CIN Lesions Research Articles

Association of lesion area measured by colposcopy and cervical neoplasia

Studies have demonstrated that the size of lesion in colposcopic imaging can be associated with the grade of CIN. We evaluated 36 patients and at the time of colposcopy, the images were recorded and analysed for lesion area measurement. A ROC curve was used to obtain the area under the curve and to determine the best cut-off values between area lesion (pixels2) and biopsy result. Fisher's exact test was performed (p < .05). Half of the sample had a cervical biopsy showing HPV or LSIL, and 18 (50%)a biopsy showing HSIL or invasive cancer. HSIL and invasive cancer were associated with a lesion area greater than 30,337.03 pixels2 (cut off) with p = .04. Thus the area of the colposcopic lesion is related to the severity of that; so small lesions can be more conservatively followed. IMPACT STATEMENT: What is already known on this subject? Studies have proposed that the size of lesion in colposcopic imaging can be associated with the grade of CIN, and the size of CIN lesions may be a factor in determining the risk of progression. What do the results of this study add? This is the first study in the literature that uses the measurement of the lesion area in pixels2 in comparison with the severity of the lesion, which provides greater accuracy of the lesion area than the mere measurement of its diameter. What are the implications of these findings for clinical practice and/or further research? The size of the lesion should be considered in the management of cervical intraepithelial lesions. This approach also leads to lower cost and is less invasive. Small lesions will have the best prognosis and would be treated in the way more conservative, bringing to the patients more comfort and less complications with the treatment.

Classification of high-grade cervical intraepithelial neoplasia by p16ink4a , Ki-67, HPV E4 and FAM19A4/miR124-2 methylation status demonstrates considerable heterogeneity with potential consequences for management.

High‐grade cervical intraepithelial neoplasia (CIN2 and CIN3) represents a heterogeneous disease with varying cancer progression risks. Biomarkers indicative for a productive human papillomavirus (HPV) infection (HPV E4) and a transforming HPV infection (p16ink4a, Ki‐67 and host‐cell DNA methylation) could provide guidance for clinical management in women with high‐grade CIN. This study evaluates the cumulative score of immunohistochemical expression of p16ink4a (Scores 0‐3) and Ki‐67 (Scores 0‐3), referred to as the “immunoscore” (IS), in 262 CIN2 and 235 CIN3 lesions derived from five European cohorts in relation to immunohistochemical HPV E4 expression and FAM19A4/miR124‐2 methylation in the corresponding cervical scrape. The immunoscore classification resulted in 30 lesions within IS group 0‐2 (6.0%), 151 lesions within IS group 3‐4 (30.4%) and 316 lesions within IS group 5‐6 (63.6%). E4 expression decreased significantly from CIN2 to CIN3 (P < .001) and with increasing immunoscore group (P trend < .001). Methylation positivity increased significantly from CIN2 to CIN3 (P < .001) and with increasing immunoscore group (P trend < .001). E4 expression was present in 9.8% of CIN3 (23/235) and in 12.0% of IS group 5‐6 (38/316). Notably, in a minority (43/497, 8.7%) of high‐grade lesions, characteristics of both transforming HPV infection (DNA hypermethylation) and productive HPV infection (E4 expression) were found simultaneously. Next, we stratified all high‐grade CIN lesions, based on the presumed cancer progression risk of the biomarkers used, into biomarker profiles. These biomarker profiles, including immunoscore and methylation status, could help the clinician in the decision for immediate treatment or a “wait and see” policy to reduce overtreatment of high‐grade CIN lesions.

