Human Papilloma Virus Attributable Head and Neck Cancer in the Sudan Assessed by p16INK4A Immunostaining

Background: The aim of this study was to screen for human papillomavirus (HPV) infections in head andneck squamous cell carcinomas (HNSCCs) using P16 immunostaining. Materials and Methods: A retrospectivestudy was performed on 150 samples from patients diagnosed with HNSCCs. HPV status was determined usingp16INK4A. Results: 31 of the 150 (20.7%) HNSCCs were HPV positive. Conclusions: A large proportion of HNSCCsin Sudan are associated with HPV infection. The fact that the prevalence of HPV is high among Sudanese patientswith head and neck cancers (HNC) has obvious implications for vaccine therapy.     

Predictive Value of Anti- E6 Oncoprotein (High Risk- Human Papilloma Virus) and p16 Ink4a for Detecting HPV in Oral Epithelial Dysplasia

Aim: The aim of this study was to evaluate the expression of human papillomavirus (HPV) in oral potentially malignant disorders (OPMD) and to examine the association of HPV in histological grades of dysplasia using p16 and Anti-E6 oncoprotein immunohistochemistry (IHC). Subjects and methods: This study focused on clinically diagnosed oral potentially malignant disorders. Clinical parameters such as age, gender, habits, occupation, duration, site, and the type of the lesions were examined and the incisional biopsy was done on the selected cases for the histopathological diagnosis. Selected cases of OPMDs were screened immunohistochemically for HPV 16 and HPV 18 (high-risk group) positivity using p16INK4a and Anti-E6 oncoprotein. The immunohistochemical p16 expression was evaluated based on (a) percentage of p16 positive cases and (b) pattern of p16 staining in various grades of OPMD. Results: Anti-E6 oncoprotein (HR-HPV) expression level was only detected in 11 cases (37%), and positive expression of p16 was found in three cases (10%), with variation in cell proportion and intensity. Subsequently, the association between p16 expression level and clinicopathological characteristic factors was analyzed and a significant association was found between age and histopathology. Conclusion: There was an association between HPV and OPMD. Both biomarker tests, HPV E6 and p16 immunocytochemistry had a specific role in the detection of HR-HPV. Anti-E6 immunocytochemistry can be a valuable test with higher specificity for HPV DNA detection in oral epithelial dysplasia without losing sensitivity.     

Lack of Association between High-risk Human Papillomaviruses and Oral Squamous Cell Carcinoma in Young Japanese Patients

Background: Human papillomaviruses (HPV) may play an important role as one of the possible etiologiesof oral squamous cell carcinoma (OSCC). The present study aimed to investigate the association between HPVand OSCC in young Japanese patients by examining the presence of HPV DNA and surrogate markers in OSCCtissues. Materials and Methods: Forty young patients with OSCC whose surgical specimens were available wereanalyzed and compared with 40 patients randomly recruited from a pool of patients aged >40 years. HPV DNAwas detected using the polymerase chain reaction-based AMPLICOR® HPV test, and surrogate markers ofHPV infection were analyzed using immunohistochemical techniques to detect p16INK4a and p53. Results: Onlytwo (5%) young patients and one (2.5%) older patient were positive for HPV DNA. p16INK4a overexpressionwas identified in six (15%) young patients. p53 staining levels were not high in tissues of most young patients(27 patients, 67.5%). HPV DNA status did not significantly correlate with p16INK4a expression levels. Profiles ofincreased levels of p16INK4a expression with diminished levels of p53 staining were not associated with the presenceof HPV DNA. The combined p53 with p16INK4a profiles were significantly correlated with alcohol consumptionin younger patients (p=0.006). Conclusions: Results of the present study indicate that HPV is less likely to causeOSCC in young Japanese patients, and the p16INK4a expression level is not an appropriate surrogate marker forHPV infection in OSCC.     

