Expression of E6/E7 mRNA from 'high risk' human papillomavirus in relation to CIN grade, viral load and p16INK4a

Detection of E6/E7 mRNA expression with real-time nucleic acid sequence-based amplification assay (NASBA) method (PreTect HPV-Proofer) from high-risk types of human papillomaviruses (HR-HPV) were compared with the presence of viral load, determined with quantitative real-time PCR in 80 cervical samples. Results regarding positivity and typing were in agreement using the two methods. However, there was no correlation between viral loads for HPV 16 or 18/45 and oncogene expression. Among 15 women with low grade atypia detected at a population-based cytology screening, and scored as 'within normal limits' according to histopathology, 14% were positive for oncogene expression, whereas 71% were HR-HPV positive. A correlation was observed between HR-HPV oncogene expression and high scores of p16(INK4a) positivity. Since HPV-Proofer detects full-length E6/E7 mRNA, a positive result should correlate with presence of integrated HPV, loss of HPV replication and stabilized E6/E7 full-length mRNA expression. Such expression from integrated HR-HPV generates a high and stable expression of full-length E6 proteins, which explains why a positive HPV-Proofer result was independent of viral load and correlate with high expression of p16(INK4a). Thus, E6/E7 oncogene expression analysis yielded information, which is consistent with and will complement the results from a real-time PCR method in a clinical prognostic procedure.     

Triage options to manage high-risk human papillomavirus-positive women: A population-based cross-sectional study from rural China

Improvement in managing HPV-positive women is urgently needed. Based on a population-based study which included 2112 women aged 49 to 69 from Shanxi, China, we aimed to evaluate the clinical performance of multiple triage strategies based on liquid-based cytology (LBC), p16^INK4a, viral load and partial genotyping, as a single or combined strategy for detecting cervical intraepithelial neoplasia grade 2/3 or higher (CIN2+/CIN3+) in women who tested positive by Hybrid Capture 2 (HC2). Among 452 HC2-positive women, the test positivity of LBC (ASC-US+), p16^INK4a, HPV16/18 and HPV16/18/31/33/45 were 39.6%, 38.5%, 18.0% and 40.0%, respectively. Compared to LBC (ASC-US+) triage, a single triage strategies using p16^INK4a or extended genotyping (SureX HPV16/18/31/33/45) achieved comparable sensitivity (relative sensitivity: 1.08, 95% confidence interval [CI]: 0.93-1.26 and 0.96, 95% CI: 0.76-1.22) and specificity (relative specificity: 1.05, 95% CI: 0.96-1.14 and 1.02, 95% CI: 0.92-1.14) for CIN3+. Viral load triage using a ≥50 RLU/CO cut-point also yielded similar results with LBC (ASC-US+). Among combined triage strategies, HPV16/18 genotyping with reflex p16^INK4a showed higher sensitivity and slightly lower specificity than LBC (ASC-US+) for CIN3+ detection, however, the differences were not statistically significant. Of note, after a negative result by p16^INK4a or LBC among HPV16/18 negative women, the posttest probability of CIN3+ was lower than 1%. Our study suggested that p16^INK4a, extended genotyping and increased viral load cut-point could be promising alternatives to cytology triage. Combined triage algorithms of HPV16/18 with reflex p16^INK4a or cytology, if negative, are associated with the substantial low posttest risk sufficient to release women to next screening round.     

Combined p16INK4a and human papillomavirus testing improves the prediction of cervical intraepithelial neoplasia (CIN II-III) in Thai patients with low-grade cytological abnormalities