CORRELATION OF BIOPSY AND AgNOR COUNTS IN PREMALIGNANT AND MALIGNANT LESIONS OF CERVIX

Background- Cervical cancer is the commonest cancer among women in developing countries. Screening methods like cervical cytology has brought down the incidence of cervical cancer. AgNOR represents Nucleolar organizer regions stained with silver and are related to cell proliferation rate and tumor malignant potential. The present study was aimed at evaluating the diagnostic role of AgNOR in cervical carcinogenesis. Material and Methods- The present study was conducted in the Pathology department of Gajra Raja Medical College, Gwalior from January 2019 to May 2020 on 500 patients who attended the Gynae OPD. Women of suspected cervical pathology and who underwent Pap smear test followed by subsequent biopsies were included. Results- AgNOR count in chronic cervicitis was 2.72±0.32, in CIN I was 4.02±0.38, in CIN II was 5.47±0.21 in CIN III was 5.59±0.18, in Well differentiated KSCC was 6.48±0.40 and in adenocarcinoma was 6.93±0.21. The study showed that the mean AgNOR count per nucleus was signicantly higher in CIN and malignant cervical lesions. Conclusion- AgNOR count is reproducible, simple, efcient and inexpensive method which can be used as an adjunct to histopathology and cytology especially in differentiating doubtful cases of CIN.

Human Papillomavirus Distribution in Women with Abnormal Pap Smear and/or Cervical Intraepithelial Neoplasia in Vaccination Era. A Single-Center Study in the North Italian Population.

Time trends prevalence of human papillomavirus (HPV) genotypes including negative and untypable infections were analyzed during a 15-year period (2005–2019) among 5807 subjects with abnormal pap-smears and/or cervical intraepithelial neoplasia (CIN). The rates of HPV16 dropped by 13% every 3 years (Prevalence Ratio, PR = 0.87, 95% CI = 0.82–0.93) in the CIN1 biopsy, while HPV16 status was unchanged over time in the CIN2+ biopsy. In CIN1 lesions, there was a corresponding increase of HR-HPV types unrelated to nonavalent vaccine. The rates of HPV 18, 31, and 52, decreased by 35% (PR = 0.65, 95% CI = 0.54–0.79), 19% (PR = 0.81, 95% CI = 0.73–0.91), and 21% (PR = 0.79, 95% CI = 0.73–0.86) every 3-year interval in CIN2+, respectively. Overall, the prevalence of negative/untypable HPV specimens in the entire database increased from 9.6% (129/1349) in the period 2011–2013 to 17.6% (161/913) and 28.4% (224/790) in the 2014–2016 period and in the 2017–2019 period, respectively (PR = 1.69, 95% CI = 1.52–1.88). HPV 16 prevalence decreased significantly among subjects with low-grade cervical squamous lesions. A significant increase of both HPV types unrelated to nonavalent vaccination and negative/untypable HPV infections was reported. The prevalence of HPV types among subjects with abnormal pap smears in Northern Italy is changing. Many variables including demographic factors and possibly vaccination could be responsible for this modification.

Age-related distribution of uncommon HPV genotypes in cervical intraepithelial neoplasia grade 3

AimCervical cancer prevention guidelines include Human Papillomavirus (HPV) test, cytology, and HPV-16/18 typing for triage to determine the risk of cervical intraepithelial neoplasia (CIN) grade 3 as the best proxy of cervical cancer risk. In doing that, they do not consider how age can modify the type-specific risk of CIN3. The present study aimed to evaluate the age-related distribution of HPV genotypes affecting the risk-assessment in cervical cancer screening programs: non-screening-type-HPV and non-HPV-16/18 in unvaccinated women with CIN3. MethodsRetrospective multi-institutional study, including HPV genotyped women with CIN3 on cone histology treated between 2014 and 2019. The sample was divided into three categories of age: <30, 30–44, ≥45. HPV genotypes were grouped in non-screening-type-HPV (not-including genotypes 16/18/31/33/35/39/45/51/52/56/58/59/66/68) and non-HPV-16/18. Associations and trends between different age-groups and HPV genotypes were measured. Results1332 women were analyzed. Non-screening-type-HPV CIN3 were 73 (5.5%). Non-HPV-16/18 were found in 417 participants (31.3%). Women over 45 associated with non-screening-type HPV [odds ratio (OR) = 1.87, 95% confidence interval (CI) 1.07–3.25; p = 0.027]. Non-screening-type-HPV prevalence increased significantly with age (3.9% vs 5.1% vs 9.0%, p = 0.016). Women under 30 showed a lower rate of non-HPV-16/18 (OR = 0.65, 95% CI 0.47–0.89; p = 0.007). There was a positive trend with age of non-HPV-16/18 CIN3 (23.6% vs 32.1% vs 38.0%, p = 0.0004). ConclusionThe proportion of CIN3 lesions unrelated to genotypes detected by primary screening tests increased with age. This implies that age probably modifies the risk of CIN3 and possibly of cancer associated with HPV types. The risk-based recommendation should take into consideration age to define the management of HPV positive women.