Human Papillomavirus 16 and 18 Infection in Oral Cancer in Thailand: A Multicenter Study

Objectives: To identify the prevalence of high-risk human papillomavirus (HPV) genotypes 16 and 18 among patients with oral squamous cell carcinoma (OSCC) in Thailand and investigate the associations of p16 expression and HPV16/18 with the demographic, clinicopathologic, and risk parameters. Materials and Methods: A total of 403 formalin-fixed paraffin-embedded OSCC specimens from four centers in four regions were obtained. p16 expression was evaluated by immunohistochemistry. The detection of HPV16/18 DNA was performed by polymerase chain reaction.  Results: Of all, 172 specimens (42.7%) were presented with amplifiable extracted DNA. Among these, 62.8% were positive for p16, 8.1% were positive for HPV16/18, and 5.8% were positive for both methods. Of all HPV-positive specimens, HPV18 was detected in 57.1%; HPV16 in 14.3%; and HPV16 and 18 (co-infection) in 28.6%. The prevalence of HPV16/18 varied between centers, with the highest rate in the northern center (20.0%). There was no significant correlation between p16 expression and HPV16/18. There were no significant associations of p16 expression and/or HPV16/18 with all variables. Conclusions: The prevalence of HPV16/18 infection in OSCC geographically varied in Thailand, with the highest rate in the northern region. Poor correlation between p16 and HPV16/18 suggests p16 not be used as a surrogate marker for HPV-positive OSCC.     

Human papillomavirus infection as a prognostic factor in carcinomas of the oral cavity and oropharynx

Although studies have established human papillomaviruses (HPVs) as a risk factor for oral and oropharyngeal cancer, it is not clear whether viral infection affects survival in head and neck malignancies. This investigation examined the relationship between HPV and survival in carcinomas of the oral cavity and oropharynx. Formalin-fixed, paraffin-embedded tumor specimens from 139 newly diagnosed cases were tested for HPVs by PCR and DNA sequencing. Patient and tumor characteristics were obtained from questionnaires, pathology reports and cancer registries. Odds ratios (ORs) and relative risks (RRs) were based on logistic and Cox regression models, respectively. HPVs were detected in 21% of the tumors; 83% were HPV-16. Greater risk of HPV infection was associated with males (OR = 2.9, 95% CI = 1.0-8.6), a history of oral-genital sex (OR = 4.2, 95% CI = 1.5-11.7), and oropharyngeal tumors (OR = 10.4, 95% CI = 3.5-31.2). As tobacco usage increased, the odds of HPV detection decreased (OR = 0.97/pack-year, 95% CI = 0.96-0.99). HPV infected patients had better overall survival (RR = 0.3, 95% CI = 0.1-0.8) than those with HPV-negative tumors. There was an interaction between gender and HPV for overall (p = 0.05) and disease-specific (p = 0.03) survival that suggested that HPV infected males had better prognosis than HPV-negative males, but this was not the case among females. HPV status was identified as an independent prognostic factor in oral and oropharyngeal cancers. This result appeared to be gender-specific, suggesting the need for further study of the interaction between HPV and gender on survival.     

Frequent p16INK4a promoter hypermethylation in human papillomavirus-infected female lung cancer in Taiwan

Inactivation of p16INK4a gene through promoter hypermethylation has been frequently observed in non small cell lung cancer; however, various studies have shown a controversial correlation between p16INK4a hypermethylation and cigarette smoking. Our recent report showed that human papillomarvirus (HPV) 16/18 infections were associated with the development of nonsmoking female lung cancer in Taiwan and we further speculated that HPV infection may be linked with p16INK4a hypermethylation. To verify the influence of environmental exposure, including cigarette smoking, environmental carcinogen exposure and HPV infections on p16INK4a hypermethylation, tumors from 162 lung patients, including 67 smoking males, 41 nonsmoking males and 58 nonsmoking females, were subjected to p16INK4a hypermethylation analysis by methylation-specific PCR. As the results showed, p16INK4a hypermethylation was detected in 40 (59.7%) of 67 smoking male, 15 (36.6%) of 41 nonsmoking male and 35 (60.3%) of 58 nonsmoking female lung tumors. This result seemed to reveal that gender and cigarette smoking both possess an equal influence on p16INK4a hypermethylation. This result also led to a speculation that HPV infection may promote p16INK4a hypermethylation in nonsmoking female lung cancer patients. From our data, p16INK4a hypermethylation frequency in nonsmoking female lung tumors with HPV infection was as high as 70% (30 of 43) compared to those without HPV infection (33%; 5 of 15). In fact, the correlation between HPV infection and p16INK4a hypermethylation was only observed in nonsmoking female lung tumors (p = 0.017), but not in smoking male or nonsmoking male lung tumors. Moreover, the reverse correlation between p16INK4a immunostaining and p16INK4a promoter hypermethylation was also only observed in nonsmoking female lung tumors. These results strongly suggested that the involvement of HPV infection in lung tumorigenesis of nonsmoking female cancer patients in Taiwan may be mediated at least in part through the increase of hypermethylation to cause p16INK4a inactivation.     

