HPV mRNA Research Articles

Molecular Diagnosis for Nodal Metastasis in Endoscopically Managed Cervical Cancer: The Accuracy of the APTIMA Test to Detect High-risk Human Papillomavirus Messenger RNA in Sentinel Lymph Nodes

Study ObjectiveTo evaluate the feasibility and accuracy of a commercially available test to detect E6/E7 mRNA of 14 subtypes of high-risk HPVs (APTIMA; Hologic, Bedford, MA) in the sentinel lymph nodes of CC patients laparoscopically operated. DesignProspective pilot study. SettingThe study was conducted in the Department of Advanced Operative and Oncologic Gynecology, Asklepios Hospital, Hamburg, Germany. Patients54 women with HPV-positive CC submitted to laparoscopic sentinel node biopsy alone or sentinel node biopsy followed by systematic pelvic and/or para-aortic endoscopic lymphadenectomy. InterventionsAll removed sentinel lymph nodes (SLNs) underwent sample collection by cytobrush for the APTIMA assay before frozen section. MeasurementsResults obtained with the HPV mRNA test were compared with the definitive histopathological analysis of the SLNs and additional lymph nodes removed. ResultsA total of 125 SLNs (119 pelvic and 6 paraaortic) were excised with a mean number of 2.3 SLNs per patient. Final histopathologic analysis confirmed nodal metastases in 10 SLNs from 10 different patients (18%). All the histologically confirmed metastatic lymph nodes were also HPV E6/E7 mRNA positive, resulting in a sensitivity of 100%. Four histologically free sentinel nodes were positive for HPV E6/E7 mRNA, resulting in a specificity of 96.4%. ConclusionThe HPV E6/E7 mRNA assay in the SLNs of patients with CC is feasible and highly accurate. The detection of HPV mRNA in 4 women with negative SLNs might denote a shift from microscopic identification of metastasis to the molecular level. The prognostic value of this findings awaits further verification.

Mucosal alpha-papillomaviruses are not associated with esophageal squamous cell carcinomas: Lack of mechanistic evidence from South Africa, China and Iran and from a world-wide meta-analysis.

Epidemiological and mechanistic evidence on the causative role of human papillomaviruses (HPV) in esophageal squamous cell carcinoma (ESCC) is unclear. We retrieved alcohol- and formalin-fixed paraffin-embedded ESCC tissues from 133 patients seropositive for antibodies against HPV early proteins, from high-incidence ESCC regions: South Africa, China and Iran. With rigorous care to prevent nucleic acid contamination, we analyzed these tissues for the presence of 51 mucosotropic human alpha-papillomaviruses by two sensitive, broad-spectrum genotyping methods, and for the markers of HPV-transformed phenotype: (i) HPV16/18 viral loads by quantitative real-time PCR, (ii) type-specific viral mRNA by E6*I/E6 full-length RT-PCR assays and (iii) expression of cellular protein p16(INK4a). Of 118 analyzable ESCC tissues, 10 (8%) were positive for DNA of HPV types: 16 (4 tumors); 33, 35, 45 (1 tumor each); 11 (2 tumors) and 16, 70 double infection (1 tumor). Inconsistent HPV DNA+ findings by two genotyping methods and negativity in qPCR indicated very low viral loads. A single HPV16 DNA+ tumor additionally harbored HPV16 E6*I mRNA but was p16(INK4a) negative (HPV16 E1 seropositive patient). Another HPV16 DNA+ tumor from an HPV16 E6 seropositive patient showed p16(INK4a) upregulation but no HPV16 mRNA. In the tumor tissues of these serologically preselected ESCC patients, we did not find consistent presence of HPV DNA, HPV mRNA or p16(INK4a) upregulation. These results were supported by a meta-analysis of 14 other similar studies regarding HPV-transformation of ESCC. Our study does not support the etiological role of the 51 analyzed mucosotropic HPV types in the ESCC carcinogenesis.

