p16 as an independent marker for detection of high-risk HPV in oral submucous fibrosis and oral squamous cell carcinoma.

BACKGROUND: An alarming increase in incidence of high-risk human papillomavirus (HPV) positive tumors in head and neck squamous cell carcinoma (HNSCC) by 25% and 70% in oropharyngeal HNSCC cannot be ignored. The early oncogenes of HPV, E6, and E7 play a key role in carcinogenesis. HPV associated tumors have a better clinical outcome and a favorable prognosis. The p16 expression has high concordance with other methods of HPV detection, ascertaining p16 as a surrogate marker for HPV. OBJECTIVE: To assess the immunohistochemical expression of p16 in oral submucous fibrosis (OSF) and oral squamous cell carcinoma (OSCC) with and without coexistent OSF as a marker for high-risk HPV detection. Materials and. METHODS: Tissue blocks of 70 cases including normal, OSF, OSCC with and without OSF were subjected to IHC staining with a p16INK4A monoclonal antibody. (Biogenex, San Roman). The p16 expression was noted according to percent positivity and pattern. The data were tabulated, statistically analyzed using the Chi-square test and the P value was assessed. RESULTS: The percentage of p16 positive cells raised from normal to OSF to OSCC with and without OSF. In addition, a shift from nuclear to cytoplasmic expression from normal to OSCC was noted with a statistical significance (P < 0.001). However, no statistical significance was established with any clinicopathologic parameters except age (P = 0.012) and habits (P= 0.023). CONCLUSION: The presence of HPV using p16 was not detected in OSF but was positive in OSCC. Altered pattern of expression from normal to OSF to OSCC indicates promising use of p16 as a diagnostic marker.

Comparative study between the Hybrid Capture II test and PCR based assay for the detection of human papillomavirus DNA in oral submucous fibrosis and oral squamous cell carcinoma.

BACKGROUND: Oral malignancy is a major global health problem. Besides the main risk factors of tobacco, smoking and alcohol, infection by human papillomavirus (HPV) and genetic alterations are likely to play an important role in these lesions. The purpose of this study was to compare the efficacy of HC-II assay and PCR for the detection of specific HPV type (HPV 16 E6) in OSMF and OSCC cases as well as find out the prevalence of the high risk HPV (HR-HPV) in these lesions. METHODS AND MATERIALS: Four hundred and thirty patients of the potentially malignant and malignant oral lesions were taken from the Department of Otorhinolaryngology, Moti Lal Nehru Medical College, Allahabad, India from Sept 2007-March 2010. Of which 208 cases were oral submucous fibrosis (OSMF) and 222 cases were oral squamous cell carcinoma (OSCC). The HC-II assay and PCR were used for the detection of HR-HPV DNA. RESULT: The overall prevalence of HR-HPV 16 E6 DNA positivity was nearly 26% by PCR and 27.4% by the HC-II assay in case of potentially malignant disorder of the oral lesions such as OSMF. However, in case of malignant oral lesions such as OSCC, 32.4% HPV 16 E6 positive by PCR and 31.4% by the HC-II assay. In case of OSMF, the two test gave concordant result for 42 positive samples and 154 negative samples, with an overall level of agreement of 85.4% (Cohen's kappa = 66.83%, 95% CI 0.553-0.783). The sensitivity and specificity of the test were 73.7% and 92.05% (p < 0.00). In case of OSCC, the two test gave concordant result for 61 positive samples and 152 negative samples, with an overall level of agreement of 88.3% (Cohen's kappa = 79.29, 95% CI 0.769-0.939) and the sensitivity and specificity of the test were 87.14% and 92.76% (p < 0.00). CONCLUSION: This study concluded that slight difference was found between the positivity rate of HR-HPV infection detected by the HC-II and PCR assay in OSMF and OSCC cases and the HC II assay seemed to have better sensitivity in case of OSCC.

Prevalence of viral (HPV, EBV, HSV) infections in oral submucous fibrosis and oral cancer from India.