High-Risk Human Papillomavirus Testing, Genotyping, and Histopathologic Follow-up in Women With Abnormal Glandular Cells on Papanicolaou Tests.

This study examined the association of high-risk human papillomavirus (hrHPV) status and HPV genotype with histopathologic follow-ups in women with an atypical glandular cell (AGC) interpretation. Cases with AGC interpretation on a Papanicolaou (Pap) test were retrieved along with hrHPV testing, genotyping, and histologic follow-up results if available. A total of 561 AGC cases were identified, with histologic follow-up available for 471 cases (84%). The follow-up diagnoses included benign or reactive changes (60% of cases), low-grade cervical intraepithelial neoplasia (18%), high-grade cervical intraepithelial neoplasia (CIN2-3; 7%), cervical carcinoma (5%), and other malignancies (10%). Tests for hrHPV were positive in 128 of 426 (30%) cases, including HPV16 (30%), HPV18 (14%) and other HPV subtypes (56%). A positive hrHPV result significantly increased the risk of developing CIN2-3 or cervical carcinoma (odds ratio, 24.6; 95% CI, 9.9-58.9) and HPV16 or HPV18 further increased the risk (odds ratio, 49.5; 95% CI, 17.7-123.7). Our data demonstrate that in women with an AGC Pap interpretation, a positive hrHPV result, especially type 16 or 18, is associated with an increased risk of developing cervical CIN2-3 or higher lesions, suggesting potential implications of hrHPV testing for the management of patients with an AGC result on a Pap test.

Progression of CIN1/LSIL HPV Persistent of the Cervix: Actual Progression or CIN3 Coexistence.

Objective The natural history of the CIN1 lesions is characterized by an elevated rate of spontaneous regression (80%), some authors recognize a capacity to progress to HSIL in 10% of cases, and other authors do not recognize the capacity of progression of LSIL (CIN1). This study was aimed to evaluate the incidence of progression to HSIL (CIN3) in women with a histological diagnosis of LSIL (CIN1). Furthermore, to this end, we studied the histological outcomes of cone specimens collected by the LEEP. Methods All the data were retrospectively analyzed. All participants underwent a follow-up of 4 years, during which each woman underwent an HPV test and genotyping, cervical cytological sampling, or biopsy every six months. The endpoint was the histological confirmation of CIN3 lesions in any moment during follow-up. Results Progression to CIN3 occurred in 7 cases (1,5%). Analyzing the histological exams of the cones of the 7 cases that progressed to CIN3, we found the coexistence of CIN1 and CIN3 lesions in all cases. Conclusion After 4 years of follow-up, only 1.5% (7/475) of the women with LSIL developed CIN3, all within the first two years of follow-up, and were immediately treated. The most likely explanations for “progression” from LSIL to HSIL are (1) actual progression, (2) underdiagnosis of HSIL on initial biopsy, (3) overdiagnosis of HSIL on follow-up biopsy/cone, and (4) CIN3 arose de novo. Analyzing the histological exams of the cones of the 7 cases that progressed to high-grade, we found the coexistence of CIN1 and CIN3 lesions in all cases. Some recent studies have shown that a viral genotype corresponds to different lesions in the same cervix; therefore, CIN1 coexisting with CIN3 does not always indicate progression of CIN1. Other authors have doubted the capacity of LSIL to progress.

Hybrid capture 2 and cobas® 4800: Comparison of performance of two clinically validated tests for human papillomavirus primary screening of cervical cancer.

