Consanguinity Protecting Effect Against Breast Cancer among Tunisian Women: Analysis of BRCA1 Haplotypes.

The purpose of this study is to assess the effect of consanguinity on breast cancer incidence in Tunisia. We conducted a case-control study to evaluate the involvement of heterozygote and homozygote haplotypes of BRCA1 gene SNPs according to consanguinity among 40 cases of familial breast cancer, 46 cases with sporadic breast cancer and 34 healthy controls. We showed significant difference in consanguinity rate between breast cancer patients versus healthy controls P = 0.001. Distribution of homozygous BRCA1 haplotypes among healthy women versus breast cancer patients was significantly different; p=0.02. Parental consanguinity seems to protect against breast cancer in the Tunisian population.

Wild-type genotypes of BRCA1 gene SNPs combined with micro-RNA over-expression in mammary tissue leading to familial breast cancer with an increased risk of distant metastases' occurrence.

Germ line deleterious mutations of BRCA1 gene are not the unique factor that could inactivate BRCA1 protein which leads to familial breast cancer onset with distant metastases' occurrence. The present research explores the role that could be assigned to BRCA1 SNPs to inactivate BRCA1 protein and therefore to the occurrence of familial breast cancer with an increased risk of distant metastases' occurrence. The presence or the absence of BRCA1 protein was first analyzed by applying the immunohistochemistry technique to the tumors with sporadic and familial breast cancer. Then, a case-control study was conducted including 40 patients with familial breast cancer, 46 ones with sporadic breast cancer and 34 healthy controls based on the genotyping of nine BRCA1 SNPs (c.442.58delT, c.2082C>T, c.2311T>C, c.2612C>T, c.3113A>G, c.3119G>A, c.3548A>G, c.4308T>C and 4837A>G) via direct sequencing. Finally, the functional role that could be assigned to these SNPs was focused upon. miRbase site was used as a bioinformatics tool to predict potential micro-RNAs (miRs) targeting SNPs that are associated with familial breast cancer according to the results of this research. These predicted miRs were confirmed by Q-PCR analysis and correlated with BRCA1 protein expression among patients along with potential distant metastases. Clinical outcome showed that distant metastasis concerned 45 % of familial breast cancer patients and 19.5 % with sporadic breast cancer. Analysis of BRCA1 protein expression revealed a negative staining among 46.6 % of familial breast cancer patients and only 16.6 % within sporadic breast cancer ones. The association of four variants was identified within BRCA1 gene (c.442.58 delT, c.2311T>C, c.2612C>T and c.4308T>C) to familial breast cancer across their wild genotypes. miR-1179 was selected as potential miR that targets the region of BRCA1 mRNA containing the c.2311T>C variant within the TT genotype. The expression of miR-1179 was significantly associated with familial breast cancer patients without BRCA1 deleterious mutations compared to those with sporadic breast cancer according to TT genotype along with BRCA1 negative staining and according to the occurrence of distant metastases. Combination between TT genotype of c.2311T>C and miR-1179 over-expression could generate a lack of BRCA1 protein leading to a high risk of familial breast cancer with distant metastases.

CD99 and HLA-II immunostaining in breast cancer tissue and their correlation with lymph node metastasis.

In an attempt to better unfold the antitumor immune response and invasion strategies perused by tumor cells, markers such as CD99 and HLA-II have been stained in breast tumors, some of them turned out to be important for prognosis and its outcome. CD99 is involved in the intracellular transport of HLA-II proteins. The expression of HLA-II and CD99 molecules has been demonstrated in a broader range of neoplastic tissues, including some epithelial tumors. In the present work, we stained CD99 and HLA-II in breast malignant and non-malignant tissues sections obtained from biopsies resected surgically from 80 Tunisian women. Data implied that CD99 marks malignant tissue significantly as compared to non-malignant breast tissue. HLA-II staining allowed determining the correlation between breast cancer and HLA-II with cytoplasmic localization. CD99 and HLA-II immunostaining was also examined in correlation with two of the most important breast cancer prognostication in routine clinical practice, the lymph node stage and the histological assessment. Results let suggest that CD99(+)HLA-II(-) is a marker of worst prognostic since this phenotype is strongly linked to lymph node metastasis in breast cancer.