Roles of Human Papillomaviruses and p16 in Oral Cancer

Head and neck cancer, including oral cancer, is the sixth most common cancer in humans worldwide. Morethan 90% of oral cancers are of squamous cell carcinoma type. Recent studies have shown a strong relationshipbetween human papillomavirus (HPV) infection and head and neck cancer, especially oropharyngeal squamouscell carcinoma (OPSCC) and oral squamous cell carcinoma (OSCC). Moreover, the incidence of HPV-relatedOSCC appears to be on the rise while HPV-unrelated OSCC tends to have stabilized in the past decades. p16, atumor suppressor gene, normally functions as a regulator of the cell cycle. Upon infection with high-risk typesof HPV (HR-HPV), particularly types 16, 18, 31, 33, 34, 35, 39, 51, 52, 56, 58, 59, 66, 68, and 70, the expressionof p16 is aberrantly overexpressed. Therefore, the expression of p16 is widely used as a surrogate marker forHPV infection in head and neck cancer.     

Comparison of p16INK4a Immunocytochemistry with the HPV Polymerase Chain Reaction in Predicting High Grade Cervical Squamous Intraepithelial Lesions

Aim: To compare p16INK4a immunocytochemistry with the HPV polymerase chain reaction in predictinghigh grade cervical squamous intraepithelial lesions. Materials and Methods: This diagnostic case-control studywas conducted from January 2010 until December 2010. We obtained 30 samples, classified according to thedegree of cervical intraepithelial neoplasia (CIN): 11 samples for CIN 1, 9 samples for CIN 2, and 10 samples forCIN 3. HPV PCR, p16INK4a immunocytochemistry, and histopathological examination were performed on allsamples. Statistical analysis was conducted using SPSS 20.0. Results: In predicting CIN 2-3, we found p16INK4ato have similar specificity and positive predictive value as HPV PCR (95%, 97.2% vs 96.7%), but better sensitivity(87.5% vs 72.5%) and negative predictive value (82.1% vs 67.6%). The most prevalent types of high-risk HPVin our study were HPV 33, 35, 58, 52, and 16. Conclusions: p16INK4a has better diagnostic values than HPVPCR and may be incorporated in the triage of ASCUS and LSIL to replace HPV PCR. Genotype distribution ofHPV differs in each region, providing a challenge to develop HPV vaccines based on the epidemiology of HPVin that particular region.     

Assessment of the Prognostic Value of Methylation Status and Expression Levels of FHIT,GSTP1 and p16 in Non-Small Cell Lung Cancer in Egyptian Patients

Background: Methylation of tumor suppressor genes has been investigated in all kinds of cancer. Tumorspecific epigenetic alterations can be used as a molecular markers of malignancy, which can lead to betterdiagnosis, prognosis and therapy. Therefore, the aim of this study was to evaluate the association between genehypermethylation and expression of fragile histidine triad (FHIT), glutathione S-transferase P1 (GSTP1) andp16 genes and various clinicopathologic characteristics in primary non-small cell lung carcinomas (NSCLC).Materials and Methods: The study included 28 primary non-small cell lung carcinomas, where an additional28 tissue samples taken from apparently normal safety margin surrounding the tumors served as controls.Methylation-specific polymerase chain reaction (MSP) was performed to analyze the methylation status ofFHIT, GSTP1 and p16 while their mRNA expression levels were measured using a real-time PCR assay withSYBR Green I. Results: The methylation frequencies of the genes tested in NSCLC specimens were 53.6% forFHIT, 25% for GSTP1, and 0% for p16, and the risk of FHIT hypermethylation increased among patientswith NSCLC by 2.88, while the risk of GSTP1 hypermethylation increased by 2.33. Hypermethylation of FHITgene showed a highly significant correlation with pathologic stage (p<0.01) and a significant correlation withsmoking habit and FHIT mRNA expression level (p<0.05). In contrast, no correlation was observed between themethylation of GSTP1 or p16 and smoking habit or any other parameter investigated (p>0.05). Conclusions:Results of the present study suggest that methylation of FHIT is a useful biomarker of biologically aggressivedisease in patients with NSCLC. FHIT methylation may play a role in lung cancer later metastatic stages whileGSTP1 methylation may rather play a role in the early pathogenesis.     