Thailand is in the process of developing a national cervical screening program. This study examined p16INK4a staining and HPV prevalence in abnormal cervical samples with atypical squamous cells of undetermined significance (ASCUS) and low-grade squamous intraepithelial lesion (LSIL), to evaluate the efficacy of combined HPV and p16INK4a detection to predict CIN II-III. Totals of 125 ASCUS and 87 LSIL cases were re-evaluated by Pap test and cervical cells of ASCUS and LSIL cases were prepared on slides for p16INK4a detection by immunocytochemistry. HPV genotyping of DNA extracts was performed by GP5+/6+ PCR and reverse line blot hybridization. Histopathologic tests were performed to identify cervical lesion. Total of 212 cases were diagnosed to normal (20), ASCUS (112), LSIL (78) and HSIL (2). HPV was detected in ASCUS (49/112, 43.8%), LSIL (60/78, 76.9%) and HSIL (2/2, 100%) cases. The majority of HPV positive samples typed for high-risk HPV. 55.7% (107/192) of abnormal cases (ASCUS, LSIL and HSIL) were positive p16INK4a. For the 111 HPV DNA positive cases, 34 of 49 (69.4%) ASCUS cases and 49 of 60 (81.7%) LSIL cases were p16INK4a positive. 140 biopsies were taken and histological classified: CIN negative (65 cases), CIN I (56 cases) and CIN II-III (19 cases). HPV DNA detection predicted CIN II-III with sensitivity and specificity of 84% and 49%, whereas p16INK4a staining showed higher sensitivity (89.5%) and specificity (56.2%). The prediction of CIN II-III was significantly better by combination of positive HPV DNA and p16INK4a with 93.8% sensitivity and 59.2% specificity. Detection of HPV DNA combined with p16INK4a in cervical cells can predict CIN II-III and may improve the screening diagnosis of Thai women at risk for CIN II-III or cancer.     

高危型HPV负荷量和p16INK4A蛋白监测评估宫颈锥切术治疗CIN

[目的]评价高危型人乳头状瘤病毒HPV负荷量的检测和p16INK4A蛋白的表达在预测宫颈上皮内瘤变(CIN)宫颈锥切术后残存病变或复发中的意义.[方法]回顾性分析142例2008年10月至2010年12月因CIN行宫颈锥形切除术治疗患者的临床资料.所有患者均于宫颈锥形切除术前6个月以内和术后6～12个月进行HPV负荷量检测,并采用免疫组化方法检测HPV DNA阳性患者宫颈细胞中p16INK4A蛋白表达.[结果]宫颈锥切术前,随着CIN级别的上升,HPV负荷量以及p16INK4A蛋白表达均明显增强(P＜0.05).但在宫颈锥切术后,HPV负荷量和p16INK4A蛋白表达明显降低,宫颈锥切术前和术后两者之间差异有统计学意义(P＜0.05).[结论] HPV负荷量持续增高和p16INK4A蛋白持续呈强阳性是宫颈锥切术后发生残存病变或复发的高危因素,在监测HPV负荷量的同时检测p16INK4A蛋白的表达,对判断宫颈锥切术后发生残存病变或复发有重要意义.     

Validation of p16INK4a as a marker of oncogenic human papillomavirus infection in cervical biopsies from a population-based cohort in Costa Rica.

Due to the high prevalence of cancer-associated types of human papillomavirus (HPV) and the poorly reproducible histologic classification of low-grade lesions, identifying infected women at highest risk for cancer prior to neoplastic progression remains a challenge. We therefore explored the utility of p16INK4a immunostaining as a potential diagnostic and prognostic biomarker for cervical neoplasia using paraffin-embedded tissue blocks (punch biopsies and loop electrosurgical excision procedures) obtained from women referred to colposcopy during the enrollment phase of the Guanacaste Project (1993 to 1994). All blocks from 292 women selected by HPV status (HPV negative, nononcogenic HPV positive, or oncogenic HPV positive) and representing the diagnostic spectrum of the population [normal to precancer: cervical intraepithelial neoplasia (CIN) 3] were immunostained for p16INK4a using the p16INK4a research kit based on the monoclonal antibody clone E6H4 (MTM Laboratories, Heidelberg, Germany). For CIN3, the sensitivity of diffuse p16INK4a immunostaining was 100% and the specificity was 95%. For CIN2, the sensitivity and specificity for diffuse staining were 81.1% and 95.4%, respectively. Generalized to the 10,000-woman cohort, this translated to positive predictive value and negative predictive value of 13.9% and 100% for CIN3, respectively, and 20.4% and 99.7% for CIN2 or CIN3, respectively. Of women with an initial diagnosis of less than CIN2 for whom follow-up data for up to 5 to 7 years were available, 44% with diffuse staining developed persistent infection (CIN2 or CIN3). Whereas our data support the diagnostic potential for p16INK4a, further prospective studies with detailed follow-up determining the prognostic capacity of this marker are needed.     