Epidemiological, clinical, and virological characteristics of women with genital warts in Greece

This is a prospective study of the epidemiological, clinical, and virological characteristics of cases of genital warts in a Greek University Hospital. The women completed a questionnaire regarding their medical and sexual history and underwent cervical cytology, HPV DNA typing, mRNA testing, colposcopy, Chlamydia testing, and proctoscopy. Univariate and multivariate analyses were performed. The most commonly detected types were type 6 (36.1%) and 16 (24.3%). E6/E7 mRNA testing was positive in 21.5%. Concurrent cervical intraepithelial neoplasia grade 2 or worse was found in 11.1% and intra-anal warts in 10.4%. For chlamydial infection the number of sexual partners was a significant predictor. Women with warts infected with types 6 and 11 constituted only 37.5% of the total. This could have a negative effect on the efficacy of vaccination in reducing the incidence of the disease. Based on the present findings the authors recommend cytology and colposcopy for all women with genital warts.

Multifaceted usage of HPV related tests and products in the management of cervical cancer--a review.

HPV viruses are integral to the development of cervical cancer. The pathogenesis has been extensively studied. To date, numerous HPV tests and products have been developed and successfully utilized in diagnosis, treatment and prevention of cervical cancer. The HPV DNA test, when combined with other routine cervical cancer screening and diagnostic tests namely exfoliative cytology, visual inspection with acetic acid (VIA) and colposcopy has increased the detection rate of cervical cancer. HPV DNA products could also be measured in other body fluids like urine, lymph node tissue, and serum. HPV association could also be quantified by measuring other parameters like HPV mRNA, viral load, viral integration and methylation status. Vaccination against HPV has been found to decrease the incidence of cervical cancer. Further, therapeutic vaccines for cervical cancer against HPV continue to evolve. All these findings pertaining to HPV could possibly decrease the incidence of cervical cancer in the near future. This review aims to give an overview of the HPV tests and products in use and those under trial currently.

The Application of Classification and Regression Trees for the Triage of Women for Referral to Colposcopy and the Estimation of Risk for Cervical Intraepithelial Neoplasia: A Study Based on 1625 Cases with Incomplete Data from Molecular Tests.

Objective. Nowadays numerous ancillary techniques detecting HPV DNA and mRNA compete with cytology; however no perfect test exists; in this study we evaluated classification and regression trees (CARTs) for the production of triage rules and estimate the risk for cervical intraepithelial neoplasia (CIN) in cases with ASCUS+ in cytology. Study Design. We used 1625 cases. In contrast to other approaches we used missing data to increase the data volume, obtain more accurate results, and simulate real conditions in the everyday practice of gynecologic clinics and laboratories. The proposed CART was based on the cytological result, HPV DNA typing, HPV mRNA detection based on NASBA and flow cytometry, p16 immunocytochemical expression, and finally age and parous status. Results. Algorithms useful for the triage of women were produced; gynecologists could apply these in conjunction with available examination results and conclude to an estimation of the risk for a woman to harbor CIN expressed as a probability. Conclusions. The most important test was the cytological examination; however the CART handled cases with inadequate cytological outcome and increased the diagnostic accuracy by exploiting the results of ancillary techniques even if there were inadequate missing data. The CART performance was better than any other single test involved in this study.

Role of E6/E7 mRNA test in the diagnostic algorithm of HPV-positive patients showing ASCUS and LSIL: clinical and economic implications in a publicly financed healthcare system

Objective: Colposcopy is widely used to triage women with mild cervical abnormalities. However, this approach is associated with low specificity and predictive value. The efficacy of E6/E7 mRNA test for this purpose has been demonstrated, but studies estimating its cost-effectiveness are still lacking. Given the limited healthcare financial resources, such an evaluation is a priority. Methods: We analyzed the clinical history of 432 women referred to colposcopy and colposcopy-directed biopsy for persisting ASCUS and LSIL, and compared three alternative triage protocols: immediate colposcopy; reflex HPV DNA testing and HPV DNA plus mRNA tests in sequence. Results: Molecular tests in sequence significantly reduce colposcopy referral, cost for assessed women, and cost for CIN2 detected. On the other hand, incremental cost-effectiveness ratio of this protocol was the highest. Conclusion: Our preliminary data, providing an estimation of the economic burden deriving from the introduction of E6/E7 mRNA test in the triage algorithm of patients with mild cervical abnormalities, may be useful for future healthcare policy.