胃癌p16基因外显子1甲基化状态的研究

目的 研究抑癌基因p165‘CpG岛甲基化状态及其与胃癌发生、发展及浸润转移的关系。方法 对46例胃癌组织和30例良性病变胃粘膜（对照组）分别采用甲基化敏感性限制性内切酶（Hpa Ⅱ,MspⅠ,SacⅡ)酶切后PCR扩增进行p16基因甲基化多位点检测。结果 46例胃癌组织,p16基因甲基化13例（28．3％）,30例对照组无p16基因甲基化,两者比较有显著性差异（P＜0．05）。23例胃癌浸润未达肌层患者p16基因甲基2例（8．7％）,23例达肌层或浆膜层患者p16基因甲基化11例（47．8％）,两者比较有显著性差异（P＜0．05）。20例高中分化胃癌患者无p16基因甲基化,17例低分化胃癌p16基因甲基化13(76．5％）,两者比较差异有显著性（P＜0．05）。18例呈结节生长型胃癌p16基因甲基化6例（33．3％）,28例局限溃疡或溃疡浸润26例胃角、胃窦癌p16基因甲基化8例（30．8％）,两者比较无显著性差异（P＞0．05）。9例伴肝或淋巴结转移者胃癌p16基因甲基4例（44．4％）,37例无转移者p16基因甲基化9例（24．3％）,两者比较无显著性差异（P＞0．05）。结论 抑癌基因p16 5‘CoG岛甲基化在胃癌发生中起重要作用,并与胃癌浸润深度、分化程度有关,但与肿瘤的生长方式、部位及转移无关。     

P16ink4a Subcellular Expression Patterns in Colorectal Adenocarcinoma,Adenoma and Non-Neoplastic Tissue Samples

Background: Colorectal cancer (CRC) is one of the most common neoplasms with high mortality at advanced stages worldwide. Thus diagnosis of CRC at an early stage with sensitive molecular methods is a high priority. The aim of this study was to evaluate P16ink4a subcellular expression patterns in colorectal adenocarcinoma, adenoma and non-neoplastic tissue samples. Methods: A total of 137 colorectal formalin fixed paraffin-embedded tissue blocks from the pathology archives of Ali-Ebne-Abitaleb central hospital, Zahedan, Iran, were examined in three groups: adenocarcinoma (n= 63), adenoma (n= 38) and non-neoplastic (n= 36). The subcellular expression pattern was determined by immunocytochemistry. Data analysis was performed using Kruskal-Wallis and Fisher exact tests with the significance level set as p˂0.05. Results: P16ink4a subcellular localization was observed in three different patterns, nuclear+cytoplasmic (73.33%), cytoplasmic (13.33%) and nuclear (13.33%). In most samples, nuclear+cytoplasmic was the predominant subcellular pattern. However, a significant difference in P16ink4a subcellular expression patterns was observed along the non-neoplastic, adenoma, adenocarcinoma sequence (p˂0.001). An association with the histological tumor type was also noted (p=0.021). Conclusion: Considering variation in localization of P16ink4a under different pathological conditions, P16ink4a night be sensitive prognostic biomarker for benign colon lesions. Its use may improve strategies for screening, prognostic assessment and management of patients with CRC. Further studies are recommended in this field.     

