Investigation of human JC and BK polyomaviruses in breast carcinomas.

We have previously showed the presence of the simian virus 40 (SV40) and the mouse mammary tumor virus (MMTV)-like in a significant proportions of Tunisian breast carcinomas. However, to date there are no published studies concerning evaluation of the possible implication of the human polyomaviruses JC (JCV) and BK (BKV) in breast carcinomas. The presence of JCV and BKV DNA was investigated by PCR in a 123 primary breast carcinomas and matched adjacent non-tumor breast tissues. The results were correlated to clinicopathological and virological parameters. JCV T-antigen DNA was detected in 23% of breast carcinoma cases; however, all cases were negative for BKV. JCV T antigen PCR products were further confirmed as authentic JCV genome by direct sequencing. JCV was found in invasive ductal carcinomas (28/112 cases) but not in invasive lobular carcinomas (0/5) or medullary carcinomas (0/6). JCV DNA presence correlates inversely with the expression of estrogen (P = 0.022) and progesterone (P = 0.008) receptors. JCV DNA presence correlates also with "triple negative" phenotype (P = 0.021). With regard to virological data, a trend toward an inverse correlation was noted between the presence of JCV and SV40 (P = 0.06). Moreover, significant correlation was found between multiple viral infection (JCV, and/or SV40, and/or MMTV-like in the same tumor) and "triple negative" phenotype (P = 0.001) and also with p53 accumulation (P = 0.028). To the best of our knowledge, this is the first study demonstrating the presence of JCV in a subset of breast carcinomas. Also our results suggest that "triple negative" breast carcinomas are viral-related tumors.

DNA methylation patterns in EBV-positive and EBV-negative Hodgkin lymphomas.

PURPOSE: Hodgkin lymphoma (HL) is characterized by the presence of Hodgkin and Reed-Sternberg cells. Epstein-Barr virus (EBV) infection is thought to play an important role in the development of HL. Although epigenetic alterations, such as aberrant DNA methylation, are known to contribute to the pathogenesis of various malignancies, little is known about such alterations in HL and their putative relationships with EBV infection. METHODS: We investigated promoter methylation patterns of seven tumor-associated genes in 53 primary HL cases using methylation-specific PCR (MS-PCR). Concomitantly, the EBV infection status was assessed using PCR, in situ hybridization and immunohistochemistry. RESULTS: The gene promoter hypermethylation frequencies observed were 77.3 % for P16, 58.5 % for RASSF1A, 50.9 % for CDH1, 45.3 % for DAPK, 43.4 % for GSTP1, 37.7 % for SHP1 and 24.3 % for MGMT. SHP1 gene promoter hypermethylation was more frequently observed in patients at extreme ages (i.e., ≤ 15 and >54 years) than in adult patients (p = 0.006) and in patients with B symptoms (p = 0.03). Interestingly, most of the analyzed gene promoters were more frequently hypermethylated in EBV-negative than in EBV-positive cases, in particular the DAPK gene promoter (58 % versus 27 %, p = 0.04). Furthermore, hypermethylation of multiple gene promoters (≥ 3) was encountered more frequently in females than in males (86 % versus 57 %, p = 0.04), whereas EBV-positive cases were more common among males than females (55 % versus 30 %, p = 0.02). CONCLUSIONS: Our results indicate that epigenetic changes frequently occur in both EBV-positive and EBV-negative HL. The rates of these changes were found to vary according to clinico-pathological parameters. These observations probably reflect the multitude of factors involved in HL development and the complexity of their interactions with genetic and/or hormonal factors.

Increased DNA methyltransferase 1 protein expression correlates significantly with intestinal histological type and gender in gastric carcinomas.