HPV DNA, p16INK4a and mRNA E6*I detection in anogenital cancers

HPV infection is a necessary cause for cervical cancer, and it is also causally linked with a range of different attributable fractions to other anogenital and head and neck cancers. The highest HPV DNA involvement in cancers other than cervix has been described in anal cancer (88.3%), followed by vaginal cancers (74.3%), peneal (33.0%) and vulvar (28.6). In high grade pre-neoplastic lesions from vulvar, vaginal, peneal and anal sites a very high HPV DNA detection rate is observed (over 85%). Agreement with biomarkers measuring the viral transforming activity is not 100%. The contribution of these additional markers is not well established when global estimates of HPV-attributable fractions (AF) in human cancer are provided. Objectives To evaluate the contribution of HPV type specific DNA, p16 INK4a and type specific mRNA E6*I in anogenital cancers derived from a large repository of parafin-embedded specimens. Methods HPV DNA (SPF-10/LiPA25), p16 INK4a and type specific mRNA E6*I are being evaluated in cancer and precancerous lesions of the vulva, vagina, anus and penis. The HPV contribution is estimated by considering different scenarios of biomarkers positivity and correcting for sensitivity drops. Preliminary results p16 INK4a positivity among HPV DNA positive cases was 98% in cervical cancer; 95% in anal cancer; 87% in cancer of the vagina; 83% in cancer of the vulva; and 70% in peneal cancer. The HPV contribution varies depending on the markers used e.g. in vulva cancer contribution would range from 29.4% if considering a positive staining with p16 INK4a , 27% with HPV DNA or 22% both combined. The meaning of discrepant results between biomarkers is still subject of discussion. Comprehensive analyses on HPV DNA, p16 INK4a and mRNA E6*I in anogenital locations will be provided. Discussion The best approach to estimate HPV contribution in a given tumor has relied mainly on the viral DNA detection. A better understanding of the additional information obtained by other markers like p16 INK4a or mRNA may refine our estimates of HPV contribution in related cancers at the population level.

Risk Factors for Locoregional Recurrence After Transoral Robotic Surgery for HPV+ Oropharyngeal Squamous Cell Carcinoma

DNA compared to mRNA suggests that RT induces viral activation and may explain the improved outcome seen in HPV-associated oropharyngeal cancer compared to uninfected patients. Furthermore, at the completion of RT HPV DNA and mRNA are undetectable in the saliva of most patients and 4 to 6 weeks later. Author Disclosure: T.R. Hollen: None. S.B. Zhang: None. W.M. Mendenhall: None. R.J. Amdur: None. A. Zhang: None. P. Okunieff: M. Stock; Paul Okunieff, MD, is a share stockholder in Enterade, LLC, Eva Pharmaceuticals, DiaCarta, LLC, and GainPep.

No causal association identified for human papillomavirus infections in lung cancer.

Human papillomavirus (HPV) infections have been implicated in lung carcinogenesis, but causal associations remain uncertain. We evaluated a potential causal role for HPV infections in lung cancer through an analysis involving serology, tumor DNA, RNA, and p16 protein expression. Association between type-specific HPV antibodies and risk of lung cancer was examined among 3,083 cases and 4,328 controls in two case-control studies (retrospective) and one nested case-control study (prospective design). Three hundred and thirty-four available tumors were subjected to pathologic evaluation and subsequent HPV genotyping following stringent conditions to detect all high-risk and two low-risk HPV types. All HPV DNA-positive tumors were further tested for the expression of p16 protein and type-specific HPV mRNA. On the basis of the consistency of the results, although HPV11 and HPV31 E6 antibodies were associated with lung cancer risk in the retrospective study, no association was observed in the prospective design. Presence of type-specific antibodies correlated poorly with the presence of the corresponding HPV DNA in the tumor. Although nearly 10% of the lung tumors were positive for any HPV DNA (7% for HPV16 DNA), none expressed the viral oncogenes. No association was observed between HPV antibodies or DNA and lung cancer survival. In conclusion, we found no supportive evidence for the hypothesized causal association between HPV infections and lung cancer.