HIV阳性女性患者P16INK4a蛋白表达与HPV感染相关性分析

[目的]了解HIV阳性感染者宫颈脱落细胞P16INK4a蛋白表达与HPV感染的相关性。[方法]采用通用引物PCR和反向点杂交技术建立的检测方法,对182例HIV阳性感染者和300例HIV阴性感染者检测23种HPV亚型,包括18种高危型和5种低危型。同时采用免疫组织化学技术(S-P染色法)检测宫颈脱落细胞中P16INK4a蛋白表达情况。[结果]HIV阳性女性HPV感染率(36.8%)及P16INK4a蛋白阳性表达率(18.7%)较HIV阴性者(19.3%、4.3%)高(P均为0.000)。HIV阳性患者中,HPV感染阳性者宫颈脱落细胞P16INK4a蛋白表达率为43.3%,HPV阴性者4.3%(P=0.000);HPV多型感染者宫颈脱落细胞P16INK4a蛋白表达率(60.9%)较HPV单型感染者高(34.1%)(P=0.042);高危型HPV感染者宫颈脱落细胞P16INK4a蛋白表达率(80.6%)较低危型HPV感染者高(0)(P=0.000)。HIV阴性患者中,HPV感染阳性者P16INK4a蛋白阳性表达率(5.2%)较HPV阴性者(1.3%)高(P=0.721);所检HPV阴性患者均为单一HPV感染;高危型HPV感染患者P16INK4a蛋白阳性表达率(40.0%)较低危型(1.9%)明显增高(P=0.018)。[结论]HIV阳性女性中HPV感染阳性、多型HPV感染及高危型HPV感染宫颈细胞P16INK4a蛋白的表达率更高。     

P16INK4a as an adjunct marker in liquid-based cervical cytology

Cytological screening for cervical cancer is hampered by high false negative rates. Inter-observer reproducibility needs optimizing. The potential of p16(INK4a) as a biomarker for cervical lesions was examined in a study of liquid-based cytology (LBC), HPV DNA testing by MY09/MY11 consensus PCR and type-specific PCRs and p16(INK4a) immunocytochemistry on a series of 291 patients selected from routine screening. Comparison of the number of p16(INK4a) immunoreactive cells/1,000 cells exhibited a significantly higher mean count in HSIL (8.80 +/- 1.13) than other cytological groups. The mean count of LSIL (1.09 +/- 0.18) was significantly higher than that of the negative group (0.82 +/- 0.40). ASC-H and HSIL combined showed a significantly higher mean count (6.46 +/- 1.17) than negative, ASC, ASC-US and LSIL. The mean count of immunoreactive cells/1,000 cells was significantly higher in HPV16 positive samples (3.22 +/- 0.72) than in samples containing infections with types of unknown malignant potential (0.83 +/- 0.26) or HPV negative samples (1.17 +/- 0.41). The mean count in infections with other high-risk HPV types (2.55 +/- 0.52) was significantly higher than that in HPV negative samples. Receiver-operating characteristic curves yielded a test accuracy (area under curve) of 0.76, 0.79, 0.88 and 0.95 for ASCUS, LSIL, ASC-H/HSIL and HSIL, respectively. Thresholds for 95% sensitivity were at 0.005, 0.007, 0.098 and 0.445 immunopositive cells/1,000 cells for ASCUS, LSIL, ASC-H/HSIL and HSIL, respectively. The 95% specificity threshold for the detection of HSIL was at 1.87 immunopositive cells/1,000 cells. P16(INK4a) immunocytochemistry can be used as an adjunct to LBC in cervical screening, because it has a good diagnostic accuracy to discriminate HSIL and ASC-H from other lesions. It could be used as a surrogate marker of high-risk HPV infections.     