To compare, in a primary human papillomavirus screening setting, two different validated human papillomavirus tests, considering their analytical and clinical screening performances. In Tuscany, a human papillomavirus screening program was implemented in 2013. Hybrid capture 2 (Qiagen) was used for testing until May 2016, when it was replaced by the cobas® 4800 human papillomavirus test (Cobas; Roche). We evaluated the performance of Hybrid capture 2 and Cobas on: the same screening population in two different periods (before and after changing to Cobas); the same Hybrid capture 2-positive consecutive samples. Discordant samples (Hybrid capture 2-positive/Cobas negative) were typed on the L1 gene (reverse line blot, AB Analitica) and E6/E7 genes (BD Onclarity assay). In the considered time period (n = 37,775), human papillomavirus positivity was 9.8% and 7.4%, respectively, for Hybrid capture 2 and Cobas (p < 0.0001). At immediate colposcopy, the cervical intraepithelial neoplasia, grade 2 positive predictive value was, respectively, 23.8% and 34% (p < 0.001). At one-year recall, human papillomavirus persistence was, respectively, 40.6% and 62.2% (p < 0.0001). Of Hybrid capture 2-positive re-tested samples (n = 620), 32.4% were Cobas negative. Of discordant samples typed on L1, 7% were positive for the 12 high-risk human papillomavirus. Of the samples found to be negative for the 12 high-risk human papillomavirus types on L1, 14.5% were positive on E6/E7 typing. Among the discordant samples, the only two cervical intraepithelial neoplasia (CIN) grade 3 lesions were non-high-risk human papillomavirus positive on both L1 and E6/E7 typing. At baseline, Hybrid capture 2 showed greater human papillomavirus positivity and a lower CIN2+ positive predictive value than Cobas, which was more specific than Hybrid capture 2 in detection of high-risk human papillomavirus: 80% of discordant samples were confirmed as high-risk human papillomavirus negative. This higher analytical specificity determined the non-identification of two CIN3 lesions.

Prognostic role of immunohistochemical overexpression of the p16 protein in women under the age of 35 and diagnosed with HSIL (CIN2) subjected to "cervix sparing" excision.

To evaluate the role of immunohistochemical staining overexpression of p16 protein (p16 IHC) as a prognostic factor of persistence or recurrence of intraepithelial disease after excision procedure in young women diagnosed with HSIL (CIN2). 62 women with a histological diagnosis of HSIL (CIN2) subjected to "cervix sparing" excisional procedure were included in this retrospective study. All had age less than or equal to 35 years, negative history of immunosuppression, available follow-up, and assessment of the resection margins state. Immunohistochemical staining for the p16 protein was evaluated on reviewed and confirmed HSIL (CIN2) histological specimens with negative resection margins. The post-treatment follow-up, including cytology, colposcopy, and histology, ranged from a minimum of 6 months to a maximum of 60 months. The persistence or recurrence of SIL during the follow-up period was based on histologic referral and defined as "the presence of SIL", "the presence of HSIL" and "progression to HSIL (CIN3)". 31/62 patients were positive for immunostaining (p16 IHC+), and 31/62 were negative (p16 IHC-). Persistence or recurrence after excision occurred more frequently within the p16 IHC+ than in p16 IHC- group, both as SIL (29% p16 IHC- vs. 32.3% p16 IHC+, p = 0.783) and HSIL (6.5% p16 IHC- vs. 12.9% p16 IHC+, p = 0.671). None of the patients in the p16 IHC- group showed progression to CIN3 for the entire observation period, whereas 9.7% of p16 IHC+ women progressed to CIN3 lesion (p = 0.042). The p16 IHC positivity showed a significant association with progression to CIN3 in 5 years of follow-up (p = 0.029) and with the presence of SIL after two years of follow-up (p = 0.031). The differences between the two groups increased after two years post-treatment: the p16 IHC- patients still had SIL only in 3.2% of cases and no longer had HSIL, while the p16 IHC+ women still showed SIL in 19.4% and HSIL in 6.5% of cases. The negative predictive value (NPV) of p16 IHC in predicting SIL's presence after treatment increased with the severity of the lesion (NPV for SIL 70.97%, for HSIL 93.55%, for CIN3 100%). The study suggests that young patients with p16 IHC- HSIL (CIN2) have a better post-excisional course of the cervical intraepithelial disease compared to p16 IHC+ women and that p16 IHC could have prognostic utility during the long-term follow-up, especially in forecasting progression to CIN3 in consideration of the high NPV (up to 100%). The efficacy of the adjuvant HPV vaccination in the management of HSIL (CIN2) p16+ young women is to be evaluated as part of the fertility-sparing treatment.

