Case report: Compound heterozygous variants detected by next-generation sequencing in a Tunisian child with ataxia-telangiectasia.

Ataxia-telangiectasia (A-T) is an autosomal recessive primary immunodeficiency disorder (PID) caused by biallelic mutations occurring in the serine/threonine protein kinase (ATM) gene. The major role of nuclear ATM is the coordination of cell signaling pathways in response to DNA double-strand breaks, oxidative stress, and cell cycle checkpoints. Defects in ATM functions lead to A-T syndrome with phenotypic heterogeneity. Our study reports the case of a Tunisian girl with A-T syndrome carrying a compound heterozygous mutation c.[3894dupT]; p.(Ala1299Cysfs3;rs587781823), with a splice acceptor variant: c.[5763-2A>C;rs876659489] in the ATM gene that was identified by next-generation sequencing (NGS). Further genetic analysis of the family showed that the mother carried the c.[5763-2A>C] splice acceptor variant, while the father harbored the c.[3894dupT] variant in the heterozygous state. Molecular analysis provides the opportunity for accurate diagnosis and timely management in A-T patients with chronic progressive disease, especially infections and the risk of malignancies. This study characterizes for the first time the identification of compound heterozygous ATM pathogenic variants by NGS in a Tunisian A-T patient. Our study outlines the importance of molecular genetic testing for A-T patients, which is required for earlier detection and reducing the burden of disease in the future, using the patients' families.

Novel and recurrent BRCA1/BRCA2 germline mutations in patients with breast/ovarian cancer: a series from the south of Tunisia.

BACKGROUND: The incidence of breast cancer (BC) and/or ovarian cancer (OC) is increasing in Tunisia especially in young women and mostly those with family history. However, the spectrum of BRCA mutations remains little explored in Tunisian patients in particular in the southern region. METHODS: We sequenced the entire coding regions of BRCA1and BRCA2 genes using next generation sequencing (NGS) in 134 selected patients with BC and/or OC. RESULTS: Among the 134 patients, 19 (14.17%) carried pathogenic mutations (10 are BRCA1 mutation carriers and 9 are BRCA2 mutation carriers) that are mainly frameshift index (76.9%). Interestingly, 5 out of the 13 variants (38.46%) were found at least twice in unrelated patients, as the c.1310-1313 delAAGA in BRCA2 and the c.5030_5033 delCTAA that has been identified in 4/98 BC patients and in 3/15 OC patients from unrelated families with strong history of cancer. Besides recurrent mutations, 6 variant (4 in BRCA1 and 2 in BRCA2) were not reported previously. Furthermore, 3 unrelated patients carried the VUS c.9976A > T, (K3326*) in BRCA2 exon 27. BRCA carriers correlated significantly with tumor site (p = 0.029) and TNBC cases (p = 0.008). In the groups of patients aged between 31 and 40, and 41-50 years, BRCA1 mutations occurred more frequently in patients with OC than those with BC, and conversely BRCA2 carriers are mostly affected with BC (p = 0.001, and p = 0.044 respectively). CONCLUSIONS: The overall frequency of the BRCA germline mutations was 14.17% in patients with high risk of breast/ovarian cancer. We identified recurrent mutations as the c.1310_1313 delAAGA in BRCA2 gene and the c.5030_5033 delCTAA in BRCA1 gene that were found in 4% and 20% of familial BC and OC respectively. Our data will contribute in the implementation of genetic counseling and testing for families with high-risk of BC and/or OC.

Identification of novel candidate genes by exome sequencing in Tunisian familial male breast cancer patients.

Male Breast Cancer (MBC) is a rare and aggressive disease that is associated with genetic factors. Mutations in BRCA1 and BRCA2 account for 10% of all MBC cases suggesting that other genetic factors are involved. The aim of the present study is to screen whole BRCA1 and BRCA2 exons using the Ampliseq BRCA panel in Tunisian MBC patients with family history. Furthermore, we performed exome sequencing using the TruSight One sequencing panel on an early onset BRCA negative patient. We showed that among the 6 MBC patients, only one (MBC-F1) harbored a novel frameshift mutation in exon 2 of the BRCA2 gene (c.17-20delAAGA, p.Lys6Xfs) resulting in a short BRCA2 protein of only 6 amino-acids. We selected 9 rare variants after applying several filter steps on the exome sequencing data. Among these variants, and based on their role in breast carcinogenesis, we retained 6 candidate genes (MSH5, DCC, ERBB3, NOTCH3, DIAPH1, and DNAH11). Further studies are needed to confirm the association of the selected genes with family MBC.