Determination of viral load thresholds in cervical scrapings to rule out CIN 3 in HPV 16, 18, 31 and 33-positive women with normal cytology

An increased high-risk human papillomavirus (hrHPV) viral load in cervical scrapings has been proposed as a determinant for high-grade cervical intraepithelial neoplasia (CIN) and cervical cancer (> or =CIN 2), but data so far for HPV types different from HPV 16 are limited and inconsistent. In addition, a viral load threshold to distinguish hrHPV positive women without > or =CIN 2 still has not been defined. Here, we used baseline cervical scrapings of women with normal cytology participating in a large population-based cervical screening trial (i.e. POBASCAM) who were GP5+/6+-PCR positive for 4 common hrHPV types, i.e. HPV 16, 18, 31 or 33, as a reference to arbitrarily define various viral load thresholds (i.e. 25th, 33rd, 50th, 67th and 75th percentiles of the lowest viral load values) for distinguishing women having single infections with these types without high-grade CIN. Viral load assessment was performed by real time type-specific PCR. The viral load threshold values were subsequently validated on abnormal cervical scrapes of 162 women with underlying, histologically confirmed CIN lesions containing 1 of these 4 hrHPV types. All 59 women with CIN 3 had viral load levels that were higher than those of 33% of the women with normal cytology containing the respective hrHPV type detectable by GP5+/6+-PCR (i.e. higher than the 33rd percentile of viral load). By using this 33rd percentile viral load cut-off, sensitivity for CIN 3 of 100% (95% CI 93.9-100) was obtained. Hence, application of this viral load threshold would increase the specificity of HPV testing for HPV 16, 18, 31 and 33-associated prevalent CIN 3 without the cost of a marked reduction in sensitivity. In practice, on the basis of viral load analysis, a less aggressive management can be foreseen for 33% of the women with normal cytology participating in a population-based screening program who are GP5+/6+-PCR positive for HPV 16, 18, 31 or 33.     

Physical state and viral load as predictive biomarkersfor persistence and progression of HPV16-positive cervical lesions: results from a population based long-term prospective cohort study

Persistent infection with a high risk (hr) human papillomavirus (HPV) has been established as the main cause of cervical cancer and high-grade cervical intraepithelial neoplasia (CIN3). Because most infections are transient, testing for hrHPV lacks specificity and has a low positive predictive value. It has been suggested that additional parameters like viral load and physical status of the viral genome could improve the effectiveness of HPV-based screening. We investigated the association between HPV16 viral load and physical state with viral persistence or risk of incident CIN3 or worse in a population-based prospective cohort study comprising 8656 women (20-29 years). All participants had two gynecological examinations two years apart and were followed through the nationwide Danish Pathology Data Bank (median follow-up: 12.9 yrs). Seventynine cervical swabs from women with a persistent HPV16 infection were available for analysis. For comparison we selected a random age-matched sample of transiently HPV16 infected women (N=91). Persistently infected women with incident CIN3 or cancer (CIN3+; N=31) were compared to women with normal cytology during follow up (non-progressors; N=39). Quantitative real-time PCR for HPV16E6, E2 and IFNb1 was done to determine the HPV16 viral load and the E2/E6 ratio was used as a surrogate marker for integration. Women with normal cytology who became persistently HPV16 infected had a significantly lower HPV16 load at baseline than women who cleared the infection (median 4.72 copies/cell versus median 20.0 copies/cell, respectively; p=0.0003). There was no difference in viral load at enrollment between women who progressed to CIN3+ and women who stayed cytologically normal (p=0.85). At the second examination viral load tended to be higher in women who progressed, but the difference was not statistically significant (p=0.39). The E2/E6 ratio was shown to be lower in the persistently infected group (p<0.0001) already at the first examination, but no difference between non-progressors and CIN3+ cases was observed at any of the two examinations (p=0.61 and 0.86). Lower viral load and integration of the viral genome are predictive for the persistence of HPV16 DNA, but not for the progression of a persistent HPV16 infection to CIN3+ in women with normal cytology.     