Comparison of the performance of the NucliSENS EasyQ HPV E6/E7 mRNA assay and HPV DNA chip for testing squamous cell lesions of the uterine cervix

This study aims to evaluate the clinical performance of the NucliSENS EasyQ assay and compare it with HPV DNA genotyping for the detection of high-grade squamous intraepithelial lesions (HSIL) and cancer in a Korean population. In 188 total thin prep samples, the remaining fluid after cytology slide preparation was tested with Goodgene HPV DNA chips and the NucliSENS EasyQ HPV E6/E7 messenger RNA (mRNA) assay. The sensitivity and specificity of each test were calculated with HSIL and squamous cell carcinoma (SCC) as the disease endpoint. Out of the 188 samples, 139 (74%) were positive for DNA of 14 HPV types, while 57 (30%) cases were positive for E6/E7 mRNA. The DNA test was positive in cytology cases of SCC, HSIL, and atypical squamous cell. The mRNA test yielded results of 75%, 74%, 60%, 56%, and 29% positivity in abnormal cytology cases of SCC, HSIL, atypical squamous cells – cannot exclude HSIL, atypical squamous cells of undetermined significance, and low-grade squamous intraepithelial lesion, respectively. In normal cytology cases, the positivity rates were 9% and 53% for the mRNA and DNA tests, respectively. For detection of HSIL and SCC, the sensitivity of the mRNA test was 74.36% and that of the DNA test was 100%, while the specificities of the tests were 85% and 40.83%, respectively. These findings suggest that the HPV E6/E7 mRNA assay can overcome the shortcoming of low specificity of DNA assays for clinical detection of high-grade cervical lesions and malignancies.

The importance of HPV testing in cervical cancer prevention

Introduction: Cervical cancer is second most common malignant tumor of female reproductive system in the world. HPV infection is present in 99% of cases, therefore very important in its development.Material and method: Our research was conducted in Primary health care center for women in Podgorica and included 100 women aged 20-60. During the examination PAP testing and colposcopy were performed to every participant. HPV mRNA detection test with differentiation of types 16, 18, 31, 35, 55 was done in cases of abnormal PAP test result, or suspected clinical or colposcopic examination accompanied by normal PAP test.Results: PAP testing showed II group in 72 patients, IIIa in 19 and IIIb in 3. HPV testing was performed in 26 patients (in which 4 with PAP II result). It was positive for presence of high-oncogenic types in 15 patients (57,69%) and negative in 11 (42,31%). In patients with detected lesion by examination or colposcopy, we performed biopsy. Results showed CIN I in 1 patient, CIN II in 2, CIN III in 5 and invasive carcinoma in 1 patient.Conclusion: Our research showed great importance of HPV testing in patients with false negative PAP tests; its importance as diagnostic marker in prognosis of illness, as well as the fact that HPV testing, as basic one, without examination and PAP testing, doesn’t make sense as it would increase the number of false negative results.

High-Risk Human Papillomavirus Messenger RNA Testing in Physician- and Self-Collected Specimens for Cervical Lesion Detection in High-Risk Women, Kenya

Little is known about the performance of physician-versus self-collected specimens for high-risk human papillomavirus (hrHPV) messenger RNA (mRNA) testing or risk factors for hrHPV mRNA positivity in physician- versus self-collected specimens. We compared the performance of hrHPV mRNA testing of physician- and self-collected specimens for detecting cytological high-grade squamous intraepithelial lesions or more severe (≥HSIL) and examined risk factors for hrHPV mRNA positivity in female sex workers in Nairobi. From 2009 to 2011, 344 female sex workers participated in this cross-sectional study. Women self-collected a cervicovaginal specimen. A physician conducted a pelvic examination to obtain a cervical specimen. Physician- and self-collected specimens were tested for hrHPV mRNA and sexually transmitted infections using APTIMA nucleic acid amplification assays (Hologic/Gen-Probe Incorporated, San Diego, CA). Cervical cytology was conducted using physician-collected specimens and classified according to the Bethesda criteria. Overall hrHPV mRNA prevalence was similar in physician- and self-collected specimens (30% vs. 29%). Prevalence of ≥HSIL was 4% (n = 15). Overall sensitivity of hrHPV testing for detecting ≥HSIL was similar in physician-collected (86%; 95% CI, 62%-98%; 13 cases detected) and self-collected specimens (79%; 95% CI, 55%-95%; 12 cases detected). Overall specificity of hrHPV mRNA for ≥HSIL was similar in both physician-collected (73%; 95% CI, 68%-79%) and self-collected (75%; 95% CI, 70%-79%) specimens. High-risk HPV mRNA positivity in both physician- and self-collected specimens seemed higher in women who were younger (<30 years), had Trichomonas vaginalis or Mycoplasma genitalium infections, or had more than 8 years of educational attainment. Self-collected specimens for hrHPV mRNA testing seemed to have similar sensitivity and specificity as physician-collected specimens for the detection of ≥HSIL among high-risk women.