Efficient combination of Human Papillomavirus Genotyping for the triage of women with Atypical Squamous Cells of Undetermined Significance in Chinese rural population: A population-based study.

Objective: In this prospective, population-based study, we evaluated the utility of high-risk human papillomavirus (HR-HPV) genotyping for triaging women with atypical squamous cells of undetermined significance (ASC-US) in the Chinese rural area.Methods: A total of 40,000 women were recruited from rural areas of Shanxi Province, China, between June 2014 and December 2014. Women with Pap results of ASC-US underwent HPV genotyping, colposcopy and histopathological examination. For those with normal cervixes or cervical intraepithelial neoplasia (CIN) 1 on the initial evaluation, a 2-year follow-up study was performed.Results: The reporting rate of ASC-US was 5.76% (2,304/40,000) in the study population. The detection rates of CIN 2 or above (CIN2+) and CIN 3 or above (CIN3+) in women with ASC-US were 7.28% and 1.75%, respectively. HPV 16 (39.53%), HPV 58 (17.83%), and HPV 52 (15.50%) were the three most prevalent HR-HPV genotypes among all women with ASC-US cytology. The five most common HR-HPV genotypes in CIN3+ lesions were HPV16, HPV58, HPV33, HPV31 and HPV18. Compared with the 15 HR-HPV testing, genotyping for a combination of HPV16/18/31/33/58 increased specificity significantly with virtually no loss of sensitivity for detecting CIN2+ and CIN3+ lesions, as well as significantly reduced colposcopy referral rate (23.15% vs 33.70%, p<0.01). In addition, in the 2-year follow-up period, women with infection of HPV16, 18, 31, 33 or 58 genotypes were the most likely population (92%, 23/25) to develop CIN2 lesion.Conclusion: Our results demonstrate that genotyping for a combination of HPV16/18/31/33/58 provides a more efficient and cost-effective model to risk-stratify women with ASC-US in the Chinese rural population.

DNA ploidy measurement and human papillomavirus in abnormal cervical cytology.

To compare the efficacy of high-risk human papillomavirus (HR-HPV) and DNA image cytometry (DNA-ICM) status for identifying high-grade cervical intraepithelial neoplasia or worse (≥CIN2). This cross-sectional study was performed in women undergoing follow-up procedure after a previous abnormal cervical cytology. Cervical cells were collected for HPV detection and DNA ploidy measurement. Biopsy samples were taken for histological confirmation. Sensitivity and specificity values for ≥CIN2 detection with HR-HPV and DNA-ICM were determined. HR-HPV was present in 74.5% of the women. The most frequent HPV infection was HPV 16, followed by HPV 31, 33 and 58. Aneuploidy was observed in 60.6% of all cases. Referral cytology revealed 78.0% sensitivity and 68.6% specificity for detecting a≥CIN2 lesion. The HR-HPV test alone showed 92.7% sensitivity, albeit it was not statistically different from DNA-ICM (88.1%, P>.05). Positivity for HPV or DNA-ICM resulted in 100% sensitivity. Higher specificity was observed for the combination of HR-HPV and DNA-ICM (88.6%), with no difference from DNA-ICM alone (85.7%, P>.05). DNA-ICM or HR-HPV positivity identified all cases of≥CIN2 in women undergoing follow-up procedure after a previous abnormal cervical cytology. Routine cervical cancer screening could be improved by the incorporation of DNA-ICM as a complementary method to primary screening to identify which women need closer follow-up.