Role of P16(INK4a) expression in identifying CIN2 or more severe lesions among HPV-positive patients referred for colposcopy after abnormal cytology

BACKGROUND: p16 is strongly overexpressed in dysplastic cervical cells because of the transforming activity of the E7 oncogene of all high-risk human papillomavirus (HR-HPV) types and may be easily revealed by immunochemistry: p16 may, therefore, be considered a surrogate marker for the activated oncogene expression of HR-HPV in dysplastic cervical cells. METHODS: HPV and p16(INK4a) testing were performed in a consecutive series of 283 patients with abnormal cytology referred to colposcopy assessment or follow-up. Triage of patients to colposcopy by HPV or HPV and p16 testing was simulated, and the relative sensitivity, specificity, and positive predictive value (PPV) of HPV and p16 testing for > CIN2 lesions was determined as well as the cost balance of the two triage types. RESULTS: Compared to current protocol, triage by HPV testing reduced the number of colposcopies by 44.2%, but also reduced the > CIN2 detection rate by 10.7%, and was associated with a cost of euro 54.16 per assessed woman and of euro 613.20 per > CIN2 detected. Compared with current protocol, triage by HPV and p16 testing combined reduced the number of colposcopies by 73.1%, but reduced > CIN2 detection rate by 21.5%, and was associated with a cost of euro 54.73 per woman assessed and of euro 704.09 per > CIN2 detected. CONCLUSIONS: Triage by HPV and p16 improves considerably the PPV of diagnostic assessment, but decreases > CIN2 detection rate, and is associated with substantially higher costs. Further decrease of molecular immunochemistry testing due to technological progress may allow HPV and p16 testing to become a cost effective procedure in the future.     

p16INK4A immunohistochemical staining and predictive value for progression of cervical intraepithelial neoplasia grade 1: A prospective study in China

p16^INK4A is strongly expressed in tissues diagnosed as cervical intraepithelial neoplasia (CIN) and cancer in women infected with human papillomavirus (HPV), but few prospective studies have evaluated p16^INK4A as a marker for the risk of low‐grade CIN (CIN1) progression. We investigated the prevalence of p16^INK4A immunostaining by CIN grade and whether overexpression of p16^INK4A in CIN1 predicts future risk for high‐grade CIN in Chinese women. 6,557 Chinese women aged 30–49 years were screened from 2003 to 2005 using cytology and carcinogenic HPV test. Colposcopy was performed on women with any abnormal result. p16^INK4A Immunostaining was performed on biopsies from all women with CIN1, as well as randomly selected women with normal or CIN grade 2 and worse (CIN2+) biopsies. Women with CIN1 were followed up without treatment. Colposcopy was performed on all untreated women at a 2‐year interval. The prevalence of p16^INK4A staining was 2.7%, 42.7%, 75.5%, 79.6% and 100% among women with normal, CIN1, 2, 3 and cancer biopsies, respectively (p < 0.001). HPV positivity was strongly associated with p16^INK4A staining [odds ratios (OR) = 12.8; 95% confidence intervals (CI): 5.2–31.6]. p16^INK4A staining of CIN1 biopsies at baseline was associated with an increased risk of finding high‐grade CIN over 2 years of follow‐up (OR = 1.43; 95% CI: 0.52–3.91). The two‐year cumulative incidence of CIN2+ for p16^INK4A positive women was higher at 10.71% than for p16^INK4A negative women at 1.30% (crude RR = 8.25, 95% CI: 1.02–66.62). p16^INK4A overexpression is strongly associated with grade of CIN and risk of progression to high‐grade CIN in women with low‐grade lesions.     

p16INK4a免疫细胞化学检测在筛查宫颈癌中的作用

目的 探讨p16^INK4a免疫细胞化学检测在筛查宫颈癌及其癌前病变中的作用。方法 选择220例宫颈液基细胞学剩余标本,制作液基薄片进行p16^INK4a 免疫细胞化学检测,随访组织活检结果,并与高危人乳头瘤病毒（HR—HPV）DNA检测结果进行对照。结果 p16^INK4a在宫颈细胞学诊断的鳞状细胞癌（SCC）、鳞状上皮内高度病变（HSIL）、鳞状上皮内低度病变（LSIL）、非典型鳞状细胞-小除外上皮内高度病变（ASC—H）和非典型鳞状细胞-不能明确意义（ASC—US）病例的阳性表达率分别为100．0％（7／7）、92．2％（107／116）、24．3％（17／70）、100．0％（14／14）和36．4％（4／11）。150例p16^INK4a阳性者中,111例有组织活检诊断,其中宫颈上皮内瘤变（CIN）2级及以上病变者97例（87．4％）;70例p16^INK4a阴性者中,18例有组织活检诊断,无一例CIN2及以上病变。p16^INK4a在CIN2及以上病变与在CIN1之间的阳性表达率差异有统计学意义（P〈0.01）,而HR-HPV DNA的阳性率在两者之间差异无统计学意义。结论 p16^INK4a在宫颈HSIL及以上病变中高表达,有利于高危病例的筛选。     

