finally, a number of novel benign variations were observed in both genes. This is the first report of HNPCC analysis in the Greek population, further underscoring the differences observed in the various geographic populations.

A modified mutagenic PCR-RFLP method for K-ras codon 12 and 13 mutations detection in NSCLC patients.

Evidence from many investigators has shown that mutations in the first exon of K- ras gene occur at elevated frequencies in lung, pancreatic and colon carcinoma and seem to be of prognostic importance. The aim of this study was to develop an effective method for the detection of K- ras mutations in codons 12 and 13 in non-small-cell lung cancer (NSCLC) patients in order to investigate correlation with clinical outcome. DNA was extracted from tumour and neighbouring non-neoplastic lung tissues from 70 patients and screened for codon 12 and 13 mutations. We applied a mutagenic PCR-restriction fragment length polymorphism for both codon 12 and 13 mutation detection. Codon 12 mutation was identified in 20% of NSCLC patients, whereas no codon 13 mutation was detected. As expected, the respective non-neoplastic tissues exhibited no mutations. We observed an increased codon 12 mutation prevalence in adenocarcinoma comparing to other types of carcinomas. Follow-up for 29 patients with a mean time of 12 months indicates an increased relapse rate in NSCLC patients with the K- ras codon 12 mutation. Furthermore, a trend towards increased percentage of mutant samples was observed in the advanced stage group of patients. We provide evidence that our approach is a fast and reliable method for screening K- ras exon 1 mutations in tumour samples from NSCLC patients.