P16INK4A和PTEN在高危型HPV相关宫颈癌组织中的表达及意义

背景与目的:高危型人乳头状瘤病毒(high-risk human papillomavirus,HR-HPV)是宫颈癌最主要的致病因素,目前研究发现,在宫颈上皮癌变过程中,P16INK4A的异常表达和HR-HPV感染密切相关;同时,另一抑癌基因PTEN也参与了宫颈上皮肿瘤的形成.本研究旨在探讨宫颈上皮癌变过程中P16INK4A、PTEN表达与HR-HPV感染的关系及其意义.方法:应用免疫组织化学方法检测P16INK4A蛋白和PTEN蛋白在30例正常宫颈组织、11例原位癌、24例宫颈浸润癌组织中的表达.用第二代杂交捕获法(HC-2)检测每一病例的13种HR-HPV DNA.结果:HR-HPV和P16INK4A阳性率浸润癌组(91.7%、87.5%)和原位癌组(90.9%、81.8%)都明显高于正常宫颈组(30.0%、6.7%),差异有统计学意义(P＜0.001).P16INK4A过表达(中、强阳性)和HR-HPV阳性同时出现有30例,其中原位癌组9例,浸润癌组21例;两者同时阴性有23例,其中正常宫颈组20例,原位癌组1例,浸润癌组2例.相关性分析结果显示,HR-HPV感染与P16INK4A表达呈正相关(rs=0.690,P＜0.001).26例PTEN中、强阳性表达均在正常宫颈组,其阳性率在浸润癌组(37.5%)和原位癌组(36.4%)明显低于正常宫颈组(83.3%),差异有统计学意义(P＜0.01).相关性分析证实,HR-HPV感染与PTEN表达的相关性无统计学意义(rs=-0.174,P=0.167).结论:在HR-HPV感染的宫颈癌中,P16INK4A出现过表达,其蛋白的肿瘤抑制功能不明显;PTEN独立于HR-HPV途径,以其功能下调促进宫颈癌的发生、发展.     

结肠癌细胞株、癌组织及正常结肠粘膜中P16基因异常甲基化率的差异

本文采用Southernblot法鉴定了结肠癌细胞株、癌组织及正常结肠粘膜组织中P16基因的甲基化情况。结果在6株结肠癌细胞株中有5株(833%)P16基因CpG岛呈异常甲基化状态,在18例结肠癌组织中13例(722%)呈异常甲基化状态。与之相对,18例正常结肠粘膜组织中仅5例(278%)表现为异常甲基化,提示在结肠癌中P16基因因异常甲基化而失活,这为结肠癌的基因治疗提供了新策略、新目标。     

Correlation between human papillomavirus and p16 overexpression in oropharyngeal tumours: a systematic review

Background:

A significant proportion of squamous cell carcinomas of the oropharynx (OP-SCC) are related to human papillomavirus (HPV) infection and p16 overexpression. This subgroup proves better prognosis and survival but no evidence exists on the correlation between HPV and p16 overexpression based on diagnostic measures and definition of p16 overexpression. We evaluated means of p16 and HPV diagnostics, and quantified overexpression of p16 in HPV-positive and -negative OP-SCCs by mode of immunohistochemical staining of carcinoma cells.

Methods:

PubMed, Embase, and the Cochrane Library were searched from 1980 until October 2012. We applied the following inclusion criteria: a minimum of 20 cases of site-specific OP-SCCs, and HPV and p16 results present. Studies were categorised into three groups based on their definition of p16 overexpression: verbal definition, nuclear and cytoplasmatic staining between 5 and 69%, and ⩾70% staining.

Results:

We identified 39 studies with available outcome data (n=3926): 22 studies (n=1980) used PCR, 6 studies (n=688) used ISH, and 11 studies (n=1258) used both PCR and ISH for HPV diagnostics. The methods showed similar HPV-positive results. Overall, 52.5% of the cases (n=2062) were HPV positive. As to p16 overexpression, 17 studies (n=1684) used a minimum of 5–69% staining, and 7 studies (n=764) used ⩾70% staining. Fifteen studies (n=1478) referred to a verbal definition. Studies showed high heterogeneity in diagnostics of HPV and definition of p16. The correlation between HPV positivity and p16 overexpression proved best numerically in the group applying ⩾70% staining for p16 overexpression. The group with verbal definitions had a significantly lower false-positive rate, but along with the group applying 5–69% staining showed a worse sensitivity compared with ⩾70% staining.

Conclusions:

There are substantial differences in how studies diagnose HPV and define p16 overexpression. Numerically, p16 staining is better to predict the presence of HPV (i.e. larger sensitivity), when the cutoff is set at ⩾70% of cytoplasmatic and nuclear staining.     

