A-1012G promoter polymorphism of vitamin D receptor gene is associated with psoriasis risk and lower allele-specific expression.

Psoriasis is caused by a combination of genetic, immunologic, and environmental factors. The vitamin D receptor (VDR) is involved in antiproliferative and prodifferentiation pathways in keratinocytes and exerts immunosuppressive effects. We aimed to investigate possible associations between VDR polymorphisms and psoriasis susceptibility and to evaluate functional effects of potential psoriasis-associated polymorphisms. We genotyped 108 patients with psoriasis and 268 healthy controls at 5 VDR polymorphisms (A-1012G, FokI, BsmI, ApaI, and TaqI) by TaqMan allelic-discrimination real-time polymerase chain reaction. We found a significant increased overall risk of psoriasis for the VDR A-1012G promoter polymorphism (odds ratio [OR]=2.43, 95% confidence interval [CI]: 1.15-5.13; p=0.05). A significant higher frequency (p=0.035) of the A allele was found in psoriatic cases compared with controls. In a case-case analysis, a statistically significant association between A-1012G and family history emerged (p=0.033). Furthermore, a significant association of A-1012G risk genotypes with a lower expression of VDR mRNA emerged (p=0.0028). Our data show that VDR promoter A-1012G polymorphism is associated with psoriasis risk and suggest that this polymorphism may modulate psoriasis risk by affecting VDR expression.

Association between the BRCA2 N372H variant and male breast cancer risk: a population-based case-control study in Tuscany, Central Italy.

BACKGROUND: Male breast cancer (MBC) is a rare disease and little is known about its aetiology. Germ-line mutations of BRCA2 and, at lower frequency, of BRCA1 are implicated in a relatively small proportion of MBC cases. Common polymorphic variants in BRCA1 and BRCA2 genes may represent breast cancer (BC) susceptibility alleles and could be associated with a modestly increased risk of MBC at population level. Considering the relevant role of BRCA2 in MBC, we investigated whether the BRCA2 N372H variant, representing the only common non-synonymous polymorphism in BRCA2, might modulate the risk of BC in male populations. METHODS: A case-control study was performed comparing a population-based series of 99 MBC cases, characterized for BRCA1 and BRCA2 mutations, with 261 male population controls, all residing in Tuscany, Central Italy. All MBC cases and controls were genotyped for the BRCA2 N372H allele by TaqMan allelic discrimination assays. To evaluate the genotype specific risk of the BRCA2 N372H variant, MBC carriers of germ-line BRCA1/2 mutations were excluded from the analyses. RESULTS: No association emerged in univariate and age-adjusted analyses. Age-specific analyses suggested an increased risk for the HH homozygous genotype in subjects younger than 60 years. A statistically significant interaction emerged between this genotype and age (p = 0.032). When analyses were restricted to MBC cases enrolled in the first 4 years following diagnosis, a recessive model showed a significantly increased risk of MBC in HH subjects younger than 60 years (OR = 5.63; 95% CI = 1.70;18.61). CONCLUSION: Overall, our findings, although based on a relatively small series, suggest that the BRCA2 HH homozygous genotype might be positively associated with an increased risk of MBC in men younger than 60 years.

A complex pattern of mutations and abnormal splicing of Smad4 is present in thyroid tumours.

Sensitivity to transforming growth factor-beta is impaired in thyroid tumours. Similar to Mad -- Mother Against Decapentaplegic-(Smad)4 is frequently altered in cancers, but its involvement in this system is unknown. We analysed 56 thyroid tumours of various histotypes for Smad4 mutations by PCR-SSCP and sequencing, linking them to Smad4 reactivity as examined by immunohistochemistry (IHC), and 29 of them also for abnormalities in RNA expression due to alternative splicing. In all, 15/56 cases (27%), both benign and malignant lesions, harbour alterations of Smad4 coding sequence. We found several novel intragenic mutations (13 missense, two silent, one frameshift and one large insertion-deletion), with high incidence in the linker region. A subset of mutated tumours failed to express Smad4 protein by IHC. We have also detected four alternatively spliced tumour-associated Smad4 isoforms, lacking portions of the linker region, and three more due to unreported internal exon-exon rearrangements. Smad4 is both frequently mutated and deregulated by aberrant splicing in thyroid tumours and these alterations may contribute as an early event to thyroid tumorigenesis.

Different expressivity of BRCA1 and BRCA2: analysis of 179 Italian pedigrees with identified mutation.

Mutations in BRCA1 and BRCA2 show different expressivity with respect to cancer risk, and allelic heterogeneity may be present in both genes. We collected 179 pedigrees with identified germline mutation (104 BRCA1 and 75 BRCA2), ascertained in six collaborating centers of the Italian Consortium for Hereditary Breast and Ovarian Cancer. Significant heterogeneity was detected for several variables, and a logistic regression model including age of diagnosis in the proband, presence of ovarian cancer in the family, presence of prostate or pancreatic cancer in the family, and presence of male breast cancer in the family proved to be effective in predicting the presence of a mutation in a gene rather than the other. Excess of familial aggregation of both breast and ovarian cancer was observed in both genes. Proportion of ovarian cancer was increased in the 5' portion of BRCA1, and presence of prostate or pancreatic cancer in a family was correlated with presence of ovarian cancer in BRCA2.

