Cervical Intraepithelial Neoplasia grade 2 biopsy: Do p16INK4a and Ki-67 biomarkers contribute to the decision to treat? A cross-sectional study

ABSTRACT BACKGROUND: Managing cervical intraepithelial neoplasia grade 2 (CIN2) is challenging, considering the CIN2 regression rate, perinatal risks associated with excisional procedures, and insufficient well-established risk factors to predict progression. OBJECTIVES: To determine the ability of p16INK4a and Ki-67 staining in biopsies diagnosed with CIN2 to identify patients with higher-grade lesions (CIN3 or carcinoma). DESIGN AND SETTING: Cross-sectional study conducted at a referral center for treating uterine cervical lesions. METHODS: In 79 women, we analyzed the correlation of p16INK4a and Ki-67 expression in CIN2 biopsies with the presence of a higher-grade lesions, as determined via histopathology in surgical specimens from treated women or via two colposcopies and two cytological tests during follow-up for untreated women with at least a 6-month interval. The expression of these two biomarkers was verified by at least two independent pathologists and quantified using digital algorithms. RESULTS: Thirteen (16.8%) women with CIN2 biopsy exhibited higher-grade lesions on the surgical excision specimen or during follow-up. p16INK4a expression positively and negatively predicted the presence of higher-grade lesions in 17.19% and 86.67% patients, respectively. Ki-67 expression positively and negatively predicted the presence of higher-grade lesions in 40% and 88.24% patients, respectively. CONCLUSIONS: Negative p16INK4a and Ki67 immunohistochemical staining can assure absence of a higher-grade lesion in more than 85% of patients with CIN2 biopsies and can be used to prevent overtreatment of these patients. Positive IHC staining for p16INK4a and Ki-67 did not predict CIN3 in patients with CIN2 biopsies.

Diagnostic value of combined detection of p16 and human papillomavirus typing in cervical intraepithelial neoplasia / 肿瘤研究与临床

Objective:To explore the application value of combined detection of p16 and human papillomavirus (HPV) typing in the diagnosis of cervical intraepithelial neoplasia (CIN).Methods:A total of 8 346 patients aged between 25 years old and 65 years old at Baoji Central Hospital of Shaanxi Province from February 2019 to February 2020 were selected. There were 2 882 patients with cervical lesions diagnosed by colposcopy biopsy. Patients were divided into the different groups based on the age range, and then the condition of HPV infection in all age groups was analyzed. Taking biopsy as the gold standard and according to the pathological results, the detection rate of p16 and HPV typing and the diagnostic value of the single and combined detection in CIN were also analyzed.Results:The age group with the highest positive rate of p16 and HPV was 31-40 years old [47.42% (1 014/2 427) and 36.84% (894/2 427), respectively], followed by 41-50 years old group [30.15% (907/2 942) and 28.11% (827/2 942)], and there were statistically significant differences in positive rate of p16 and HPV in all age groups (all P < 0.05). Among 2 882 patient with cervical lesions diagnosed by pathological examination, there were 2 572 cases (89.24%) of p16 positive, and 2 169 cases (75.26%) of HPV positive. With the disease progression of cervical lesions, the positive rate of p16 and HPV was gradually increased, and the positive rate of p16 of inflammation, CINⅠ, CINⅡ, CIN Ⅲ, cervical squamous cell carcinoma (SCC) was 11.68% (23/197), 94. 85% (1 105/1 165), 93.57% (771/824), 96.76% (538/556), 96.43% (135/140), respectively; the positive rate of HPV was 17.77% (35/197), 77.60% (904/1 165), 80.22% (661/824), 80.40% (447/556), 87.14% (122/140), respectively, and HPV infection was mostly HPV16/18 infection type with the disease progression of cervical lesions. The sensitivity, specificity, positive predictive value and negative predictive value in detecting CIN of HPV was 75.26%, 81.13%, 67.78% and 86.14%, respectively; the sensitivity, specificity, positive predictive value and negative predictive value in detecting CIN of p16 was 89.24%, 84.74%, 75.51% and 93.72%, respectively; the diagnostic efficacy of p16 was higher than that of HPV in detecting CIN, and the difference was statistically significant ( P < 0.05). The sensitivity, specificity, positive predictive value and negative predictive value of HPV combined with p16 in detecting CIN was 94.10%, 91.33%, 85.12%, 96.71%, which were higher compared with those of single detection (all P < 0.05). Conclusions:HPV infection mainly occurs in women aged 31-40 years old followed by 41-50 years old, and the infected population of CIN tends to be younger. p16 is superior to HPV in detecting the diagnostic efficacy of CIN; combined detection of p16 and HPV can increase the sensitivity and specificity, reduce the rate of misdiagnosis, and can play a key clinical value in early diagnosis and treatment of CIN.

