Comparison Between In Situ Hybridization and Real-time PCR Technique as a Means of Detecting the Integrated Form of Human Papillomavirus 16 in Cervical Neoplasia

Fujii, Takuma; Masumoto, Nobuo; Saito, Masafumi; Hirao, Nobumaru; Niimi, Shinichi; Mukai, Makio; Ono, Akiko; Hayashi, Shigenori; Kubushiro, Kaneyuki; Sakai, Eiichi; Tsukazaki, Katsumi; Nozawa, Shiro

doi:10.1097/01.pas.0000162755.84026.9f

Cited by 40 publications

(46 citation statements)

References 27 publications

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“…23,24 All assays were performed using the Chromo 4 DNA Engine cycler (MJ Research, Bio-Rad, CA). Briefly, 20 ll of PCR amplification was done in duplicate contain- ing 100 ng of extracted genomic DNA, 10 ll of 23 TaqMan Universal PCR Master Mix (Applied Biosystems, Foster, CA) and 1 ll of 203 primer/probe.…”

Section: Hpv Viral Load By Real-time Quantitative Pcrmentioning

confidence: 99%

Host and viral factors in relation to clearance of human papillomavirus infection: A cohort study in Taiwan

Lai

Chao

Chang

et al. 2008

Intl Journal of Cancer

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Human papillomavirus (HPV) persistence is essential for cervical cancer development. We accrued nested-cohort subjects from a population-based study to investigate the host and viral factors related to outcome of HPV infection. Women (age ≥ 30 years old) with HPV-positive but normal cytology and negative colposcopy were invited to participate. After signing informed consent, every participant completed a structured questionnaire and had 6-monthly follow-ups of Pap smear, HPV testing and colposcopy. Total and type-specific HPV clearance rates as well as host and viral factors associated with clearance in 3-year longitudinal followup were analyzed. Adjusted hazard ratios (HRs) of progression to ≥ cervical intraepithelial neoplasia (CIN) 2 according to baseline HPV of the women with normal cytology were calculated from national registry database. Among the 626 eligible women, 526 (median age 47, 29-75) were enrolled and 412 returned for followup at least once. The median follow-up of enrolled subjects was 23 months (range 6.8-39). The 3-year cumulative total HPV clearance rate was 49.0% (95% confidence interval [CI]: 43.3-54.7%). The median 3-year cumulative type-specific HPV clearance rate was 50.0% (range 0-100.0%) with a median time to clearance of 12.4 months (6.4-24.5). Older age was associated with significantly decreased total HPV clearance and decreased type-specific clearance in HPV-18 and -53, while high viral load was associated with decreased total and type-specific clearance. After adjusting confounding variables, the HR of developing CIN2 in baseline HPVpositive women was 34.0-fold (95% CI: 15.5-74.7) as compared to HPV-negative women. ' 2008 Wiley-Liss, Inc.Key words: human papillomavirus; genotype; viral load; clearance Cervical cancer is one of the leading cancers in female worldwide. 1 There is strong epidemiological and molecular biological evidence indicating that human papillomavirus (HPV) plays a central role in the etiology of cervical cancer. 2,3 Over 90% of cervical cancers and their precursor lesions are attributable to HPV infection. 4,5 However, the prevalence of genital-tract HPV infections has been reported to range from 1.4 to 25.6% in cytologically normal population. 6 It is widely believed that persistent HPV infection is necessary for the development of cervical cancer. 3,7,8 Incorporation of HPV testing into cervical cancer screening has been advocated, although specificity is decreased with expected increase in sensitivity in early detection of high-grade cervical neoplasms. 9-11 However, identifying infection of high-risk HPV causes anxiety in the individual woman. Because of lack of adequate knowledge in predicting outcome, the appropriate management is unclear except periodical follow-up.Several 8,13 HPV variants 17 have been reported associated with HPV persistence/decreased clearance, whereas ever users of oral contraceptives were associated with faster clearance. 15 Multiple infections, viral load 15 and smoking 8 were unrelated to clearance in other longitudinal follow-u...