Comparison of the sensitivities of three commercial assays for detection of the high risk HPV types 16, 18 and 45

The relative and analytical sensitivity of the APTIMA HPV test (AHPV, broad-spectrum, target amplification) and the PreTect HPV-Proofer (type-specific, target amplification) for the detection of HPV mRNA in various cell lines was compared. Equivalent relative sensitivity for the HPV 16-containing cell lines (2.5cells/ml with both CaSki and SiHa) was observed for the mRNA assays – and similar sensitivities were observed for the detection of HPV 18 (HeLa) and 45 (MS751); ranging from 2.5cells/ml (Proofer) to 25cells/ml (APTIMA). In relation to analytical sensitivity, again, the mRNA assays showed similar sensitivities to each other, ranging from 0.1 to 1cell per reaction for APTIMA and 0.1 to 10cells per reaction for PreTect HPV-Proofer (depending on cell line). Both mRNA assays consistently achieved a higher analytical sensitivity than a DNA based comparator – the Hybrid Capture 2 High-Risk HPV DNA test (hc2). This study indicates that mRNA tests had high analytical sensitivity, higher than a well established DNA-test based when using cell lines as target. Implications for clinical application are discussed.

Detection of specific HPV subtypes responsible for the pathogenesis of condylomata acuminata

BackgroundThe low-risk human papillomavirus types 6 and 11 are responsible for approximately 90% of anogenital wart cases, with approximately 190,000 new and recurrent cases reported in the UK in 2010. The UK has recently selected the quadrivalent HPV vaccine, which conveys protection against both HPV6 and HPV 11, as part of its immunisation programme for 2012 and it is expected that this will reduce disease burden in the UK. The aims of the study were to evaluate current strategies used for the monitoring of HPV infection in genital warts and to assess the suitability of laser-capture microdissection (LCM) as a technique to improve the understanding of the natural history of HPV types associated with genital wart lesions.MethodsDNA and RNA were extracted from whole wart, surface swabs and LCM sections from 23 patients. HPV types present were determined using the Linear Array HPV Genotyping Test (Roche), with HPV DNA viral load and mRNA expression investigated using qPCR and qRT-PCR, respectively.ResultsResults indicated that swabbing the surface of warts does not accurately reflect potential causative HPV types present within a wart lesion, multiple HPV types being present on the surface of the wart that are absent in the lower layers of tissue isolated by LCM. Although it was shown that HPV DNA viral load does not directly correlate with HPV mRNA load, the presence of both DNA and mRNA from a single HPV type suggested a causative role in lesion development in 8/12 (66.6%) of patients analysed, with dual infections seen in 4/12 (33.3%) cases. HPV 6 and HPV 11 were present in more than 90% of the lesions examined.ConclusionsSurface swabbing of warts does not necessarily reflect the causative HPV types. HPV type specific DNA and mRNA loads do not correlate. HPV 6 and 11 were likely to be causally involved in over 90% of the lesions. Dual infections were also found, and further studies are required to determine the biological and clinical nature of dual/multiple infections and to establish the relationship of multiple HPV types within a single lesion.

Abstract 3482: Directly Detect High Risk HPV Oncogenes E6/E7 mRNAs from Pap Smear Samples without RNA Purification and RT-PCR.