Cervical squamous intraepithelial lesions grades 1 and 2: value of an alternative morphological classification and of p16 immunohistochemistry in the prediction of clinical outcome / Lesões intraepiteliais escamosas cervicais graus 1 e 2: valor de uma classificação morfológica alternativa e do estudo imuno-histoquímico com p16 na predição do desfecho clínico

ABSTRACT Introduction: Diagnostic reproducibility and determination of prognostic factors in cervical intraepithelial neoplasias grades 1 and 2 are still relevant problems in the daily practice of gynecological histopathology. Objective: To correlate the value of morphological reclassification and of p16 immunoexpression in cervical intraepithelial neoplasias grades 1 and 2 with clinical outcome. Materials and methods: Sixty-six patients were included (34 with cervical intraepithelial neoplasia grade 1, and 32 with grade 2); an immunohistochemical study with p16 and reclassification according to the Lower Anogenital Squamous Terminology (LAST) Consensus and by the alternative proposal of Herfs and Crum were done; unfavorable outcome was defined as a subsequent histologic diagnosis of cervical intraepithelial neoplasia grade 3 or invasive squamous cell carcinoma. Results: We observed superior performance of the alternative morphological classification (p = 0.002) to determine unfavorable outcome. We also detected superior performance of p16 in the same determination (p = 0.002). Conclusion: The use of an alternative morphological classification is promising; in the context of the use of immunohistochemical antibodies as biomarkers, p16 showed good sensitivity and negative predictive value in the determination of cases in which the outcome was unfavorable.

RESUMO Introdução: A reproducibilidade diagnóstica e a determinação de fatores prognósticos em neoplasias intraepiteliais cervicais graus 1 e 2 ainda são problemas relevantes na prática diária da histopatologia ginecológica. Objetivo: Correlacionar o valor da reclassificação morfológica e da imunoexpressão do marcador p16 em neoplasias intraepiteliais cervicais graus 1 e 2 com o desfecho clínico. Materiais e métodos: Incluídas 66 pacientes (34 com neoplasia intraepitelial cervical grau 1 e 32 com grau 2); realizou-se estudo imuno-histoquímico com p16 e reclassificação segundo o Consenso Lower Anogenital Squamous Terminology (LAST) e a proposta alternativa de Herfs e Crum; desfecho desfavorável foi definido como diagnóstico histológico subsequente de neoplasia intraepitelial cervical grau 3 ou carcinoma de células escamosas invasivo. Resultados: Observamos performance superior da classificação morfológica alternativa (p = 0,002) para determinação de desfecho desfavorável. Também detectamos performance superior do marcador p16 na mesma determinação (p = 0,002). Conclusão: A utilização de uma classificação morfológica alternativa é promissora. No âmbito da utilização dos anticorpos imuno-histoquímicos como biomarcadores, o p16 apresentou boa sensibilidade e valor preditivo negativo na determinação dos casos em que o desfecho foi desfavorável.

Significance of p16/Ki-67 double immunocytochemical staining in cervical cytology ASCUS, LSIL,and ASC-H / 中华妇产科杂志

Objective To investigate the application value of p16/cell proliferation associated nuclear antigen (Ki-67) double-staining and human papillomavirus mRNA in the cytological screening.Methods Two hundred and fifty-one cases who suffered from atypical squamous cell of undetermined significance (ASCUS),low-grade squamous intraepithelial lesion (LSIL),atypical squamous cell-cannot exclude high-grade squamous intraepithelial lesion (ASC-H) in ThinPrep cytologic test (TCT) were collected in Peking University First Hospital between October 2015 and March 2016.And p16/Ki-67 double-staining and hybrid capture Ⅱ (HC-Ⅱ) detection were performed on the cervical cells.The result was compared with the pathological result of colposcope guided biopsy.All statistical analysis was completed by Stata 12.0 statistical software analysis.The results of diagnostic tests were described by using the sensitivity,specificity,positive predictive value,negative predictive value,and the area under the receiver operating characteristic (ROC) curve.Results (1) One hundred and eight cases of liquid based cytology diagnosis of ASCUS patients,the positive rate of p16/Ki-67 was 13.9％ (15/108),102 cases of liquid based cytology diagnosis of LSIL patients,the positive rate of p16/Ki-67 was 21.6％ (22/102),41 cases of liquid based cytology diagnosis of ASC-H patients,the positive rate of p16/Ki-67 was 39.0％ (16/41),compared amongthree groups,the difference was statistically significant (x2=78.516,P＜0.05);cervical exfoliated cells p16/Ki-67 expression rate was 13.0％ (28/215) in cervical low-grade lesions [cervical intraepithelial neoplasia (CIN)Ⅰ],which was 69.4％ (25/36) in high level lesions (CIN Ⅱ-Ⅲ),the difference was statistically significant (x2=7.932,P＜0.05).(2) The specificity of p16/Ki-67 detection and diagnosis were higher than those of HC-Ⅱ in ASCUS,LSIL,and ASC-H (89.8％ vs 71.4％,83.3％ vs 15.6％,88.9％ vs 40.7％;all P＜0.05),meanwhile,the positive predictive value of p16/Ki-67 detection and diagnosis exceed those of HC-Ⅱ in ASCUS,LSIL,and ASC-H (33.3％ vs 26.3％,31.8％ vs 12.6％,81.3％ vs 38.5％;all P＜0.05).Moreover,the ROC curve of p16/Ki-67 were bigger than those of HC-Ⅱ in ASCUS,LSIL,and ASC-H (0.799 vs 0.696,0.708 vs 0.531,0.909 vs 0.561;all P＜0.05).Conclusion For patients with cytological diagnosis of ASCUS,LSIL,and ASC-H,p16/Ki-67 double staining method could be used as an effective method to assist in the diagnosis of high-grade cervical lesions,and the screening efficiency is superior to that of high-rist HPV.

