• Asian Pacific Journal of Cancer Prevention
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• v.15 no.24
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• pp.10803-10808
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• 2015

Background: The role of $p16^{INK4A}$ expression in uterine cervix cancer is well established. In the remaining female lower genital tract cancers, the importance of $p16^{INK4A}$ up-regulation is less clear. In our study, we analyzed the role of $p16^{INK4A}$ expression and HPV infection in carcinomas of the vulva and the vagina in Tunisian women. Materials and Methods: We conducted a retrospective study of 30 carcinomas including 15 vulvar squamous cell carcinomas (SCCs) and 15 vaginal SCCs. Immunohistochemistry was used to determine $p16^{INK4A}$ expression. HPV detection and typing was by in situ hybridization. Results: $p16^{INK4A}$ expression was detected in 86.7% of vaginal SCCs with a strong and diffuse immunostaining in 60% of cases, and also in 73.3% of vulvar SCCs with focal immunoreactivity in 53.3% The association between $p16^{INK4A}$ expression and HPV infection was significant in vaginal SCCs (p=0.001) but not vulvar SCCs (p>0.05). Conclusions: $p16^{INK4A}$ expression could be used as a useful marker for HPV positivity in vaginal SCCs similar to that described in uterine cervix cancers. However, our data support the presence of 2 different mechanisms for $p16^{INK4A}$ expression in HPV-related and HPV-unrelated vulvar carcinomas.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.3
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• pp.1165-1168
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• 2015

Objective: To investigate the incidence of malignant transformation and P53 and P16 expression in teratomatous skin of ovarian mature cystic teratoma. Materials and Methods: Data on ovarian teratoma specimens in nearly 10 years were reviewed. P53 and P16 expression were detected by immunohistochemistry in 25 cases of teratomatous skin of ovarian mature cystic teratoma, 20 cases of squamous cell carcinoma and 2 cases of squamous cell carcinoma originated from teratomatous skin. Results: Of 1913 cases of ovarian mature cystic teratoma in nearly 10 years, only two cases of squamous cell carcinoma were found in teratomatous skin, with malignant transformation rate of 0.1045%. P53 expression was detected in 2 cases squamous cell carcinoma originated from teratomatous skin and P16 overexpression in one. There were no expressions of P53 and P16 in 25 cases of teratomatous skin of ovarian mature cystic teratoma. Of 20 cases of squamous cell carcinoma P53 overexpression (positive rate of 55%) was detected in 11 cases, P16 overexpression (positive rate of 35%) in 7 cases. The positive rates of P53 and P16 expression in squamous cell carcinomas were significantly higher than that in the teratomatous skins (p< 0.001, p= 0.002). Conclusions: There was low risk of malignant transformation in teratomatous skin of ovarian mature cystic teratoma which can be explained by lower P53 and P16 expressionin teratomas than that in squamous cell carcinoma.

• IMMUNE NETWORK
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• v.4 no.4
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• pp.237-243
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• 2004

Background: The normal functions of the cell cycle inhibitor p16INK4a are frequently inactivated in many human cancers. Over 80% of hepatocellular carcinoma (HCC) cases lack a functional p16/Rb pathway. p16/Rb pathway, as well as p53 pathway, is considered as one of key components of tumor suppression. Methods: To study the roles of p16INK4a in HCC, a stable cell line expressing exogenous p16 was generated from SNU-449 hepatocellular carcinoma cells lacking endogenous p16, and suppression subtractive hybridization (SSH) was performed in parallel with the control cells. Results: 1) SSH identifies fibronectin (FN1), crystallin ${\alpha}B$ (CRYAB), Rac1, WASP, RhoGEF, and CCT3 as differentially-expressed genes. 2) Among the selected genes, the up-regulation of FN1 and CRYAB was confirmed by Northern blot, RT-PCR and by proteomic methods. Conclusion: These genes are likely to be associated with the induction of stress fiber and stabilization of cytoskeleton. Further studies are required to clarify the possible role of p16 in the signal transduction pathway.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.7
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• pp.3201-3206
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• 2016

Purpose: To evaluate the expression of p16 and Ki67 in cervical intraepithelial neoplasia (CIN) and cancer. Materials and Methods: We performed a immunohistochemical study of p16 and Ki67 in 243 cervical tissues - 53 non-dysplastic lesions, 106 CIN1, 61 CIN2/3 and 23 squamous cell carcinomas. The expression of p16 and Ki67 was interpreted independently by 2 researchers and the sensitivity and specificity to detect clinically significant lesions (${\geq}CIN2$) were determined. Results: The overall agreement results of positive or negative immunostaining of intra-inter observer variability were 0.659 for p16 and 0.808 for Ki67. p16 expression was demonstrated in 91.3% of invasive carcinomas, 78.7% of CIN2/3, 10.4% of CIN1 and 9.4% of non-dysplasic lesions. The corresponding Ki67 expression was: 100% of all invasive carcinomas, 75.4% of CIN2/3, 22.6% of CIN1, and 11.3% with non-dysplasia. The expression was significantly different between CIN2/3 vs CIN1 for both p16 and Ki67 (p-values <0.001 both), and cancer vs CIN2/3 for Ki67 (p-value 0.008). The differences were not significant between CIN1 vs non-dysplasia (p-values 1.000 for p16 and 0.130 of Ki67), and cancer vs CIN2/3 for p16 (p value 0.219). The sensitivity and specificity to detect > CIN2 were 84.5% and 90.5% by p16 and 82.1% and 88.6% by Ki67. Conclusions: The rates for 16 and Ki67 expression were directly associated with the severity of cervical lesions. Significant differences in these markers expression may be useful in cases with equivocal histologic features among cervical intraepithelial lesions, but not between CIN1 and non-dysplastic lesions. The two markers had high sensitivity and specificity in determining >CIN2.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.8
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• pp.4149-4154
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• 2016