Hereditary breast cancer in Middle Eastern and North African (MENA) populations: identification of novel, recurrent and founder BRCA1 mutations in the Tunisian population.

Germ-line mutations in BRCA1 breast cancer susceptibility gene account for a large proportion of hereditary breast cancer families and show considerable ethnic and geographical variations. The contribution of BRCA1 mutations to hereditary breast cancer has not yet been thoroughly investigated in Middle Eastern and North African populations. In this study, 16 Tunisian high-risk breast cancer families were screened for germline mutations in the entire BRCA1 coding region and exon-intron boundaries using direct sequencing. Six families were found to carry BRCA1 mutations with a prevalence of 37.5%. Four different deleterious mutations were detected. Three truncating mutations were previously described: c.798_799delTT (916 delTT), c.3331_3334delCAAG (3450 delCAAG), c.5266dupC (5382 insC) and one splice site mutation which seems to be specific to the Tunisian population: c.212 + 2insG (IVS5 + 2insG). We also identified 15 variants of unknown clinical significance. The c.798_799delTT mutation occurred at an 18% frequency and was shared by three apparently unrelated families. Analyzing five microsatellite markers in and flanking the BRCA1 locus showed a common haplotype associated with this mutation. This suggests that the c.798_799delTT mutation is a Tunisian founder mutation. Our findings indicate that the Tunisian population has a spectrum of prevalent BRCA1 mutations, some of which appear as recurrent and founding mutations.

Are HLA DQB1 alleles correlated with breast cancer histopronostic parameters in Tunisia?

BACKGROUND: Tumor cells express surface structures different from normal cells. These structures may be recognized by the immune system, which ensure anti-tumoral surveillance. Antigenic presentation requires HLA molecules role. Since, these molecules are encoded by a high polymorphic system, immune response can be modulated according to HLA genotype. So, HLA polymorphism could be correlated with tumor escape from anti-tumor immunosurveillance. AIM: We have aimed to search for possible associations between HLA DQB1 alleles and the histoprognostical parameters in breast cancer in the Tunisian population. METHODS: DQB1 alleles were determined by PCR-SSO molecular typing in 100 healthy matched and unrelated Tunisian female and 87 Tunisian women with breast cancer. RESULTS: Allelic distribution between the two studied groups showed no significant associations between this locus and the occurrence, the EE grade and the lymph node invasion of breast cancer in the Tunisian population. CONCLUSION: This result may be explained by the fact that cancer is a multifactoral disease due to several interacting factors that might change from one population to another.

[Polymorphism of the mitochondrial microsatellite 303-315 in breast cancer in Tunisia]. / Polymorphisme du microsatellite mitochondrial 303-315 dans le cancer du sein en Tunisie.

AIM: The aim of this work was to study the correlation between the mitochondrial microsatellite, situated between the nucleotides 303 and 315 of the mitochondrial genome and the breast cancer in Tunisia. MATERIALS AND METHODS: We have analyzed, by PCR-sequencing, the polymorphism of a mitochondrial microsatellite in 40 Tunisian patients and 39 healthy Tunisian donors. Comparisons of epidemiologic and sequences data were performed by chi-2 test. RESULTS: We have revealed, for this mitochondrial microsatellite, seven different haplotypes in patients and five different haplotypes in controls. The haplotypic distribution was not significant between patients and controls but a negative association between one of these haplotypes (309+C 315+C) and the lymph node invasion was found. CONCLUSION: The haplotype 309+C 315+C is negatively correlated with lymph node invasion of breast cancer in Tunisia.

[Penetrance of BRCA1 gene mutation and DNA mitochondrial in Tunisian breast cancer occurrence]. / Pénétrance des mutations du gène BRCA1 et de l'ADN mitochondrial dans l'occurrence du cancer du sein en Tunisie.

AIM: The aim of this study is to evaluate the implication of BRCA1 gene and the mitochondrial micro satellite (situated between 303 and 315 positions) mutations in the occurrence of breast cancer in Tunisia. METHODS: Nine Tunisian patients with hereditary breast cancer have been analyzed. For each patient, total genomic DNA was extracted and used as a template for the amplification of 24 exons of the BRCA1 gene and an hyper variable mitochondrial region. The obtained products were purified and automatically sequenced. RESULTS: The results revealed five types of mutations for the micro satellite situated between the 303 and 315 positions and two deleterious BRCA1 mutations for two unrelated patients which present the same mitochondrial mutation (315.insC) suggesting his implication in the modulation of the BRCA1 deleterious mutations penetrance.