CONCLUSION: There is a high prevalence of human papilloma viruses (HPV) in oral submucous fibrosis (OSMF) and the etiologic implication of this finding warrants further studies. OBJECTIVE: The prevalence of oral squamous cell carcinoma (OSCC) and OSMF is high in India, and the diseases are partly attributed to high consumption of betel quid containing areca nut and tobacco. This study investigated the prevalence of HPV, herpes simplex virus (HSV), and Epstein­Barr virus (EBV) DNA in two groups of patients using betel quid with tobacco, those with OSMF (n = 12) and those with OSCC (n = 62). METHODS: DNA was extracted from all the samples and viral genome was examined by PCR/DNA sequencing. HPV-positive samples were analyzed separately for the high-risk types HPV 16 and 18. RESULTS: HPV DNA, HSV DNA, and EBV DNA were detected in 11 (91%), 1 (8%), and 3 (25%) of the 12 samples from patients with OSMF compared with 15 (24%), 3 (5%), and 18 (29%), respectively, from 62 patients with OSCC. HPV 16 and 18 DNA was detected in 8/12 (67%) in the OSMF group and 10/62 (16%) in the OSCC group. The difference between presence of HPV DNA in OSMF and OSCC groups was statistically significant, while the difference between HSV and EBV DNA content in OSMF and OSCC groups was insignificant.

Human papillomaviruses in oral squamous cell carcinoma and pre-cancerous lesions detected by PCR-based gene-chip array.

Human papillomavirus (HPV) infection is a significant risk factor for uterine cervical carcinoma. Many studies have also demonstrated the presence of HPV in oral epithelia tissue, but the role of HPV infection in oral squamous cell carcinoma (OSCC) is still controversial. The aim of this study was to determine the frequency and type of HPV in OSCC and oral pre-cancerous lesions. DNA samples were collected by cytobrushing from 51 patients with OSCC, 46 with oral pre-cancerous lesions and 90 normal controls. Nested polymerase chain reaction and gene-chip arrays were used to identify the HPV types in the samples. In pre-cancerous lesions, there was a higher frequency of HPV of any type (14/46, OR = 2.844, CI = 1.186-6.816, P = 0.0216) and of low-risk HPV types (9/46, OR = 5.529, CI = 1.597-19.14, P = 0.0096) than in control samples. The prevalence of high-risk types was significantly higher in OSCC than in control lesions (11/51 vs 8/90, OR = 2.819, CI = 1.051-7.558, P = 0.0420) but this was not the case for HPV of any type (13/51 vs 12/90, OR = 2.244, CI = 0.9266-5.337, P = 0.1066). High-risk HPV types are prevalent in OSCC and may play a role in its progression, while low-risk types are associated with oral pre-cancerous lesions.

Demonstration of different modes of cell death upon herpes simplex virus 1 infection in different types of oral cells.

The effects of Herpes simplex virus 1 (HSV-1) infection on five different types of oral cancerous cells (neck metastasis of gingival carcinoma (GNM) cells and tongue squamous cells of carcinoma (TSCCa) and non-cancerous cells (buccal mucosal fibroblasts (BF), gingival fibroblasts (GF), oral submucosal fibrosis cells (OSF)) and one type of non-oral cancerous cells (KB cells) were investigated. In HSV-1-infected cells the cell viability, CPE, viral antigens accumulation, caspase-3 activity, annexin V binding and DNA fragmentation were estimated. Three different forms or pathways of cell death were considered: apoptosis (the presence or rise of caspase-3 activity, DNA fragmentation and annexin V binding), slow cell death (the presence or rise of DNA fragmentation, the absence or decline of caspase-3 activity and annexin V binding), and necrosis (the absence of decline of caspase-3 activity, DNA fragmentation and annexin V binding). The viability of all cell types, except for KB cells, was reduced by the infection. CPE and viral antigens data demonstrated that all six types of cells could be infected with HSV-1. Upon HSV-1 infection there occurred (i) a classical apoptosis in GF cells, (ii) apoptosis in the early phase of infection and necrosis in the late phase of infection in GNM and TSCCa cells, (iii) slow cell death followed by necrosis in BF and OSF cells (however, these cells showed a different type of CPE), (iv) a classical slow cell death in KB cells. It is hypothesized that HSV-1 infection has a potential to induce several distinct pathways leading to cell death or several forms of cell death. Moreover, more than one pathway may be involved in the death of particular cell type. As HSV-1 was demonstrated to infect different oral and non-oral cells and cause different pathways or forms of cell death, the safety of using HSV-1 as a vector for gene therapy should be re-considered.