Application of p16/Ki-67 immunocytochemistry in triage of patients with atypical squamous cells of undetermined significance / 中华病理学杂志

Objective@#To investigate the clinical value of p16/Ki-67 immunocytochemistry in patients with atypical squamous cells of undetermined significance(ASC-US).@*Methods@#One hundred and seventy-one cases of thin-prep cytology test (TCT) diagnosed as ASC-US underwent p16/Ki-67 immunocytochemistry. All patients had colposcopy and biopsy from March 2015 to January 2016. Ninety of the 171 cases underwent high-risk HPV test at the same time.@*Results@#p16/Ki-67 immunocytochemistry was positive in 43.9% (75/171) of the 171 cytology samples; the sensitivity and specificity of p16/Ki-67 immunocytochemistry were 77.6%(52/67) and 77.9%(81/104) in detecting CIN2+ , and the positive and negative predictive value were 69.3%(52/75) and 84.4%(81/96), respectively. The sensitivity and specificity in diagnosing CIN2+ were 100.0%(34/34) and 10.7%(6/56) for HPV test, and the positive and negative predictive value were 40.5%(34/84) and 6/6. p16/Ki-67 immunocytochemistry showed lower sensitivity but obviously higher specificity than high-risk HPV detection.@*Conclusion@#p16/Ki-67 immunocytochemistry is a good triage test for identifying CIN2+ in ASC-US specimens.

Human Papillomavirus Genotyping and p16INK4a Expression in Cervical Lesions: A Combined Test to Avoid Cervical Cancer Progression

Cervical cancer is a major public health problem in Morocco. The cervical cancer has a long precancerous period that provides an opportunity for the screening and treatment. Improving screening tests is a priority goal for the early diagnosis of cervical cancer. This study was conducted to evaluate the combination of p16INK4a protein expression, human papillomavirus (HPV) typing, and histopathology for the identification of cervical lesions with high risk to progress to cervical cancer among Moroccan women. A total of 96 cervical biopsies were included in this study. Signal amplification in situ hybridization with biotinylated probes was used to detect HPV. Immunohistochemistry was used to evaluate the expression of p16INK4a protein. HPV DNA was detected in 74.0% of the biopsies (71/96). Of the seventy-one positive HPV cases, we detected 67.6% (48/71) of high risk (HR)-HPV (HPV 16 and 18), 24% of low risk-HPV (HPV 6 and 11), 1.4% intermediate risk-HPV (HPV 31, 33, and 35), and 7% coinfections (HPV 6/11 and 16/18). Overexpression of p16INK4a protein was observed in 72.9% (70/96) of the biopsies. In addition, p16INK4a protein detection was closely correlated with recovery of HR HPV. Our result showed that p16INK4a expression level is correlated with HR-HPV status.

The natural history of cervical intraepithelial neoplasia Ⅰ and the clinical significance of p16INK4a protein as a marker of progression in cervical intraepithelial neoplasiaⅠ / 中华妇产科杂志