BRCA1 and BRCA2 mutation status and tumor characteristics in male breast cancer: a population-based study in Italy.

To investigate at the population level the impact of BRCA1/BRCA2 gene alterations in male breast cancer, we analyzed a population-based series of 25 male breast cancer cases from Florence, Central Italy. We combined mutational screening with the study of germ-line allele transcript levels and of tumor-associated losses of heterozygosity. Screening by protein truncation test and single-strand conformational polymorphism assay, followed by sequencing, revealed 4 pathogenetic mutations (4 of 25 = 16%; 95% confidence interval, 5-37%), 1 in BRCA1 and 3 in BRCA2, including mutations recurring in Central Italy (BRCA1 3345delAG and BRCA2 6696delTC). The a priori probability of carrying a mutation, estimated using BRCAPRO software, showed a good agreement between expected and observed mutations (14% versus 16%). A 7-fold association between germ-line mutations and family history of breast-ovarian cancer emerged. To investigate associations between BRCA1/BRCA2 status and clinicopathological characteristics, we analyzed the histopathological and immunophenotypic parameters of the tumors. A significant association emerged between mutation carrier status and high histological grade (P = 0.02). Furthermore, one BRCA2 carrier was affected with Paget's disease, an extremely rare male breast cancer histotype. Overall, BRCA1/2 mutations were observed to be strongly associated with positive c-erbB-2 immunostaining (P = 0.004). To evaluate germ-line allele expression, we used primer extension assays targeting frequent BRCA1 and BRCA2 polymorphisms. A BRCA2 allele transcript imbalance was found in one of four heterozygotes tested, all of them negative for germ-line mutations. BRCA1 transcript imbalances were not detected in nine heterozygotes analyzed. Losses of heterozygosity at one or more of nine loci in the BRCA2 region were found in 8 of 22 tumors tested. Interestingly, a case that was negative for BRCA1/BRCA2 germ-line mutations and that had a priori mutation probability <10% showed loss of heterozygosity at all three of the intragenic BRCA2 markers analyzed, which could be related to a somatic involvement of BRCA2. No losses of heterozygosity were detected at BRCA1. In conclusion, constitutional BRCA1/BRCA2 mutations accounted for 16% of the male breast cancer cases in this area of Central Italy. The detection of a BRCA2 germ-line transcript imbalance and of a somatic loss of BRCA2 among the cases that resulted negative for germ-line mutations suggests a role of this gene more relevant than indicated by conventional mutational analysis. A distinct pattern of characteristics indicative of aggressive behavior, including high-grade and c-erbB-2 expression, was evident in tumors from germ-line BRCA2 mutation carriers. This suggests that phenotypic characteristics may contribute to the identification of hereditary BRCA2-related male breast cancers and that these tumors might share a unique molecular pathway of cancer progression.

Subcellular localization of the BRCA1 gene product in mitotic cells.

The product of the hereditary breast cancer susceptibility gene BRCA1 is a multifunctional protein involved in the maintenance of genomic integrity, in transcriptional coactivation, and in the control of cell growth. BRCA1-deficient cells manifest chromosomal instability. During mitosis, BRCA1 is known to interact with gamma-tubulin in the centrosomes, key elements of the mitotic spindle. Using confocal microscopy and immunogold electron microscopy, we investigated the distribution of endogenous BRCA1 relative to mitotic spindle markers in breast cancer cells. By confocal analysis, BRCA1 and beta-tubulin colocalized to microtubules of the mitotic spindle and to the centrosomes. Immunogold electron microscopy confirmed these results and further revealed that BRCA1 and alpha-tubulin codistributed to the walls of the centrioles and to pericentriolar fibers at centrosomes. During chromatid segregation, codistribution was also detected along individual spindle microtubules and at sites of insertion of microtubules on chromosomes. At cytokinesis, BRCA1 and alpha-tubulin codistributed to the midbody. Coimmunoprecipitation supported the association of full-length BRCA1 with alpha- and beta-tubulin. These results are consistent with an involvement of BRCA1 in the dynamics of the mitotic spindle and in the segregation of duplicated chromosomes.

p53 expression is decreased in primary breast carcinomas with microsatellite instability.

p53 and p185 expression in primary breast cancer with microsatellite instability (MSI) is still largely unexplored. To investigate the relationship between these oncoproteins and the pathways of genomic instability, we examined 52 primary invasive breast cancers stratified by the presence and absence of MSI. We determined the status of eight microsatellite loci using radioactive and silver staining methods, and evaluated the immunohistochemical expression of p53 and p185 in a consecutive series of Italian cancer patients characterized by clinical-pathological and biological parameters. Nineteen cases (36.5%) were MSI-positive in at least two loci. p53 was expressed in 15 cases (28.8%) and p185 in eight (15.4%). MSI-positive tumors were inversely correlated with p53 expression (p = 0.0007); in addition, the percent of p53-expressing cells decreased as the number of MSI-positive loci increased. MSI-positive tumors were correlated with a larger tumor size (p = 0.04), lymph-node metastasis (p = 0.001), and advanced clinical stage (p = 0.0006). These data demonstrate the existence of two subsets of primary breast cancers: one characterized by MSI, the other by p53 expression. MSI-positive patients had a more advanced and/or aggressive disease.