Abstract Cervical cancer is one of the most common cancers in women, with about 450,000 new cases diagnosed each year. Nearly all the cervical cancers are from Papillomaviruses (HPV) infection. HPV are DNA-based viruses that infect the skin and mucous membranes of humans and many other animals. Most cancers of the vulva and vagina are induced by oncogenic HPV types. In precancerous lesions, most HPV genomes persist in an episomal state whereas, in many high-grade lesions and carcinomas, genomes are found integrated into the host chromosome. Two viral genes, E6 and E7, are invariably expressed in HPV-positive cancer cells. Their gene products are known to inactivate the major tumour suppressors, p53 and retinoblastoma protein (pRB), respectively. In addition, E6 oncoprotein is also capable to up regulate the expression of inhibitors of apoptosis, and E6 and E7 cooperate to effectively immortalise primary epithelial cells. It has been demonstrated that HPV E6/E7 expression level plays a key role in the progression of invasive carcinoma of the uterine cervix via the deregulation of cellular genes controlling tumour cell proliferation. HPV E6/E7 oncogenes have been proved to be good biological markers for prognosis assessment and specific therapy of the disease. We have developed a sandwich nucleic acid hybridization assay using branched DNA to amplify the signals. The assay can simultaneously detect E6/E7 mRNAs of all 14 high risk HPV subtypes directly from Pap smear samples without RNA purification and RT-PCR. All HPV mRNA targets are captured through cooperative hybridization of multiple probes and probe set design determines the specificity of each HPV subtype of E6/E7 mRNA. Probe set oligonucleotides bind a contiguous region of the target E6/E7 mRNAs and selectively capture target RNAs to a solid surface during hybridization. Signal amplification is performed via sequential hybridization of Pre-Amplifier, Amplifier and label probe. The assay is highly specific and sensitive, which can detect as low as 50 transcript molecules. Citation Format: Aiguo Zhang, Lulu Zhang, Rachel Chuang, Xining Zhu, Rachel Diaz, Lily Chen. Directly Detect High Risk HPV Oncogenes E6/E7 mRNAs from Pap Smear Samples without RNA Purification and RT-PCR. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3482. doi:10.1158/1538-7445.AM2013-3482

HPV mRNA Testing in Triage of Women with ASC-US Cytology May Reduce the Time for CIN2+Diagnosis Compared with Repeat Cytology

HPV mRNA Testing in Triage of Women with ASC-US Cytology May Reduce the Time for CIN2+Diagnosis Compared with Repeat Cytology

HPV mRNA Testing in Triage of Women with ASC-US Cytology May Reduce the Time for CIN2+ Diagnosis Compared with Repeat Cytology

In delayed HPV triage women with atypical squamous cells of uncertain significance (ASC-US) cytology are retested after 6-12 months in order to decide whether they should be referred for colposcopy, further follow-up cytology or routine screening in three years. Triage using a specific HPV E6/E7 mRNA test may reduce referrals for colposcopy of women with ASC-US cytology compared to HPV DNA testing. We explored whether HPV mRNA triaging could reduce the time from ASC-US index cytology to biopsy compared with repeat cytology, and whether the positive predictive value (PPV) of the HPV mRNA test for high grade cervical intraepithelial neoplasia (CIN2+) was comparable with the PPV of repeat cytology. We used repeat cytology and the HPV mRNA test PreTect HPV-Proofer, which detects E6/E7 mRNA from HPV subtypes 16, 18, 31, 33 and 45, in the triage of women with ASC-US. We included all women from the two northernmost counties of Norway with a first ASC-US cytology during the period 2004-2008. Two triage methods were evaluated 1) only repeat cytology (n=964) and 2) both HPV mRNA testing and cytology (n=542). Histologically confirmed CIN2+ was the study endpoint. Among 1506 women with an ASC-US index cytology, 59 women (3.9%) had biopsy taken, of whom 49 women had CIN2+ (PPV 83.1%). The mean time from index ASC-US cytology until the case was resolved (biopsy or return to screening) was 10.6 months in the repeat cytology group and 7.3 months in the HPV group (P < 0.001). Of the 964 women in the group with repeat cytology only, 35 women (3.6%) had biopsy and 30 had CIN2+ (PPV 85.7%). Of the 542 women in the group with both HPV test and cytology, 24 women (4.4%) had biopsy and 19 had CIN2+ (PPV 79.2%). In triage of women with ASC-US, the HPV mRNA test significantly reduced the time from the first abnormal cytology until biopsy and had predictive values comparable with those of repeat cytology.

Cervical intraepithelial neoplasia- predictive molecular growth factors in natural history.