Objective To describe the natural history of cervical intraepithelial neoplasia(CIN)Ⅰand the biologic factors associated with the progression of CINⅠ and to analyze the predictive values of p16INK4a protein for the progression of CINⅠ. Methods From August 2010 to July 2013, 104 patients referred for abnormal cytology [≤ low-grade squamous intraepithelial lesion (LSIL); including negative for intraepithelial lesion or malignancy (NILM), atypical squamous cells of undetermined significance (ASCUS), LSIL] and high-risk (HR) HPV positive,and were diagnosed CINⅠ by colposcopy-assisted biopsy and followed at 1-year intervals in the First Affiliated Hospital of Nanjing Medical University. In order to analyze the relationship between the progression of CINⅠ with clinical biologic factors, including patient age, cervical cytology before colposcopy, loads of HR HPV, HPV L1 capsid protein, p16INK4a protein,χ2 tests was used to compare the different frequencies of factors in groups of progressed and persisted/regressed CINⅠ, then five factors with progressed CINⅠwere processed into binary logistic regression analysis. Results (1) In the first year of follow-up, among 104 patients(including 15 cases NILM,78 cases ASCUS,11 cases LSIL), 52 cases of them were NILM and HR HPV negative, 30 cases were negative for intraepithelial lesion, 10 cases were CINⅠ, 5 cases were CINⅡand 7 cases were CINⅢ. In total, 82 cases (78.8%,82/104) cases had regressed, 10 cases (9.6%,10/104) persisted, 12 cases (11.5%,12/104) progressed [including 5 cases (4.8%,5/104) progressed to CIN Ⅱ, 7 cases (6.7%,7/104) progressed to CIN Ⅲ, none progressed to invasive cancer]. (2) All patients, 88 cases of them accepted immunohistochemical detection the expression of p16INK4a protein. The result shown that 30 cases (34%,30/88) were positive and 58 cases (66%,58/88) were negative. And 94 cases accepted immunocytochemical detection the expression of HPV L1 capsid protein, 49 cases (52%,49/94) were positive and 45 cases (48%,45/94) were negative. (3) Univariate analysis showed that age of the patient, loads of HR HPV, cervical cytology before colposcopy and the expression of HPV L1 capsid protein were not risk factors of the progression of CINⅠ(all P>0.05) except for the expression of p16INK4a protein (P<0.05). Multivariable analysis found that p16INK4a protein positive was associated with progression of CINⅠ(OR=5.1,95%CI:1.162-22.387,P=0.031). (4) Thirty-one cases were p16INK4a protein positive, 8 cases (27%,8/30) of them progressed,while 4 cases (7%,4/58) of 58 cases with p16INK4a protein negative progressed,in which there were significant difference (P<0.05). The sensitivity was 75%, the specificity was 71%, the positive predictive value was 27%and the negative predictive value was 93%for progression to CINⅡ-Ⅲof p16INK4a protein staining. Conclusions The progression rate of CINⅠwith abnormal cytology (≤LSIL) and HR HPV positive was lower, and there was no progression to invasion at 1-year intervals. Immunostaining of p16INK4a protein as the risk factors of CINⅠprogression could have a role in prediction of CINⅠand the management of high-risk CINⅠ.

Distribution of Human Papillomavirus 52 and 58 Genotypes, and Their Expression of p16 and p53 in Cervical Neoplasia

BACKGROUND: This study investigates the prevalence of human papillomavirus (HPV) 52 and 58 genotypes among women residing in Busan, and the expression of p16 and p53 proteins in cervical neoplasia with HPV 52 and 58 infections. METHODS: A total of three hundred fifteen cases were analyzed using the HPV DNA chip test for HPV genotypes, and of these, we retrospectively examined p16 and p53 expression in 62 cases of cervical tissues infected with HPV 52 and 58 using immunohistochemistry. RESULTS: HPV 52 and 58 genotypes were identified in 62 (54.9%) out of 113 high-risk, HPV-infected cases. Of the cases examined, there were 19 single HPV 52 infections (16.8%), 23 single HPV 58 infections (20.4%), 4 multiple HPV 52 infections (3.5%), and 16 multiple HPV-58 infections (14.2%). Immunoreactivity of p16 and p53 was observed in 41 (66.1%) and 23 (37.1%) of the 62 cases of cervical neoplasia infected with HPV 52 and 58 genotypes, respectively. CONCLUSIONS: This study demonstrates a high prevalence of HPV 52 and 58 genotypes, in addition to HPV 16, among high-risk strains of cervical neoplasia in Korea. These findings suggest that development of more vaccines would be beneficial for the prevention of the various HPV genotypes.

Human Papillomavirus Prevalence and Cell Cycle Related Protein Expression in Tonsillar Squamous Cell Carcinomas of Korean Patients with Clinicopathologic Analysis

BACKGROUND: Human papillomavirus (HPV)-related tonsillar squamous cell carcinoma (TSCC) has recently been characterized as a distinct subset with a favorable prognosis. The prevalence and clinicopathologic significance of HPV-related TSCC in Koreans are not well known. METHODS: HPV in situ hybridization (ISH) accompanied by p53, p16, pRb, and cyclin D1 immunohistochemical staining were performed on 89 resection cases of TSCC from 2000 through 2010. RESULTS: HPV was detected by ISH in 59 of 89 cases (66.3%). HPV-positive TSCCs were more common in younger ages (p=0.005), and tumor sizes were smaller in the HPV-positive compared to the HPV-negative group (p=0.040). Positive HPV staining was significantly correlated with p16 expression (p<0.001), pRb inactivation (p=0.003), and cyclin D1 down-regulation (p<0.001) but not with p53 expression (p=0.334). Seventeen cases that showed p16-immunopositivity with HPV-negativity by ISH were retested by HPV typing; HPV DNA was not detected in all cases. There was no significant difference between HPV-positive and HPV-negative patients either in the disease-specific survival (DSS, p=0.857) or overall survival (p=0.910). Furthermore, pRb-inactivated cases showed better DSS (p=0.023), and p53-positive cases showed worse DSS (p=0.001). CONCLUSIONS: Although high HPV prevalence was noted, it was not correlated with histopathologic findings or survival benefit. In addition to p53 expression, pRb inactivation along with p16 overexpression and down-regulation of cyclin D1 are thought to be important pathogenetic steps for developing TSCCs.