AYURVEDIC MANAGEMENT OF ATROPHIC VAGINITIS WITH CERVICAL INTRAEPITHELIAL NEOPLASIA- A CASE REPORT

Vaginitis in postmenopausal women is termed as atrophic vaginitis, characterised by vaginal bleeding or spotting, vaginal discharge and discomfort or dryness in vulva. As the symptoms are not usually disclosed by women it may become chronic and can adversely affect the quality of life. In cervical intraepithelial neoplasia the squamous epithelium of cervix is replaced by the cells of varying degree of atypia. Even though CIN1 and CIN2 regress spontaneously, untreated CIN1 and CIN2 lesions may progress to invasive carcinoma. Ayurvedic management for the above conditions focuses on symptomatic relief and improvement of general health and immunity. Oral medication and Sthanika Chikitsa like Kshara Karma and Varthi play vital role in this. A 70 year old lady presented with complaints of burning sensation and itching over genital area. On examination haemorrhagic spots were observed on vaginal walls. She was sent to Early Cancer Detection Centre for cytological screening. The reports showed low grade sqamous intraepithelial lesion. She was managed with oral medications and Sthanika chikitsa. During follow up she was relieved from the symptoms like burning sensation and itching over genital area and vaginal discharge. Bleeding spots on vaginal walls were absent and cytology revealed a negative smear for intraepithelial lesion or malignancy. From this case report it is evidenced that Ayurvedic treatment modality is effective and successful for the management of Atrophic vaginitis with CIN.

Efficacy and safety of an adsorbent and anti-oxidative vaginal gel on CIN1 and 2, on high-risk HPV, and on p16/Ki-67: a randomized controlled trial

PurposeThe effect of SAM vaginal gel, a medical device containing adsorptive silicon dioxide and antioxidative sodium selenite and citric acid, on histologically-proven cervical intraepithelial neoplasia type 2 (CIN2) as well as p16 positive CIN1, and on the presence of the onco-marker p16 was investigated.Methods216 women aged 25–60 years were randomized to either receive an intravaginal daily dose of SAM gel for three 28-day periods, or be followed-up without intervention. The primary endpoint was efficacy, defined as a combined histological and cytological regression. At baseline and after 3 months participants had: a guided biopsy including p16 immunohistochemical (IHC) staining, only if a lesion was visible at colposcopy; a cervical smear for cytology, high-risk human papillomavirus (hr-HPV) and a p16/Ki-67 test. At 6 months a further cytology and p16/Ki-67 test was performed.ResultsRegression of CIN lesions was observed in 78 out of 108 patients (72.2%) in the SAM gel arm and in 27 out of 108 patients (25.0%) in the control arm. Similarly, the change in the p16/Ki-67 cytological test status was significantly in favor of the treatment arm. The prevalence of hr-HPV decreased significantly (p < 0.001) in the treatment arm, from 87.0% to 39.8%, while it slightly increased in the control arm, from 78.7% to 83.3%. At 6 months the cytological regression in the treatment group and the highly significant effect on p16/Ki-67 was still present.ConclusionSAM vaginal gel enhances the regression of cervical lesions and clears hr-HPV and p16/Ki-67 in smears significantly, thus offering an active non-destructive management to prevent cervical cancer.Trial registration numberISRCTN11009040, date of registration: 10/12/2019; https://doi.org/10.1186/ISRCTN11009040; retrospectively registered.

The Association of Molecular Biomarkers in the Diagnosis of Cervical Pre-Cancer and Cancer and Risk Factors in Senegalese.

