Dimitra Vageli

Transcriptional activation of H-ras, K-ras and N-ras proto-oncogenes in human bladder tumors

In this study we demonstrate the involvement of ras oncogenes in bladder cancer at the level of RNA overexpression. We examined 26 bladder specimens, consisting of paired tumor and adjacent normal tissue and found that H-ras transcripts were overexpressed in 39% of the specimens while K-ras and N-ras in 58% of total specimens. Each tumor specimen had a unique pattern of overexpression for the three ras genes. A competitive-RT-PCR was employed for H-ras and a beta-actin control gene was co-amplified with K-ras or N-ras genes. These results indicate that the involvement of ras oncogenes in bladder cancer could be relative to overexpression of these genes.

High-risk human papillomavirus (HPV) in parotid lesions

Although several studies have reported that oropharyngeal infection with HPV may predispose to tumorigenesis, little is known about the etiological factors of salivary gland tumors and the presence of HPV. We studied 9 parotid lesions for HPV infection including an oncocytoma, an acinic cell carcinoma, a high-grade adenocarcinoma, a low-grade polymorphous adenocarcinoma, a Warthins tumor and 2 pleomorphic adenomas, a lymphoepithelial cyst and a lipoma of the parotid gland. DNA was extracted from formalin-fixed and paraffin-embedded tissue sections. Solution PCR for HPV detection was performed using the GP5+/GP6+ primers, while HPV typing was carried out by multiplex PCR for HPV6, 11, 16, 18, and 33; positive samples were recorfirmed by PCR with specific primers for each type. Quantitative real-time PCR for the high-risk HPV genotypes 16, 18, 31, 33, 35, 52, 58 and 67 was also performed to quantitate the viral load. Finally, in situ PCR was employed with HPV16-specific primers by direct-detection method. Seven of the 9 parotid lesions were HPV positive while 6 of these 7 had been infected by HPV16 and/or HPV18 oncogenic types. High viral load of highrisk genotypes of HPV was found in the oncocytoma, in one of the pleomorphic adenomas, and in the Warthins tumor. Finally, in situ PCR indicated that HPV16 amplification occurred in the salivary gland tumors. This is the first time that highrisk HPV genotypes are detected in these histological types of parotid lesions, suggesting the possible involvement of the virus in the disease.

Transcriptional Activation of hTERT in Breast Carcinomas by the Her2-ER81-Related Pathway

Her2 and ER81 (a member of ETS family) have been suggested to cause a synergistic increase in the transcriptional activation of hTERT. Our study aimed to offer further confirmation in clinical material. We determined the mRNA levels of Her2, ER81, and hTERT, by QRT-PCR, in 43 breast carcinomas. In the specimens showing hTERT transcriptional activation, Her2 and ER81 were increased in statistically significant tumor subgroups (61% and 79% correspondingly). The 86% of specimens with both Her2 and ER81 increased expression showed hTERT transcriptional activation. Synchronous transcriptional activation of hTERT, Her2, and ER81 elevated expression was noted in 42% of the samples. In conclusion, we agree with a previous study that Her2 overexpression may increase the hTERT transcriptional activation. Our data indicate that the mechanism may involve Her2-ER81 interaction(s) and that the activation of hTERT could be mainly mediated by transcriptional activation of ER81.

hMSH2 and hMLH1 gene expression patterns differ between lung adenocarcinoma and squamous cell carcinoma: correlation with patient survival and response to adjuvant chemotherapy treatment

Background We recently showed that the mRNA levels of mismatch repair (MMR) proteins in non-small cell lung carcinoma (NSCLC) tissue specimens and the phenotypic translation of molecular MMR data refines the biology of the MMR system with consequent diagnostic implications in the clinical assessment of lung cancer patients. Methods hMLH1 and hMSH2 mRNA expression was previously evaluated by qPCR for 29 NSCLC patients (13 with squamous cell carcinoma [SQC] and 16 with adenocarcinoma [ADC]) and MMR mRNA levels were converted into clinically distinct phenotypic entities. In this study, we evaluated the correlation of the hMSH2 and hMLH1 mRNA phenotypes with patient survival and their response to adjuvant chemotherapy. Results hMSH2 and hMLH1 mRNA phenotypic distribution differed between SQC and ADC. the MMR phenotypes differed also between advanced and early stage SQC. SQC patients with an increased hMSH2 expression had a better outcome than patients with a reduced hMSH2 expression. However, ADC patients with an increased hMSH2 expression had a poor outcome compared to those with low hMSH2 levels. SQC patients with a high hMSH2 expression exhibited a better response to adjuvant chemotherapy, whereas ADC patients with high hMSH2 levels had a poor response. ADC patients with low hMSH2 levels showed good response to adjuvant chemotherapy compared to SQC patients bearing the same phenotypic profile. Conclusions Our findings show that MMR mRNA phenotypes may be added to the known biological differences between SQC and ADC. hMLH1 and hMSH2 phenotypes distributed differently according to the NSCLC stage. Distinct MMR mRNA phenotypes in SQC and ADC corresponded to patient response to adjuvant chemotherapy.