Ciclo celular, HPV e evolução da neoplasia intraepitelial cervical: seleção de marcadores biológicos / Cell cycle, HPV and cervical intraepithelial neoplasia evolution: biomarkers selection

O câncer de colo do útero é a segunda causa de câncer mais comum entre as mulheres em todo o mundo, sendo precedido por lesões precursoras denominadas neoplasias intraepiteliais cervicais (NICs), de grau 1, 2 e 3. A relação entre o papilomavírus humano (HPV) e o câncer de colo uterino já está bem estabelecida. O ciclo de vida do HPV, assim como seu mecanismo de ação sobre o ciclo celular da célula hospedeira, causam a transformação neoplásica e progressão das lesões precursoras para o câncer de colo uterino. Tal transformação está associada principalmente à expressão de dois genes do HPV: o E6 e o E7, cujos produtos interferem no controle do ciclo celular, tendo como alvos principais as proteínas p53 e pRB. Esta revisão de literatura pretende fornecer ao leitor conhecimento da ação do HPV sobre o ciclo celular, visando identificar potenciais marcadores biológicos da evolução das lesões precursoras ao câncer de colo uterino.

Cervical cancer is the second most common malignant disease in women worldwide and is preceded by cervical intraepithelial neoplasia (CIN) grades 1, 2 and 3. The relationship between human papillomavirus (HPV), cervical CIN and cervical cancer is well established. The HPV life cycle cause neoplastic transformation and CIN progression for cervical cancer. This transformation is associated with E6 and E7 HPV genes, whose products interfere with cell cycle control, mainly through action on p53 and pRB proteins. This review aims to provide to the reader an update for the understanding of the HPV effects on the cell cycle, attempting to identify potential biological markers for the evolution of precursor lesions to cervical cancer.

Immunohistochemical expression of p53, p16 and hTERT in oral squamous cell carcinoma and potentially malignant disorders

Oral carcinogenesis is a multi-step process. One possible step is the development of potentially malignant disorders known as leukoplakia and erytroplakia. The objective of this study was to use immunohistochemistry to analyze the patterns of expression of the cell-cycle regulatory proteins p53 and p16INK4a in potentially malignant disorders (PMD) of the oral mucosa (with varying degrees of dysplasia) and in oral squamous cell carcinomas (OSCC) to correlate them with the expression of telomerase (hTERT). Fifteen PMD and 30 OSCC tissue samples were analyzed. Additionally, 5 cases of oral epithelial hyperplasia (OEH) were added to analyze clinically altered mucosa presenting as histological hyperplasia without dysplasia. p53 positivity was observed in 93.3 percent of PMD, in 63.3 percent of OSCC and in 80 percent of OEH. Although there was no correlation between p53 expression and the grade of dysplasia, all cases with severe dysplasia presented p53 suprabasal immunoexpression. p16INK4a expression was observed in 26.7 percent of PMD, in 43.3 percent of OSCC and in 2 cases of OEH. The p16INK4a expression in OEH, PMD and OSCC was unable to differentiate non-dysplastic from dysplastic oral epithelium. hTERT positivity was observed in all samples of OEH and PMD and in 90 percent of OSCC. The high hTERT immunoexpression in all three lesions indicates that telomerase is present in clinically altered oral mucosa but does not differentiate hyperplastic from dysplastic oral epithelium. In PMD of the oral mucosa, the p53 immunoexpression changes according to the degree of dysplasia by mechanisms independent of p16INK4a and hTERT.

Effect of Helicobacter pylori Infection on p16, p53, CEA, EGFR Expression in Gastric Neoplasia