Background:Cervical intraepithelial neoplasia (CIN) grading is subjective and affected by substantial rates of discordance among pathologists. Although recent studies have suggested that p16INK4a may be a useful surrogate biomarker of cervical neoplasia, Ki-67 and human papillomavirus testing have also been shown to be useful in detecting neoplasia. The purpose of this study was to determine the expression of p16INK4a and Ki-67 in cervical neoplasia and its correlations with cofactors. Methods: The study involved 69 patients with and without cervical neoplasia who underwent colposcopic directed biopsy. On each patient, two samples were taken; the first was used for immunohistochemistry and the second for molecular testing, using HPV16and18 genotyping Real-Time PCR Kit. Results: The study revealed the expression level of p16INK4a and Ki-67 in a descending order, from invasive squamous cell carcinoma (SCC), CIN2/3, CIN1 and non-dysplastic lesions. Correlations showed an association between the staining of p16NK4a and Ki-67 with the increase of age (OR: 1.79 (95%IC: 0.49 – 6.55), p = 0.037) and marital status (OR: 0.17 (95%IC: 0.04 – 0.68), p = 0.003). We found that the expressions of p16INK4a and Ki-67 were significantly different between invasive SCC vs non-dysplasia (OR: 44.57 (95%IC: 4.91 – 403.91), p<0.0001). The study showed significant correlation between HPV 16and18 infection with p16 INK4a and Ki-67 expression (OR: 0.13 (95%IC: 0.03 – 0.52), p<0.0001). Strong expression of p16INK4a and Ki-67 were observed in invasive squamous cell carcinoma, moderate staining was found in CIN2/3, weak staining in CIN1 and normal histology. Conclusion:Our findings indicate that p16INK4a and Ki-67 expressions associated strongly with cervical pathology. Therefore, p16/Ki-67 could be considered as a suitable biomarker for cervical cancer screening, particularly in HPV-based screening programs.

Use of Cap Analysis Gene Expression to detect human papillomavirus promoter activity patterns at different disease stages

Transcription of human papillomavirus (HPV) genes proceeds unidirectionally from multiple promoters. Direct profiling of transcription start sites (TSSs) by Cap Analysis Gene Expression (CAGE) is a powerful strategy for examining individual HPV promoter activity. The objective of this study was to evaluate alterations of viral promoter activity during infection using CAGE technology. We used CAGE-based sequencing of 46 primary cervical samples, and quantitatively evaluated TSS patterns in the HPV transcriptome at a single-nucleotide resolution. TSS patterns were classified into two types: early promoter-dominant type (Type A) and late promoter-dominant type (Type B). The Type B pattern was more frequently found in CIN1 and CIN2 lesions than in CIN3 and cancer samples. We detected transcriptomes from multiple HPV types in five samples. Interestingly, in each sample, the TSS patterns of both HPV types were the same. The viral gene expression pattern was determined by the differentiation status of the epithelial cells, regardless of HPV type. We performed unbiased analyses of TSSs across the HPV genome in clinical samples. Visualising TSS pattern dynamics, including TSS shifts, provides new insights into how HPV infection status relates to disease state.

Health Outcomes at 1 Year After Thermal Ablation for Cervical Precancer Among Human Papillomavirus- and Visual Inspection With Acetic Acid-Positive Women in Honduras.

PURPOSEThis study aims to assess the detection of cervical intraepithelial lesions grades 2 and 3 (CIN2-3) at 1 year after treatment with thermal ablation among human papillomavirus (HPV)–positive and visual inspection with acetic acid (VIA)–positive women.METHODSAll women screened and triaged for cervical cancer at four government health facilities in Honduras who were eligible for ablative treatment were enrolled and treated with thermal ablation. Women with confirmed CIN2-3 and a subset of women with CIN1/normal diagnoses at baseline were evaluated at 12 months. Follow-up procedures included HPV testing (careHPV), VIA, directed biopsy (if VIA-positive), and Papanicolaou test (if HPV positive, VIA negative). Outcomes at 1 year included histologic or cytologic assessment of CIN lesions among those with any abnormal test.RESULTSAmong the 319 women treated with thermal ablation, baseline histologic diagnoses were available for 317. Two (0.6%) had invasive cancer, 36 (11.4%) had CIN3, 40 (12.6%) had CIN2, and 239 (75.4%) had CIN1/normal histology. Among the 127 women eligible for follow-up, 118 (92.9%) completed all study procedures at 1 year. Overall, 98 (83.1%) had no evidence of CIN2-3 or persistent low-grade infection, 13 (11.2%) had CIN1/atypical squamous cells of undetermined significance, six (5.1%) had CIN2/high-grade squamous intraepithelial lesion, and 1 (0.8%) had a persistent CIN3. No adverse events associated with thermal ablation at 1 year were registered.CONCLUSIONA high proportion of women had no evidence of CIN2-3 at 1 year after thermal ablation treatment. Thermal ablation is an alternative to cryotherapy that may facilitate greater treatment coverage and prevent unnecessary deaths from cervical cancer.