PURPOSE: Promoter hypermethylation and reduced expression of many genes have been found in gastric cancer. DNA methyltransferases are enzymes potentially affecting promoter hypermethylation. MATERIAL AND METHODS: We analyzed proteins expression of DNA methyltransferase 1 and 3b by immunohistochemistry in 47 surgically resected gastric cancer samples for which clinicopathological characteristics, patient's outcome and methylation status of 11 selected tumor-related genes have been determined. Promoter methylation status of genes was assessed by methylation specific PCR. RESULTS: We found that DNMT1 and 3b were up-regulated in gastric cancer and were detected in 51.1% and 57.4% of cases, respectively. Co-expression of DNMT1 and 3b was detected in 44.7%. Correlations analysis have showed that DNMT1 overexpression was significantly correlated with gastric cancer of intestinal histological type (P=0.01) and with gender of patient (P=0.01). However, there was no correlation between DNMT1 and DNMT3b overexpression in cancer and patients outcome. Moreover, there were no clear relations between the proteins expression of DNMT1 and 3b and DNA methylation status of genes. But co-expression of DNMT1 and DNMT3b was significantly associated with promoter hypermethylation of RAR-ß2 (P=0.04). CONCLUSIONS: Results from our study indicate that DNMT1 and 3b were overexpressed and could be involved in gastric tumorigenesis of intestinal histological type in the case of Tunisian patients.

Investigation of Epstein-Barr virus in breast carcinomas in Tunisia.

Breast carcinoma is a major cause of death among women, and the potential implication of viruses in its pathogenesis remains worth a hypothesis. The potential role of Epstein-Barr virus (EBV) in its pathogenesis is still a subject of continued discussion and investigation. The aim of this study was to evaluate the prevalence of EBV in sporadic breast cancers in Tunisia, and to determine the clinicopathological characteristics of virus-positive cases. Viral presence has been evaluated by polymerase chain reaction (PCR), in situ hybridization, and immunohistochemistry investigated on tumor tissues and their corresponding normal breast tissues collected from 123 Tunisian women with sporadic breast carcinomas. Viral status in tumors was then correlated with various clinicopathological parameters. Using specific PCR assays, EBV DNA was found in 33 (27%) out of 123 breast carcinoma cases. EBV-encoded small RNAs (EBERs) in situ hybridization was negative in the neoplastic cells, but stomal lymphocytes were positive in 4 cases. Immunohistochemistry for latent membrane protein 1 (LMP1) was negative in all cases. None of the normal breast tissues showed positive results for EBV using PCR, in situ hybridization, and immunohistochemistry. A correlation was found between EBV DNA presence and the negativity of estrogen receptor (P=0.008). However, no significant correlation was found for the other parameters investigated, including patient age, Scarff-Bloom-Richardson (SBR) histological grade, tumor size, and histological node involvement. With regard to survival data, overall and disease-free survivals were shorter in EBV-positive breast carcinoma cases than in EBV-negative ones, but this difference did not reach statistical significance. Our study indicates the presence of EBV DNA in a significant proportion of breast cancer in Tunisia. Further studies are required to elucidate the role of this virus in breast carcinogenesis.

Prevalence and characteristics of Epstein-Barr virus-associated gastric carcinomas in Tunisia.

OBJECTIVE: Epstein-Barr virus (EBV) has been linked to gastric carcinoma (GC) with worldwide geographical variations of prevalence ranging from 1 to 18% of cases. Investigations carried out in north Africa have shown that some EBV-associated types of cancers are common in this area. This study was taken to determine the prevalence of EBV-associated GC in Tunisia. METHODS: Ninety-six nonselected GC cases (male/female ratio 1.7/1, mean age 60.9 years, range: 20-88 years) were evaluated for the presence of EBV by polymerase chain reaction as well as by in-situ hybridization for EBV-encoded small RNAs (EBERs) and immunohistochemistry for LMP-1 and EBNA-2 expression. RESULTS: EBV was detected by polymerase chain reaction in 36% of cases, whereas EBERs were detected in the tumor cells in only four cases (4.1%). Immunohistochemistry for LMP-1 and EBNA-2 was negative in all cases. The mean age for patients harboring EBERs-positive GC was 55.7 years (range: 52-59 years). All EBERs-positive GC cases were males of advanced clinical stage (pT3-pT4). According to Lauren's classification, two cases were of diffuse histological type and two cases were of intestinal type. In three cases, the tumors have a proximal location and in the remaining case the tumor arises in the antrum. All EBV strains detected from EBV-associated GC were exclusively of type A and D, prototype F, and XhoI-maintained variant. CONCLUSION: We conclude that the prevalence of EBV-associated GC in Tunisia is low (4.1%), suggesting that this virus is not an important etiological factor in GC arising in north African populations. The clinicopathological profile of EBV-associated GC in Tunisia did not differ markedly from that found elsewhere.