BACKGROUND/AIMS: Gastric cancer is one of the most widespread cancers and the second leading cause of cancer-related death worldwide. Although Helicobacter pylori (H. pylori) has been classified as a type I carcinogen for gastric cancer, the exact pathway has remained indistinct. In this study, we investigated the effects of H. pylori on oncogenic proteins (epidermal growth factor receptor [EGFR], CEA), tumor suppressor (p53) and cell-cycle regulator (p16) expression, using immunohistochemical stains, in gastric neoplasias. MATERIALS AND METHODS: From April 2007 until July 2009, 166 patients with consecutive gastric neoplasias resected were retrospectively enrolled; 35 gastric dysplasias, 70 early gastric cancers and 60 advanced gastric cancers. We examined the relationship of clinicopathologic features of gastric neoplasias such as age, sex, p16, p53, EGFR, tissue CEA, TNM stage, Lauren classification, location, histologic type of neoplasia to H. pylori infection status. RESULTS: H. pylori infection detected in the samples of gastric dysplasia, early gastric cancer (EGC) and advanced gastric cancer (AGC) were 15 (41.7%), 38 (54.3%) and 33 (55.0%) samples. p53, CEA and EGFR stains expression were associated with cancer stage (P<0.05). There was no relation between the immunohistochemical stains (p16, p53, CEA, EGFR) and H. pylori infection. CONCLUSIONS: This study failed to show any relation of immunohistochemical markers of p16, p53, EGFR, CEA expressions to H. pylori infection in gastric dysplasia as well as gastric cancer. Further study is necessary to investigate the effect of H. pylori infection on p16, p53, CEA, EGFR expressions in precancerous lesions such as atrophic gastritis and intestinal metaplasia.

Estudo de p27, p21, p16 em epitélio escamoso normal, papiloma escamoso e carcinoma de células escamosas da cavidade oral / Comparative analysis of the immunohistochemistry expression of p27, p21WAF/Cip1, and p16INK4a in oral normal epithelium, squamous papilloma and squamous cell carcinoma

INTRODUÇÃO E OBJETIVO: O tipo de câncer oral mais frequente é o carcinoma de células escamosas, que corresponde a 95 por cento dos casos(9). O papiloma escamoso oral é uma neoplasia benigna normalmente associada à infecção pelo papilomavírus humano (HPV)(21). A análise da literatura mostra alterações nos genes reguladores do ciclo celular p27, p21WAF/Cip1 e p16INK4a, porém sem uma definição de seus papéis na carcinogênese oral. O objetivo foi caracterizar imuno-histoquimicamente p27, p21WAF/Cip1 e p16NK4a em epitélio escamoso normal, papilomas escamosos e carcinomas de células escamosas da cavidade oral. MÉTODOS: Imuno-histoquímica para p27, p21WAF/Cip1 e p16NK4a em 32 casos de epitélio escamoso normal, 30 casos de papiloma escamoso e 34 de carcinoma de células escamosas da cavidade oral. RESULTADOS: p27: 97,06 por cento dos casos de carcinoma de células escamosas apresentaram imunopositividade focal. O grupo papiloma escamoso apresentou 33,33 por cento e o grupo controle, 18,75 por cento. p21WAF/Cip1: 100 por cento de imunopositividade focal tanto no grupo controle como no grupo carcinoma de células escamosas, e 90 por cento no grupo papiloma escamoso. p16INK4a: 100 por cento de imunopositividade focal para os grupos controle e papiloma escamoso, e 94 por cento para o grupo carcinoma de células escamosas. CONCLUSÃO: Imuno-histoquimicamente demonstrou-se diferença significativa para p27 quando feita comparação dos grupos controle e papiloma escamoso com o grupo carcinoma de células escamosas. O p21WAF/Cip1 não demonstrou poder de diferenciar os grupos analisados. O p16INK4a apresentou imunopositividade difusa em uma minoria dos casos do grupo carcinoma de células escamosas. O grupo papiloma escamoso se comportou de maneira similar ao grupo controle em relação aos três marcadores.

INTRODUCTION: The most frequent type of oral cancer is the squamous cell carcinoma, which corresponds to 95 percent of the cases(9).The oral squamous papilloma is a benign neoplasia, commonly associated with infections caused by the human papilloma virus(21). The analysis of medical literature shows changes in cell cycle regulatory genes (p27, p21WAF/Cip1 and p16INK4a), but does not define their roles in oral carcinogenesis. Objective: Characterize the immuno-histochemical expression of p27, p21WAF/Cip1 and p16INK4a in oral normal squamous epithelium, oral squamous papilloma and oral squamous cell carcinoma. METHODS: Immuno-histochemical evaluation of p27, p21WAF/Cip1 and p16INK4a in 32 samples of oral normal squamous epithelium, 30 of oral squamous papilloma and 34 of oral squamous cell carcinoma. RESULTS: 97.06 percent of the oral squamous cell carcinoma group, 33.33 percent of the squamous papilloma group and 18.75 percent of the control group showed focal immunopositivity for p27. 100 percent of both control and oral squamous cell carcinoma groups and 90 percent of the oral squamous papilloma group showed focal immunopositivity for p21WAF/Cip1. 100 percent of both control and oral squamous papilloma groups and 94 percent of the oral squamous cell carcinoma group showed focal immunopositivity for p16INK4a. CONCLUSIONS: The study revealed a statistically significant difference for p27 expression when comparing the control and oral squamous papilloma groups with the oral squamous cell carcinoma group. p21WAF/Cip1 did not prove to be useful to differentiate the groups. p16INK4a showed diffuse immunopositivity in a minority of the oral squamous cell carcinoma cases. The oral squamous papilloma group behaved similarly to the control group as to the three markers.