Background: Cervical carcinoma is one of the main causes of mortality in women worldwide as well as in India. It occurs as a result of various molecular events that develop from the combined influences of an individual's genetic predisposition and external agents such as smoking and menstrual hygiene, for example. However, infection with human papillomavirus (HPV) is the established major risk factor. The aim of the current study was to investigate p16 CpG island methylation and establish any correlation with mRNA expression in north Indian population. Materials and Methods: We analyzed 196 woman volunteer out of which 98 were cases and 98 healthy controls. For the analysis of methylation pattern, DNA extracted from blood samples was modified with a bisulfate kit and used as template for methylation specific PCR (MSP). Quantitative real-time PCR (QRT-PCR) was performed to check mRNA expression. Results: Correlation between methylation status of p16 gene and poor menstrual hygiene was significant (p=0.006), high parity cases showed methylation of p16 gene (p=0.031) with increased risk up to 1.86 times for cervical cancer and smoking was a strong risk factor associated with cervical cancer. We analyzed methylation pattern and found 60.3% methylation in cases with low mRNA expression level (0.014) as compare to controls (1.24). It was also observed that promoter methylation of p16 gene was significantly greater in FIGO stage III. Conclusions: We conclude that p16 methylation plays an important role in cervical cancer in the north Indian population and its methylation decreases mRNA expression. It can be used as an important and consistent blood biomarker in cervical cancer patients.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.19
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• pp.8351-8359
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• 2014

Background: Recent epidemiological data have implicated human papilloma virus (HPV) infection in the pathogenesis of head and neck cancers, especially oropharyngeal cancers. Although, HPV has been detected in varied amounts in persons with oral dysplasia, leukoplakias and malignancies, its involvement in oral tongue carcinogenesis remains ambiguous. Materials and Methods: HPV DNA prevalence was assessed by PCR with formalin fixed paraffin embedded sections (n=167) of oral tongue squamous cell carcinoma patients and the physical status of the HPV16 DNA was assessed by qPCR. Immunohistochemistry was conducted for p16 evaluation. Results: We found the HPV prevalence in tongue cancers to be 51.2%, HPV 16 being present in 85.2% of the positive cases. A notable finding was a very poor concordance between HPV 16 DNA and p16 IHC findings (kappa<0.2). Further molecular classification of patients based on HPV16 DNA prevalence and p16 overexpression showed that patients with tumours showing p16 overexpression had increased hazard of death (HR=2.395; p=0.005) and disease recurrence (HR=2.581; p=0.002) irrespective of their HPV 16 DNA status. Conclusions: Our study has brought out several key facets which can potentially redefine our understanding of tongue cancer tumorigenesis. It has emphatically shown p16 overexpression to be a single important prognostic variable in defining a high risk group and depicting a poorer prognosis, thus highlighting the need for its routine assessment in tongue cancers. Another significant finding was a very poor concordance between p16 expression and HPV infection suggesting that p16 expression should possibly not be used as a surrogate marker for HPV infection in tongue cancers. Interestingly, the prognostic significance of p16 overexpression is different from that reported in oropharyngeal cancers. The mechanism of HPV independent p16 over expression in oral tongue cancers is possibly a distinct entity and needs to be further studied.

• Clinical and Experimental Pediatrics
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• v.53 no.7
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• pp.753-758
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• 2010

Purpose: It has been suggested that p16 has a role in glucocorticoid (GC)-related apoptosis in leukemic cells, but the exact mechanisms have yet to be clarified. We evaluated the relationship between the GC response and p16 expression in a lymphoma cell line. Methods: We used p16 siRNA transfection to construct p16-inactivated cells by using the B-cell lymphoblast cell line NC-37. We compared glucocorticoid receptor (GR) expression, apoptosis, and cell viability between control (p16+NC-37) and p16 siRNA-transfected (p16-NC-37) cells after a single dose of dexamethasone (DX). Results: In both groups, there was a significant increase in cytoplasmic GR expression, which tended to be higher for p16+NC-37 cells than for p16- NC37 cells at all times, and the difference at 18 h was significant (P<0.05). Similar patterns of early apoptosis were observed in both groups, and late apoptosis occurred at higher levels at 18 h when the GR had already been downregulated ($P$<0.05). Cell viability decreased in both groups but the degree of reduction was more severe in p16+NC-37 cells after 18 h ($P$<0.05). Conclusion: These results suggest a relationship between GR expression and cell cycle inhibition, in which the absence of p16 leads to reduced cell sensitivity to DX.