Contribution of the BRCA1 and BRCA2 mutations to breast cancer in Tunisia.

Hereditary breast cancer accounts for 3-8% of all breast cancers, with mutations in the BRCA1 and BRCA2 genes responsible for up to 30% of these. To investigate the prevalence of BRCA1 and BRCA2 gene mutations in breast cancer patients with affected relatives in Tunisia, we studied 36 patients who had at least one first degree relative with breast and/or ovarian cancer Thirty-four 34 patients were suggestive of the BRCA1 mutation and two were suggestive of the BRCA2 mutation, based on the presence of male breast cancer detected in their corresponding pedigrees. Four mutations in BRCA1 were detected, including a novel frame-shift mutation (c.211dupA) in two unrelated patients and three other frameshift mutations--c.4041delAG, c.2551delG and c.5266dupC. Our study is the first to describe the c.5266dupC mutation in a non-Jewish Ashkenazi population. Two frameshift mutations (c.1309del4 and c.5682insA) were observed in BRCA2. Nineteen percent (7/36) of the familial cases had deleterious mutations of the BRCA1 or BRCA2 genes. Almost all patients with deleterious mutations of BRCA1 reported a family history of breast and/or ovarian cancer in the index case or in their relatives. Our data are the first to contribute to information on the mutation spectrum of BRCA genes in Tunisia, and we give a recommendation for improving clinical genetic testing policy.

Immunolocalization of BRCA1 protein in tumor breast tissue: prescreening of BRCA1 mutation in Tunisian patients with hereditary breast cancer?

BRCA1 is a tumor suppressor gene which is inactivated by mutation in familial breast and ovarian cancers. Over 300 different disease causing germ-line mutations have been described; 60% are unique to an individual family. This diversity and the large size of the gene lead us to search for a prescreening method for BRCA1 mutations. Since BRCA1 is a nuclear protein in normal cells, but reported by some authors to be cytoplasmic in breast tumor cells of patients with BRCA1 mutation, we evaluated immunohistochemistry as a prescreening technique to identify BRCA1 mutations in patients with familial presentation of breast cancer. Using a monoclonal antibody against the carboxy-terminal region of BRCA1, we performed immunohistochemistry on 18 tumor samples from patients with hereditary breast cancer. Cytoplasmic staining of BRCA1 was observed in 10 cases. Of the 18 tumors, 12 (66%) showed either BRCA mutation or BRCA1 accumulation or both, indicating that BRCA1 function might be lost in breast tumor cells not only through mutation, but also via abnormal cytoplasmic location. The immunohistochemical test used in this study would not be efficient as a pre-screening method of deleterious mutations, but it appeared useful to investigate tumor physiology.

Do we know all there is to know about Familial Adenomatous Polyposis?

Familial Adenomatous Polyposis (FAP) and Attenuated FAP (AFAP) are caused by a germline mutation in the Adenomatous polyposis coli (APC) gene. Recently, a new pathway characterized by a biallelic mutation in the MYH gene, with a recessive model of inheritance was discovered for this inherited syndrome. This report describes a Tunisian patient with an attenuated FAP phenotype, presenting seven colon polyps and an adenocarcinoma but no detectable germline mutations in the FAP target genes. A well known somatic mutation was found in the APC mutation cluster region (MCR). This case shows that further studies are needed to fully understand all the pathways of the FAP syndrome.

HLA class II polymorphism: protective or risk factors to breast cancer in Tunisia?

HLA system plays a key role in the tumor cells' escape from immune surveillance. Herein is the first report on the correlation of the susceptibility to breast cancer with HLA class II markers in Tunisia. Molecular typing of HLA-DRB1 and -DQB1 loci was undertaken for 70 Tunisian female patients. Comparison of allele and haplotype distribution between patients and 70 female control subjects reveals a negative association between HLADRB1* 07-DQB1*02 and the incidence of breast cancer in the Tunisian population.