Effects of BMI-1 over-expression on HOX family expression and cell cycle in HeLa cells / 中国病理生理杂志

AIM: To observe whether transfection of mammalian expression vector pEGFP containing the gene of B-cell specific moloney leukemia virus insertion site 1(BMI-1) could express in human cervix cancer cell line HeLa, and to detect its effect on HOX family expression and cell cycle.METHODS: pEGFP-BMI-1 was transfected into HeLa cells with Lipofectamine 2000. The expression of pEGFP-BMI-1 was determined by EGFP fluorescence and Western blotting. SYBR green I real-time RT-PCR was used to quantitate mRNA expression of P16~(INK4a), hTERT, HOXA9, HOXB4 and HOXC13. FACS analysis was used to detect the change of cell cycle.RESULTS: In HeLa cells transfected with pEGFP-BMI-1, the results of real-time RT-PCR showed that the mRNA expressions of P16~(INK4a), HOXA9 and HOXC13 were reduced to 9.2%, 10.9% and 69.7%, respectively, as compared to control HeLa cells (P<0.01). However, hTERT and HOXB4 mRNA expressions did not change significantly (P>0.05). FACS analysis showed a decrease from 65.68 % to 50.53% in G_1 population and a significant increase from 27.17% to 39.59 % in S population after transfection (P<0.01).CONCLUSION: BMI-1 over-expression in HeLa cells down-regulates mRNA expressions of P16~(INK4a), HOXA9 and HOXC13, decreases G_1 population and increases S population. Therefore, BMI-1 may be involved in carcinogenesis and cancer development.

Expression of p16INK4A, CyclinD1, Ki-67 and HPV infection in cervical squamous cell carcinoma / 肿瘤研究与临床

Objective To analyse the expression of p16INK4A, CyclinD1, Ki-67 proteins and HPV16/18, 31/33 in invasive squamous cell carcinoma of cervix(ISCC)and to indicate the significance of them in the occurrence and development of ISCC. Methods Using tissue microarray,in Situ Hybridization (ISH) and immunohistochemical method detected the expression of HPVI6/18, 31/33 and investigated the expression of p16INK4A, CyclinD1, Ki-67 in ISCC, CIN and NCE. Results was analysed by SPSS vision 13.0. Results The positive expression rate of HPV16/18, CyelinD1, Ki-67 in ISCC was markedly higher than that in CIN and NCE. Significant relationship was observed between p16INK4A protein expression and histological grade of the cancer. No significant correlation was observed between CyclinD1 protein expression and every clinical pathological parameters of ISCC. Conclusion High-risk HPV takes an important role in the development in ISCC. p16INK4A gene might not act as a suppressor-gene only on the occurrence in ISCC. Ki-67 can be looked as a valuable clinic index of diagnosing benign and malignant tumor.

The Overexpression of Histone Deacetylase 1 and Its Relationship with p16INK4a Gene Hypermethylation in Pulmonary Squamous Cell Carcinoma and Adenocarcinoma

BACKGROUND: DNA methylation and histone modification are dynamically linked in the epigenetic control of gene silencing and they play an important role in tumorigenesis. METHODS: To evaluate the role of histone deacetylase 1 (HDAC1) in the development of lung cancer and the relationship between a HDAC1 overexpression and p16INK4a hypermethylation, we performed immunohistochemical staining for HDAC1 in 76 lung cancer specimens (39 squamous cell carcinomas and 37 adenocarcinomas) that had been previously evaluated for their p16INK4a methylation status by real-time quantitative polymerase chain reaction. RESULTS: A HDAC1 overexpression (>50% of HDAC1 immunoreactive cells) was detected in 65 (85.5%) out of the 76 cases and it was more frequently seen in the squamous cell carcinomas (97.4%) than in the adenocarcinomas (73.0%) (p=0.002). The incidence of HDAC1 overexpression tended to be higher in the heavy smokers with more than 20 pack-years (p=0.067). Although there was no statistical significance, the frequency of p16INK4a hypermethylation in the cases with a HDAC1 overexpression (27.7%) tended to be higher than that in the cases without a HDAC1 overexpression (9.0%) (p=0.175). CONCLUSIONS: A HDAC1 overexpression might be involved in lung carcinogenesis, and especially in a subgroup of smoking and squamous cell carcinoma patients, and a HDAC1 overexpression may be associated with p16INK4a hypermethylation.