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Section: Hpv Viral Load By Real-time Quantitative Pcrmentioning

confidence: 99%

Host and viral factors in relation to clearance of human papillomavirus infection: A cohort study in Taiwan

Lai

Chao

Chang

et al. 2008

Intl Journal of Cancer

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“…ISH-TSA detected integrated form of HPV16/18 with sensitivity of 70.0% and specificity of 60.9%, as compared with RT-PCR. Fujii et al [22] for 53 women with cervical neoplasia or cervical cancer, compared frequency of HPV16 integrated form in exfoliated cervical cells between RT-PCR and ISH-TSA. They found a concordance rate of 86.0%.…”

Section: Discussionmentioning

confidence: 99%

“…However, they did not compare the sensitivity of RT-PCR and ISH-TSA in the assessment of viral genome state. In turn, Fujii et al [22] have shown in cervical neoplasia a strong concordance between RT-PCR and ISH-TSA according to the frequency of HPV16 integration. Therefore, large scale clinical studies are still required to validate the ISH-TSA method [7].…”

Section: Introductionmentioning

confidence: 99%

“…Values between 0 and 1 reflect a mixed form. Some authors [8,14,15], similarly to us, defined an episomal form when E2/E6 = 1 or more, whereas Fujii et al [22] established the cut-off point on the basis on RT-PCR analysis of two plasmids solutions (prepared in increments of 20%), where one represents a pure episomal, and the other a pure integrated, form. They set the E2/E6 cut-off point at level of 0.79 based on 99% confidence interval calculation.…”

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Comparison of the sensitivity and specificity of real-time PCR and <i>in situ</i> hybridization in HPV16 and 18 detection in archival cervical cancer specimens

Biesaga¹,

Szostek

Klimek³

et al. 2012

Folia Histochem Cytobiol.

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Abstract:The aim of this study was to analyze the correlation between real-time PCR (RT-PCR) treated as a reference method and in situ hybridization with tyramide amplification system (ISH-TSA) in the detection of HPV16 and 18 infection and the assessment of viral genome status. The study was performed on cervical cancer biopsies fixed in 10% neutral buffered formalin and embedded in paraffin obtained from 85 women. TaqMan--based 5'exonuclease RT-PCR with type-specific primers was used to assess HPV16 and 18 infections and genome status. Viral infection and genome status was also assessed by ISH-TSA. RT-PCR revealed 76 (89.4%), and ISH-TSA 81 (95.3%) cancers with HPV16 and 18 infections. The ISH-TSA sensitivity and specificity were: 96.1% and 11.1% compared to RT-PCR. The difference between these techniques in HPV detection was significant (p = 0.000). Among 76 HPV16/18 positive cancers in RT-PCR, there were 30 (39.5%) with integrated and 46 (60.5%) with mixed viral genome form. According to ISH-TSA, there were 39 (51.3%) samples with integrated and 37 with mixed form (48.7%). The sensitivity and specificity of ISH-TSA in genome status assessment were 70.0% and 60.9%, respectively. The difference between RT-PCR and ISH-TSA in genome state detection was not statistically significant (p = 0.391). These results suggest that ISH-TSA shows insufficient specificity in HPV detection for use in clinical practice. However, this assay could be applied for viral genome status assessment.

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“…Feyza ve ark.nın yaptığı çalışmada CIn I olgularında %50 integral patern, %27.3 mikst patern, %22.7 epizomal patern saptanmıştır (7). Epizomal HPV-16'nın servikal karsinomlarda rol oynadığı bilinmektedir (8). Epizomal form HPV enfeksiyonu varlığında bile erken dönemlerde sitolojik ve histolojik bulguların negatif olabileceği gösterilmiş, bu nedenle HPV'nin dokuya integrasyonunu göstermede in situ hibridizasyonun PCR yöntemine üstünlüğü vurgulanmıştır.…”