• Toxicological Research
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• v.20 no.2
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• pp.109-116
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• 2004

For the safety evaluation of adenovirus-mediated gene transfer, we investigated differential gene expressions after transfecting adenoviral vector containing p16 tumor suppressor gene (Ad5CMV-p16) into human non-small cell lung cancer cells. In the previous study, we showed adenovirus-mediated $p16^{INK4a}$ gene transfer resulted in significant inhibition of cancer cell growth. We investigated gene expression changes after transfecting Ad5CMV-p16, Ad5CMV (null type, a mock vector) into A549 cells by using cDNA chip and oligonucleotide microarray chip (1200 genes) which carries genes related with signal transduction pathways, cell cycle regulations, oncogenes and tumor suppressor genes. We found that $p16^{INK4a}$ gene transfer down regulated 5 genes (cdc2, cyclin D3, cyclin B, cyclin E, cdk2) among 26 genes involved in cell cycle regulations. Compared with serum-free medium treated cells, Ad5CMV-p16 changed 27 gene expressions, two fold or more on oligonucleotide chip. In addition, Ad5CMV-p16 did not seem to increase the tumorigenicity-related gene expression in A549 cells. Further studies will be needed to investigate the effect of Ad5CMV-p16 on normal human cells and tissues for safety evaluation.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.36 no.6
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• pp.453-459
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• 2010

Introduction: Ameloblastic carcinoma is a rare malignant lesion, and may arise from either carcinoma ex-ameloblastoma or de novo carcinoma. Aberrant promoter hypermethylation of the tumor-associated genes leading to their inactivation is a common event in many cancer types. The p16/CDKN2/INK4A gene and p16 5 protein are involved directly in regulating the cell cycles. Cadherins are cell adhesion molecules that modulate the epithelial phenotype and regulate tumor invasion. The aim of this study was to evaluate the roles of p16 and E-cadherin methylation and loss of p16 and E-cadherin expression in the malignant transformation of an ameloblastoma. Materials and Methods: Eight cases of ameloblastoma, including 4 benign ameloblastomas without recurrence, 2 benign ameloblastomas with recurrence and 2 carcinoma ex-ameloblastomas, were examined. The promoter hypermethylation profile of the p16 and E-cadherin genes was studied using methylation-specific polymerase chain reaction (MSP) and immunohistochemical staining for p16 and E-cadherin expression. Results: 1) Aberrant CpG island methylation of the p16 gene was detected in 3 of the 4 benign ameloblastomas without recurrence and 1 of the 2 benign ameloblastomas with recurrence. 2) Aberrant CpG island methylation of the E-cadherin gene was found in 1 of the 4 benign ameloblastomas without recurrence. 3) A loss of p16 expression was noted in 1 of 4 benign ameloblastomas without recurrence and 1 of 2 carcinoma ex-ameloblastomas. 4) A loss of E-cadherin expression was noted in 2 of the 4 benign ameloblastomas without recurrence, 1 of the 2 benign ameloblastomas with recurrence and 2 of the 2 carcinoma ex-ameloblastomas. 5) A loss of p16 expression was observed in 1 of the 4 cases showing aberrant methylation of the p16 gene. 6) A loss of E-cadherin expression was observed in 3 benign ameloblastoma case showing aberrant methylation of the E-cadherin gene. Conclusion: These results suggest that loss of E-cadherin expression related to the other genetic pathway (not methylation) might be an adjuvant indicator predicting the malignant transformation of an ameloblastoma. However, the number of samples in this study was too small and the relationship between the treatment methods and clinical course were not defined. Therefore, further study will be needed.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.16
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• pp.7161-7165
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• 2015

Background: Esophageal squamous cell carcinoma (ESCC) is a common cancer in the north east of India. The present study concerned the prevalence of human papilloma virus (HPV) in the ESCC in north eastern India and its impact on response to chemotherapy. Materials and Methods: p16 expression, a surrogate marker for HPV infection was assessed in 101 pre-treatment biopsies of locally advanced ESCC, reported from a comprehensive cancer centre in north east India, using immunohistochemistry. All patients received neo-adjuvant chemotherapy. Response was assessed clinically and histopathologically with attention to p16 expression. Results: p16 was expressed in 22% of ESCC (22 out of 101) and was more prevalent in patients who were more than 45 years of age (P=0.048). p16 positive tumors appeared more commonly in the upper 2/3 of the thoracic esophagus (18 in 22). Nine of the 22 (41%) p16 positive tumors achieved pathologic complete response following neo-adjuvant chemotherapy (P=0.008). There was a trend towards reduced mortality in this group (P=0.048). Some 9 of the 20 (45%) patients who achieved pathologic complete response were p16 positive. Conclusions: Expression of p16 in ESCC correlates with higher rate of pathologic complete remission in patients undergoing neo adjuvant chemotherapy and could be a predictive marker for response assessment.