Expressão de p53, p16 E COX-2 em carcinoma escamoso de esôfago e associação histopatológica / p53, p16 E COX-2 expression in esophageal squamous cell carcinoma and histopathological association

RACIONAL: O câncer de esôfago representa cerca de 2 por cento dos tumores malignos e a terceira causa mais comum de câncer do trato gastrointestinal. A associação do prognóstico do câncer de esôfago com alguns marcadores imunoistoquímicos, como as proteínas p53, p16 e a ciclooxigenase 2 (COX-2) tem sido relatada. A detecção de marcadores moleculares através de imunoistoquímica pode ser utilizada para avaliação prognóstica. OBJETIVOS: Investigar a associação entre a expressão das proteínas p53, p16 e a COX-2 com o estádio do carcinoma escamoso de esôfago. MÉTODOS: Foram analisadas 31 amostras de ressecção cirúrgica por esofagectomia diagnosticadas como carcinoma de células escamosas de esôfago e 31 amostras não-tumorais referentes a cada caso. Realizou-se a revisão histopatológica e o estádio pTNM. Amostras tumorais e não-tumorais adjacentes foram submetidas a análise imunoistoquímica para avaliar o conteúdo das proteínas p53, p16 e COX-2. Foi considerada positiva a expressão nuclear para p53 em quantidade igual ou superior a 10,00 por cento das células e presença da expressão citoplasmática de acordo com três escores (1, 2, 3) de intensidade (leve, moderada, acentuada) de imunocoloração para COX-2. RESULTADOS: Em área tumoral, as análises revelaram 48,38 por cento de positividade para p53, 16,12 por cento de positividade para p16, e 100,00 por cento de positividade escores 1+, 2+ ou 3+ para COX-2. No entanto, quando se avaliou possível relação da expressão destes marcadores com o estádio, apenas a COX-2, escore 3+ intensidade acentuada mostraram associação significativa. CONCLUSÃO: O presente estudo demonstrou que existe relação positiva entre a expressão de COX-2, escore 3+ e estádio mais avançado no carcinoma de esôfago.

BACKGROUND: The esophageal carcinoma represents about 2 percent of malignant tumors and is the third most common cause of gastrointestinal cancer. The correlation between immunohistochemistry markers, such as p53, p16 and COX-2 proteins and cancer esophageal prognosis has been suggested. AIMS: To Investigate whether the expression of p53, p16 and COX-2 proteins are associated to tumor staging. METHODS: For this purpose we proceeded immunohistochemistry assays and TMN in 31 esophageal tumor and normal tissue samples. The p53 nuclear expression was considered positive when it appears in 10.00 percent or more cells. COX-2 expression was scored according to intensity in three scores (1+, 2+, 3+). RESULTS: On the tumor samples the results presented 48.38 percent positivity for p53, 16.12 percent for p16 and 100 percent with 1+, 2+ or 3+ scores for COX-2. However, when we investigated whether the expression of p53, p16 and COX-2 proteins are related to tumor staging, only COX-2 expression, score 3+, had shown statistical significant association. CONCLUSION: Therefore, in the present study we could see positive correlation between COX-2 protein and high grade tumor as well as advanced tumor staging in esophageal carcinoma.

Cyclin D1 Overexpression, p16 Loss, and pRb Inactivation Play a Key Role in Pulmonary Carcinogenesis and have a Prognostic Implication for the Long-term Survival in Non-small Cell Lung Carcinoma Patients / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: We investigated the immunoexpressions of cyclin D1, cyclin-dependent kinase inhibitor p16 and phosphorylated retinoblastoma (p-pRb) proteins in non- small cell lung carcinoma (NSCLC) to demonstrate their key roles in tumorigenesis, their relationship with the clinicopathologic factors, and their prognostic influences on the long-term survival. MATERIALS AND METHODS: 115 surgically resected NSCLCs were immunohistochemically stained for the G1/S cell cycle proteins, with using a tissue microarray. The correlation between their immunoexpressions and the clinicopathologic prognostic factors, their inter-relationships and their single or combined effects on the long-term survival (over 5 years) were statistically analyzed by SPSS15.0. RESULTS: Loss of p16 was found in 75% of the cases and cyclin D1 overexpression and phosphorylated pRb (p-pRb) were found in 64% and 46%, respectively. Cyclin D1 overexpression was correlated with the p16 loss and pRb inactivation by phosphorylation. The p16 loss was tightly associated with p-pRb. The Kaplan-Meier survival curves disclosed that the cyclin D1-positive group and the p16-negative group showed a rapid decline of survival at the point of about 5 years after surgery and thereafter. The combined actions of cyclin D1 overexpression, loss of p16 and pRb inactivation tended to have an adverse influence on the prolonged survival. CONCLUSIONS: The observation that cyclin D1 overexpression, p16 loss and pRb inactivation were largely found in NSCLCs suggests that they play an important role in pulmonary carcinogenesis. Also, their inverse or positive correlations indicate that the G1/S cell cycle proteins may act alternatively or synergistically on the mechanisms by which tumor cells escape the G1 restriction point. Finally, their solitary or combined actions might have a long-term effect on the survival.