There is considerable controversy regarding the possible over-treatment of patients with mild Cervical Intraepithelial Neoplasia (CIN), with lesions being often excised or ablated. Thus, identifying the markers of potentially malignant lesions would be of a great prognostic value. In the current study, we hypothesized that using colposcopic, cytological and histological findings together with assessing the expression of molecular growth factors can predict CIN outcome. The study group consisted of 285 women between 19 and 81 years of age (median age, 37,8 years). The follow up were 60 months and considered 138 women: 50 women with Subclinical Papillomavirus Infection (SPI), 50 women with CIN1 and 38 women with CIN2. All patients underwent cytology, colposcopy, and sampling for subsequent testing for HPV. In cases in which colposcopy suggested the presence of suspicious lesions, biopsy specimens were taken. HPV DNA was genotyped for HPV types 16, 18, 31, 33, and 45 by multiplex PCR. Transcripts of HR HPV types 16, 18, 31, 33, and 45 were detected by the NucliSens EasyQ HPV assay. The VEGF expression was analyzed with immunohistochemistry, RNA extraction, cDNA synthesis and RT-PCR analysis and Western blot. We found that so called lymphangiogenetic switch (over expression of VEGF C and VEGFR-2) appears already in CIN 2, which is a rare observation, Persistent HPV HR infection is not only a trigger but also a maintenance factor in the cervical carcinogenesis. CIN2/3 and cervical cancer is in high percentage associated with the presence of HR DNA HPV as well as E6/E7 DNA mRNA. In CIN2/3 and cervical cancer VEGF and its receptor expression correlate with the stage of cervical carcinogenesis. Progression of cervical intraepithelial neoplasia occurs when co expression of all: HR DNA HPV, E6/E7 HR HPV mRNA and VEGF is present.

MRNA and DNA Detection of Human Papillomaviruses in Women of All Ages Attending Two Colposcopy Clinics

ObjectiveHPV infection is a common finding, especially in young women while the majority of infections are cleared within a short time interval. The aim of this study was to examine the efficacy of HPV DNA and mRNA testing in a population attending colposcopy units of two University hospitals.Methods1173 liquid based cervical samples from two colposcopy clinics were tested for HPV DNA positivity using a commercial typing kit and HPV E6/E7 mRNA positivity with a flow cytometry based commercial kit. Statistic measures were calculated for both molecular tests and morphological cytology and colposcopy diagnosis according to histology results.ResultsHPV DNA, high-risk HPV DNA, HPV16 or 18 DNA and HPV mRNA was detected in 55.5%, 50.6%, 20.1% and 29.7% of the cervical smears respectively. Concordance between the DNA and the mRNA test was 71.6% with their differences being statistically significant. Both tests’ positivity increased significantly as lesion grade progressed and both displayed higher positivity rates in samples from women under 30 years old. mRNA testing displayed similar NPV, slightly lower sensitivity but significantly higher specificity and PPV than DNA testing, except only when DNA positivity for either HPV16 or 18 was used.ConclusionsOverall mRNA testing displayed higher clinical efficacy than DNA testing, either when used as a reflex test or as an ancillary test combined with morphology. Due to enhanced specificity of mRNA testing and its comparable sensitivity in ages under 25 or 30 years old, induction of mRNA testing in young women could be feasible if a randomized trial verifies these results.

O117 HPV DNA AND mRNA EXPRESSION IN CIN2 MIMICS CIN1 MORE THAN CIN3

International Journal of Gynecology & ObstetricsVolume 119, Issue S3 p. S301-S301 Free communication (oral) presentations O117 HPV DNA AND mRNA EXPRESSION IN CIN2 MIMICS CIN1 MORE THAN CIN3 J.-L. Brun, J.-L. BrunSearch for more papers by this authorN. Nocart, N. NocartSearch for more papers by this authorV. Charbonneau, V. CharbonneauSearch for more papers by this authorO. Delorme, O. DelormeSearch for more papers by this authorM. Marty, M. MartySearch for more papers by this authorI. Garrigue, I. GarrigueSearch for more papers by this author J.-L. Brun, J.-L. BrunSearch for more papers by this authorN. Nocart, N. NocartSearch for more papers by this authorV. Charbonneau, V. CharbonneauSearch for more papers by this authorO. Delorme, O. DelormeSearch for more papers by this authorM. Marty, M. MartySearch for more papers by this authorI. Garrigue, I. GarrigueSearch for more papers by this author First published: 22 October 2012 https://doi.org/10.1016/S0020-7292(12)60547-5AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat No abstract is available for this article. Volume119, IssueS3Abstracts of XX FIGO World Congress of Gynecology and ObstetricsOctober 2012Pages S301-S301 RelatedInformation