Human papillomavirus 16 load in normal and abnormal cervical scrapes: an indicator of CIN II/III and viral clearance

The relation between human papillomavirus type 16 (HPV 16) viral load in cervical scrapes and development of high-grade cervical intraepithelial neoplasia (CIN II or III) was studied in a nested case-control study of women with normal cytology (group A) and in a cohort of women with abnormal cytology (group B). HPV 16 DNA load was determined using a quantitative real-time PCR assay. In group A, case women (women with CIN II/III, n = 12) had a significantly higher viral load than control women (women with CIN < or = I, n = 47). This resulted in an increased relative risk of women with the 50% highest viral load for development of CIN II/III (OR 7.7; CI 1.6-33). In group B, women with CIN II/III (n = 38) had a significantly higher viral load than women with CIN < or = I (n = 25). Women with the 50% highest viral load had an increased relative risk of CIN II/III (OR 3.2; CI 1.1-9.3) and a decreased chance of both viral clearance and cytologic regression. Our data suggest that in women with normal cytology an increased HPV 16 load confers an increased risk of developing a CIN lesion. A sustained high viral load is subsequently informative for progression to a high-grade CIN lesion.     

Cervical Pathology in Cytology-Negative/HPV-Positive Women: Results from Lampang Cancer Hospital,Thailand

Background: To evaluate the cervical pathology of cytology-negative/high-risk human papillomavirus(HR-HPV) positive-women. Materials and Methods: This study recruited 4,583 women aged 30-70 years whohad undergone cervical screening by liquid-based cytology and HR-HPV test (14 HR-HPV types) at LampangCancer Hospital during October 2012 to July 2013. Colposcopy was carried out in all women. Results: Onehundred and ninety-two (4.19%) women were found to be cytology-negative/HR-HPV-positive. However, 23cases were excluded because of incomplete information, leaving 169 women for further analyses. Of these 169,45 (26.6%) were infected with HPV 16/18 and 49 (29.0%) with multiple genotypes of HR-HPV. Nineteen of 169(11.24%) women were found to have CIN 2-3. No women in the present study had AIS or invasive cervical lesions.Prevalence of CIN 2-3 among women infected with HPV 16/18 was 15.6% which was higher than the 9.68% inthose with non-HPV 16/18 oncogenic types. Conclusions: Overall, 11% of cytology-negative/HR-HPV-positivewomen had significant cervical lesions. Risk of harboring such lesions was substantially increased among thosewho were HPV 16/18 positive.     

Type distribution, viral load and integration status of high-risk human papillomaviruses in pre-stages of cervical cancer (CIN)

A series of 176 archival cervical intraepithelial neoplasia (CIN) was analysed for the presence, viral load and integration status of 'high-risk' types of human papillomavirus (HR-HPV). The samples were assayed using newly developed methods based on real-time PCR. Two methods for the extraction of DNA from the paraffin-embedded biopsies were compared: a protocol based on the MagNA pure system (Roche) and a Qiagen spin column kit (Qiagen). It was possible to amplify 94% (166) of the samples. Of these, 36, 63 and 80% of the CIN I, II and III cases contained HR-HPV. HPV 16 was the most prevalent, and was found in 20, 28 and 46% of the CIN I, II and III cases, respectively. The second most frequent HR-HPV was type 33 group, and in CIN II it was as prevalent as HPV 16. The median number of copies of HR-HPV per cell was not significantly different in the CIN I, II and III cases, but there was a wide range of viral load values over several magnitudes, regardless of the grade of CIN. All samples were found to contain integrated forms of HPV 16, frequently mixed with an episomal form.     