The p16INK4a Antibody Immobilization Method for Immonosensor Application

Background: The p16INK4a is a protein that expressed in Liquid-based cervical cytology specimens and hasbeen proved link to cervical cancer. The p16INK4a could be detection by piezoelectric immunosensor and theimmobilization of the p16INK4a antibody influence the sensitivity of the piezoelectric immunosensor. Materials andMethods: 5μL mouse polyclonal antibody against p16INK4a was bound onto the surface of immonosensor throughtwo methods. (directly immobilized method; protein A method). Absorb of the p16INK4a antibody on the surfaceof immonosensor caused a shift in the resonant frequency of the immunosensor and The frequency changesrecorded showed a better reproducibility. The activity of the immobilization antibody with the directly methodand protein A method was tested with p16INK4a antigen. Results: The resonant frequency for different antibodyimmobilization methods were different, and the sensitivity for p16INK4a detection also different. Conclusions:The protein A method was found to be much more better than the directly method for the immobilization ofthe p16INK4A antibody on the gold electrode of the quartz crystal for cervical lesion detection. The Protein Amethod created more reproducible and stable immobilization antibody layers with p16INK4A antigen.     

p16INK4A Expression in Condyloma Acuminata Lesions Associated with High-Risk Human Papillomavirus Infection

Objective: The objective of this study was to discover the possible correlation between p16INK4A expression and the LR/HR-HPV infection in condyloma acuminate (CA) lesions. Materials and Method: This cross-sectional study was conducted during January-December 2017 on 33 CA patients. The expression of p16INK4A was detected by immunohistochemistry (IHC) staining. The positive interpretation was carried out by scoring which score 0 was negative, score 1 was sporadic, score 2 was focal, and score 3 was diffuses. The HPV genotypes were identified by reverse line blot, and 40 genotypes of HPV detected, including HR-HPV (HPVs 16, 18, 26, 31, 33,35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 67, 68a, 68b, 69, 73, and 82) and LR-HPV (HPVs 6, 11, 40, 42, 43, 44, 54, 55, 61, 62, 64, 70, 71, 72, 81, 83, 84, 87, 89, and 90). Results: The expression of p16INK4A was significantly correlated with HR-HPV infection. Patients infected with HR-HPV had 0.644 times higher possibility to express p16INK4A gene compared to those infected with LR-HPV. LR-HPV genotypes detected in CA patients were HPVs 6, 11, 42, 61, 54, 81, 87, 89, and 90 and HR-HPV genotypes were HPVs 18, 26, 45, 51, 52, 67, 68B, 69, and 82. LR-HPV was found in 19/33 of patients and HR-HPV was in 14/33 of patients. The expression of p16INK4A in CA lesions was diffuse in15.2% of patients, was focal in 24.2% of patients , was sporadic in 39.4% of patients were, and was negative in 21.2% of patients . In LR-HPV group, there was no diffuse expression, focal expression was observed in 15.8%, sporadic in 47.4%, and negative in 36.8%, while in HR-HPV group, p16INK4A expression was detected in all lesions , in a way that its expression was diffuse in 35.7%, focal in 35.7%, and sporadic in 28.6%. Conclusion: IHC is a routine method in histopathological diagnosis, therefore the detection of p16INK4A expression by IHC can be used as a biomarker for HR-HPV infection diagnosis.     

Upregulation of p16INK4A in Peripheral White Blood Cells as a Novel Screening Marker for Colorectal Carcinoma

Objective: Screening of colorectal cancer (CRC) is important for the early detection. CRC is relating to aging and immuno-senescence. One such senescent marker is p16INK4A expression in immune cells. The objective of the study is to investigate the protein expression of p16INK4A in peripheral white blood cells as a screening marker for colorectal cancer. Methods: A case-control studies were conducted. Cases were patients with colorectal cancer and controls were matched with cases based on age and sex. Peripheral blood was collected from patients and controls and the protein p16INK4A was measured with immunofluorescent techniques. The p16INK4A levels from cases and controls were evaluated using ROC analysis to be used as a screening marker in CRC patients. Mean fluorescent intensity of p16INK4A of cases and controls were analyzed in CD45+, CD3+ or CD14+ cells. The p16INK4A levels of cases were also correlated with clinical data. Result: Statistically significant increased expression of p16INK4A levels were found in cases compared to controls. p16INK4A in peripheral immune cells had 78% sensitivity and 71% specificity which can possibly be used as a diagnosis tool for colorectal cancer. P16INK4A-positive cell percentage and mean florescent intensity were significantly higher in CD45+ cells, CD3 positive cells and CD14 positive cells. No significant correlation was observed with the clinical data and p16INK4A level of CRC patients. Conclusion: The significant increase of p16 INK4A expression level in peripheral immune cells represents potential for use as a CRC screening marker.