Section: Cın ı Cın ıı Cın ııı Shk N (%) N (%) N (%) N (%)unclassified

Evaluation of hpv and p16 expression in the cervical intraepithelial neoplasia

Kimiloglu¹,

Gündoğar²,

Demir³

et al. 2017

JCurrPath

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Tel: 0 532 302 91 11 (J Curr Pathol. 2017;3:53-9.) ÖZET Amaç: serviks kanseri, önleme programlarına rağmen dünya genelinde kadın ölümlerinin önde gelen nedenlerindendir. bu çalışmanın amacı, servikal intraepitelyal neoplazi ve skuamöz hücreli karsinom olgularında, HPV-HR ve p16 değerlendirilmesi ile sitolojik ve histopatolojik sonuçların karşılaştırılmasıdır. Gereç ve Yöntem:Hastanemiz Patoloji Kliniğinde histopatolojik olarak servikal intraepitelyal neoplazi ve skuamöz hücreli karsinom tanısı almış 51 olgu daha önceki servikal smear sonuçları ile uyumluluk açısından değerlendirildi. silver in situ hibridizasyon yöntemi ile servikal intraepitelyal neoplazi ve skuamöz hücreli karsinom olgularında HPV-HR varlığı ve aynı doku biyopsilerinde immünohistokimyasal olarak p16 ekspresyonu değerlendirildi. Bulgular: smear sonuçlarına göre servikal intraepitelyal neoplazi düzeyinin farklılık gösterdiği saptandı. Mevcut bulgulardan smearlerinde yüksek dereceli skuamöz intraepitelyal lezyon ile ayrılamayan atipik skuamöz hücreler ve yüksek dereceli skuamöz intraepitelyal lezyon saptanmış olan olguların, doku biyopsilerinde CIn I'e nazaran daha çok CIn II ve CIn III tanısı aldıkları saptanmıştır. Aynı test ile değerlendirildiğinde HPV-HR düzeyine göre histolojik tanı tiplerinin farklılık gösterdiği saptandı. Histopatolojik incelemede CIn I olarak değerlendirilen olguların %35'inde, CIn II olarak değerlendirilen olguların %93.3'ünde, CIn III olarak değerlendirilen olguların %92.3'ünde ve skuamöz hücreli karsinom olarak değerlendirilen olguların %100'ünde p16 ile pozitif boyanma görülmüştür.Sonuç: servikal smear testi, in situ hibridizasyon yöntemi ile HPV-HR değerlendirilmesi ve immünohistokimyasal olarak p16 uygulanması, servikal intraepitelyal neoplazilerin erken tanı ve tedavisinde oldukça önemli ve gereklidir.Anahtar Sözcükler: serviks, sitoloji, Histopatoloji, HPV-HR, p16 ABSTRACT Aim: Cervical carcinoma is still one of the leading causes of mortality in women worldwide despite the preventing programmes. The aim of this study is to evaluate the comparison of results of the cytology and histopathology in cervical intraepithelial neoplasia and squamous cell carcinoma cases and to confirm the presence of HPV-HR and p16. materials and methods: We evaluated fifty one cases of cervical intraepithelial neoplasia and squamous cell carcinoma cases with their cytology results. HPV-HR was assesed by silver insitu hybridization and p16 immunohistochemical expressions in cervical intraepithelial neoplasia and squamous cell carcinoma cases. Results:The level of CIn showed a significant difference for cytological results. Cases with a cytological diagnosis of atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion and high-grade squamous intraepithelial lesion had more CIn II and CIn III diagnosis in their histological materials. Histopathological types showed statistically significant difference for HR-HPV levels. p16 positive staining was detected in 35% of CIn I, 93.3% of CIn II, 92.9% of CIn III, and 10...

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Comparison Between In Situ Hybridization and Real-time PCR Technique as a Means of Detecting the Integrated Form of Human Papillomavirus 16 in Cervical Neoplasia

Cited by 40 publications

References 27 publications

Host and viral factors in relation to clearance of human papillomavirus infection: A cohort study in Taiwan

Host and viral factors in relation to clearance of human papillomavirus infection: A cohort study in Taiwan

Comparison of the sensitivity and specificity of real-time PCR and <i>in situ</i> hybridization in HPV16 and 18 detection in archival cervical cancer specimens

Evaluation of hpv and p16 expression in the cervical intraepithelial neoplasia

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