Inhibition of NF- κ B prevents the acidic bile-induced oncogenic mRNA phenotype, in human hypopharyngeal cells

Bile-containing gastro-duodenal reflux has been clinically considered an independent risk factor in hypopharyngeal carcinogenesis. We recently showed that the chronic effect of acidic bile, at pH 4.0, selectively induces NF-κB activation and accelerates the transcriptional levels of genes, linked to head and neck cancer, in normal hypopharyngeal epithelial cells. Here, we hypothesize that NF-κB inhibition is capable of preventing the acidic bile-induced and cancer-related mRNA phenotype, in treated normal human hypopharyngeal cells. In this setting we used BAY 11-7082, a specific and well documented pharmacologic inhibitor of NF-κB, and we observed that BAY 11-7082 effectively inhibits the acidic bile-induced gene expression profiling of the NF-κB signaling pathway (down-regulation of 72 out of 84 analyzed genes). NF-κB inhibition significantly prevents the acidic bile-induced transcriptional activation of NF-κB transcriptional factors, RELA (p65) and c-REL, as well as genes related to and commonly found in established HNSCC cell lines. These include anti-apoptotic bcl-2, oncogenic STAT3, EGFR, ∆Np63, TNF-α and WNT5A, as well as cytokines IL-1β and IL-6. Our findings are consistent with our hypothesis demonstrating that NF-κB inhibition effectively prevents the acidic bile-induced cancer-related mRNA phenotype in normal human hypopharyngeal epithelial cells supporting an understanding that NF-κB may be a critical link between acidic bile and early preneoplastic events in this setting.

Mismatch DNA repair mRNA expression profiles in oral melanin pigmentation lesion and hamartomatous polyp of a child with peutz–jeghers syndrome

Mismatch DNA repair (MMR) mRNA expression analysis was performed on a biopsy of oral mucosa melanin pigmentation lesion, a hamartomatous polyp and peripheral blood derived from a 12‐year‐old child with Peutz–Jeghers Syndrome (PJS). We present a deficient MMR system, in a PJS patient, which demonstrated low mRNA levels of hMSH6 and hPMS2 and an increasing MMR deficiency from the non‐dysplastic lesion to hamartomatous polyp of PJS with a high risk of cancer. Pediatr Blood Cancer 2013;60:E116–E117.

Mismatch DNA Repair hMSH2, hMLH1, hMSH6 and hPMS2 mRNA Expression Profiles in Colorectal Carcinomas

Background: Mismatch repair (MMR) deficiency has been related with HNPCCs. So far, there is limited information on MMR mRNA profiles in sporadic colorectal carcinomas (CRCs). We previously showed that distinct MMR mRNA phenotypes were related to tumor stage and survival of patients with lung cancer or urinary bladder carcinomas. Aim: The aim of this study was to quantify hMSH2, hMLH1, hMSH6 and hPMS2 mRNA levels, in CRCs and their adjacent normal tissues (ANTs), using accurate methodology, and to correlate MMR mRNA profiles with patient or tumor characteristics. Materials and methods: We analyzed 31 fresh frozen tissue specimens of paired CRCs with their ANTs. We evaluated MMR mRNA profiles by a Q-real-time PCR, using hPBGD gene as reference control and creating a standard curve. The MMR mRNA levels were assigned as ratios MMR/hPBGD mRNAs. Relative expression of each MMR gene was given as ratios of CRCs/ANTs mRNA levels. Results: All CRCs and their ANTs expressed low hPMS2 mRNA levels while a significant proportion of CRCs (73%) and their ANTs (82%) presented low hMSH2 mRNA levels. Analysis of relative expression patterns showed that hMSH6 and hMLH1 exhibited the highest percentages of reduction (53% and 45.5%, respectively). We found a correlation of transcriptional levels between hMSH2 and hMLH1, the crucial components of MMR mechanism and between their counterparts, hMSH6 and hPMS2, in CRCs of early stages, related to gender. On the contrary, CRCs of late stages revealed a correlation between reduced levels of hMSH2 and hMSH6, MutSa components, unrelated to gender but related to lymph node metastasis. Also, reduced hMSH2, hMSH6 and hMLH1 mRNA phenotypes correlated with advanced stage, and rectal localization. Conclusion: In this study we demonstrated that MMR mRNA deficiency is a common event in sporadic CRCs. Specific profiles of MMR deficiency maybe related to tumor progression, especially in male patients.

HPV6 Infection of an InfantâÂÂs Penile Condyloma at the Urethral Meatus

Penile warts in infants are rare and condylomata acuminata at the urethral meatus are rarer. This is a first report of a 2.5 years infant with a condyloma acuminatum at the urethral meatus of penis that was found positive for HPV 6, by PCR analysis. Infant’s mother presented warts on her hands, suggesting a possible transmission of virus from mother to infant via extragenital contact, without excluding a vertical transmission.