RIP140 and LCoR expression in gastrointestinal cancers.

The transcription coregulators RIP140 and LCoR are part of a same complex which controls the activity of various transcription factors and cancer cell proliferation. In this study, we have investigated the expression of these two genes in human colorectal and gastric cancers by immunohistochemistry. In both types of tumors, the levels of RIP140 and LCoR appeared highly correlated. Their expression tended to decrease in colorectal cancer as compared to adjacent normal tissues but was found higher in gastric cancer as compared to normal stomach. RIP140 and LCoR expression correlated with TNM and tumor differentiation. Significant correlations were observed with expression levels of key proteins involved in tumor progression and invasion namely E-cadherin and Cyclooxygenase-2. Survival analysis showed that patients with LCoRlow/RIP140high colorectal tumors have a significant prolonged overall and disease-free survival. In gastric cancer, high LCoR expression was identified as an independent marker of poor prognosis suggesting a key role in this malignancy. Altogether, these results demonstrate that RIP140 and LCoR have a prognostic relevance in gastrointestinal cancers and could represent new potential biomarkers in these tumors.

Clinical and prognosis relevance of COX-2 expression in Tunisian patients with primary gastric adenocarcinoma.

BACKGROUND: Inflammation and hormonal signalling induce the cyclooxygenase-2 (COX-2) expression in various human cancers including Gastric Cancer (GC). GC remains among the human malignancies diagnosticated at advanced tumor stage and thus having a poor prognosis. COX-2 is a key protein in cancer progression which is involved in proliferation, invasion, and metastasis of tumor cells. OBJECTIVE: The aim of this study was to investigate the expression of COX-2 and its association with clinico-patholocigal parameters and survival in Tunisian GC patients and to correlate COX-2 expression with others cancer-related proteins. METHODS: The immunohistochemistry was used to study the expression of COX-2 on 93 patients with gastric adenocarcinoma. RESULTS: Our results show that COX-2 immunostaining is negative to weak in 51.6%, moderate in 33.3%, and intense in 15.1% of tumor tissues. The expression of COX-2 associated significantly with tumor differentiation (p = 0.003), and histological type (p = 0.039). Furthermore, lack of COX-2 expression is significantly associated with 1-year (p= 0.005), 2-years (p= 0.000), and 5-years (p= 0.042) relapse free survival. In addition, Cox regression model, revealed that metastasis (p= 0.014), tumor site (p= 0.013), histotype (p = 0.02), and COX-2 expression (p = 0.003) are independent factors for prognosis. Regarding the relationship between COX-2 and cancer related proteins, we found that COX-2 expression is positively associated with APC (p = 0.006), and P53 (p = 0.026), supporting a cross link between these proteins in gastric carcinogenesis. CONCLUSION: Our findings emphasize the importance of COX-2 as a potential marker of tumor progression and prognosis in GC, and that the inhibition of COX-2 activity may have a therapeutic benefit in GC.

Hypermethylation of tumor-related genes in Tunisian patients with gastric carcinoma: clinical and biological significance.

BACKGROUND: Promoter hypermethylation is an alternative mechanism of gene silencing in cancers including gastric carcinoma (GC). Its affects genes with crucial functions as tumor suppressor. METHODS: DNA methylation in the promoter of P16INK4a, DAPK, retinoic acid receptor ß (RARß2), RASSF1A, and CDH1 genes was investigated in 79 Tunisian patients with GC using methylation-specific PCR. RESULTS: The methylation frequencies vary from 31.6% for P16INK4a to 65.8% for RARß2. Hypermethylation of DAPK and CDH1 was associated with tumor grade and age (P = 0.04 and 0.034) respectively, while hypermethylation of RASSF1A correlated with TNM stage (P = 0.027). The distribution of the methylated DNA at P16INK4a, DAPK, and CDH1 promoters were different in the intestinal and diffuse histotypes of GC according to TNM. Moreover, the survival rate of patients with P16INK4a methylated status was shorter than that of patients with the unmethylated status (P log rank = 0.009). On the other hand, the hypermethylation of RARß2 correlated with COX-2 expression (P = 0.001). CONCLUSION: We showed that methylation of P16INK4a is predictive of poor prognosis and could be a useful marker. Moreover, the association between RARß2 methylation and COX-2 expression suggests a functional link between these two proteins in gastric carcinogenesis.