Persistence and load of high-risk HPV are predictors for development of high-grade cervical lesions: a longitudinal French cohort study

Oncogenic HPV types are the major cause of worldwide cervical cancer, but only a small proportion of infected women will develop high-grade cervical intraepithelial neoplasia or cancer (CIN2/3+). We performed a prospective study including 781 women with normal, atypical squamous cells of undetermined significance (ASCUS) or low-grade squamous intraepithelial lesion (LGSIL) cytology, and infected or not by high-risk (HR) HPV tested by Hybrid Capture II. Women were followed up every 6 months for a median period of 22 months. Among the HR-HPV-positive women at entry, more than half cleared their virus in 7.5 months; the clearance rate was greater for low viral loads than for high loads and also was higher in women with an initial ASCUS/LGSIL smear than in women with normal cytology. The incidence of cytologic abnormalities strongly depended on baseline viral load and HR-HPV persistence. Maintenance of cytologic abnormalities was associated with the outcome of HR-HPV status (negative or =100 pg/mL). Conversely, women who were consistently HR-HPV negative or transiently HR-HPV positive, whatever the cytology at baseline was, did not develop CIN2/3+ during follow-up. Age seemed to affect only the rate of incident HR-HPV infection. In conclusion, our data suggest that women repeatedly tested positive for HR-HPV are at risk of developing CIN2/3+, even when initial cytology is normal. A high viral load could be used as a short-term marker of progression toward precancerous lesions, although lower load does not inevitably exclude progressive disease.     

Evaluation of a new p16(INK4A) ELISA test and a high-risk HPV DNA test for cervical cancer screening: results from proof-of-concept study

p16(INK4a), a cell cycle regulation protein, accumulates in abnormal epithelial cells infected with high-risk human papilloma virus (HPV). In immunostaining studies, p16(INK4a) has shown potential as a marker of high grade cervical intraepithelial neoplasia (CIN) and invasive cervical cancer. To evaluate its potential use in cervical cancer screening, we conducted a feasibility study to compare the performance of a new enzyme linked immunosorbant assay (ELISA) for p16(INK4a) (mtm laboratories, Heidelberg, Germany) to that of the Hybrid Capture 2 (hc2) test for high-risk HPV DNA for the detection of CIN3. Three hundred and nineteen women were referred from Western Washington Planned Parenthood clinics for colposcopy examination and cervical biopsy because of abnormal Pap test results. Cervical samples were obtained from study participants for p16(INK4a) ELISA, liquid-based cytology and hc2. The order (first and second) for obtaining samples for cervical cytology and p16(INK4a) ELISA changed with every other subject. Concentrations of p16(INK4a) protein were higher when the sample was taken before the cytology. The sensitivity of p16(INK4a) ELISA (concentration > or = 8 units/ml) taken as first sample was 90.0% for CIN3, and the sensitivity of HC2 taken as a second sample was 85%. In the same group, the specificity of p16(INK4a) ELISA (46.9%) was slightly better than hc2 (35.4%) Results from this proof-of-concept study suggest that p16(INK4a) ELISA has a similar sensitivity and slightly better specificity for CIN3 compared to hc2. These findings support proceeding with a larger study with samples from a population of women presenting for routine cytology screening.     

p16INK4a在宫颈液基细胞学辅助筛查中的标记意义

目的 评价p16INK4a在宫颈液基细胞学检查中的标记意义.方法 收集74例宫颈外口和颈管的脱落细胞标本,分别进行液基细胞学检测和p16INK4a免疫细胞化学染色,并应用杂交捕获二代法检测高危人乳头瘤病毒(HR-HPV)感染.结果 74例标本中,细胞学诊断未见癌细胞或癌前病变细胞(阴性)10例,意义不明的不典型鳞状细胞(ASC-US)15例,鳞状上皮内低度病变(LSIL)28例,不除外上皮内高度病变的不典型鳞状细胞(ASC-H)5例,鳞状上皮内高度病变(HSIL)11例,鳞状细胞癌(SCC)5例.各级别病变中,HR-HPV阳性者分别为1、4、3、9、7和5例,p16INK4a免疫细胞化学染色阳性者分别为2、5、3、8、9和5例.随着宫颈病变级别的上升,HR-HPV和p16INK4a免疫细胞化学染色阳性率均增高.结论 p16INK4a免疫细胞化学染色增强了对不典型细胞的区分能力,可以提高宫颈癌筛查的准确性.     

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.