Characterization of cervical biopsies of women with HIV and HPV co-infection using p16ink4a, ki-67 and HPV E4 immunohistochemistry and DNA methylation

This study aims to characterize cervical intraepithelial neoplasia (CIN) in women living with HIV using biomarkers. Immunohistochemical (IHC) staining for human papillomavirus (HPV) E4 protein indicates CIN with productive HPV infection, whereas Ki-67 and p16ink4a indicate CIN with transforming characteristics, which may be further characterized using DNA hypermethylation, indicative for advanced transforming CIN. Cervical biopsies (n = 175) from 102 HPV positive women living with HIV were independently reviewed by three expert pathologists. The consensus CIN grade was used as reference standard. IHC staining patterns were scored for Ki-67 (0–3), p16ink4a (0–3), and E4 (0–2) and correlated to methylation levels of four cellular genes in corresponding cervical scrapes. Reference standards and immunoscores were obtained from 165 biopsies:15 no dysplasia, 91 CIN1, 31 CIN2, and 28 CIN3. Ki-67 and p16ink4a scores increased with increasing CIN grade, while E4 positivity was highest in CIN1 and CIN2 lesions. E4 positive CIN1 lesions had higher Ki-67 and p16ink4a scores and higher methylation levels compared with E4 negative CIN1 lesions. E4 positive biopsies with low cumulative Ki-67/p16 ink4a immunoscores (0-3) had significantly higher methylation levels compared with E4 negative biopsies. No significant differences in Ki-67 and p16ink4a scores and methylation levels were observed between E4 negative and positive CIN2 or CIN3 lesions. The presence of high methylation levels in scrapes of CIN lesions with IHC characteristics of both productive (E4 positive) and transforming infections (increased Ki-67/p16ink4a expression) in women living with HIV might indicate a rapid aggressive course of HPV infections towards cancer in these women.

P63 expression profile in uterine cervical lesions

Background and objective: P63 is a member of the p53 gene family. It plays an essential role in the normal development of the cervix. This study aimed to assess p63 expression in different cervical lesions and the correlation between their expression in cervical intraepithelial neoplastic changes and invasive cervical cancer. Methods: A total of 62 cervical tissues were performed for p63 immunostaining, including 15 samples of non-neoplastic reactive cervicitis, 27 cervical intraepithelial neoplastic changes, and 15 invasive cervical cancer cases as well as five samples of normal cervical tissue as control. Results: P63 in normal cervical squamous epithelium had restricted staining pattern in the nuclei of basal cells. Reactive cervicitis staining ranged from restricted staining pattern in basal cells to mild increase in staining intensity involving upper parabasal cells. P63 was positive in all CIN lesions, and staining intensity increased significantly with increasing severity of cervical intraepithelial lesions compared with non-neoplastic epithelium. All cervical squamous cell carcinoma had high staining intensity while adenocarcinoma and small cell carcinoma were stained negative. Conclusion: P63 can be used as a marker for the distinction between reactive cervicitis and cervical intraepithelial changes. It is also of value for grading of CIN lesions and differentiation between squamous cell carcinoma, adenocarcinoma, and small cell carcinoma.

Clinical performance of the aptima HPV assay in 4196 women with positive high-risk HPV and ASC-US cytology: A large women hospital experience.

Despite Aptima assay as the latest US Food Drug Administration (FDA)-approved high-risk human papillomavirus (hrHPV) test has been implemented as an adjunct in cervical cancer screening for years, histological follow-up data remain limited with respect to its performance in women with ASC-US Pap tests and positive hrHPV results. Cases with results of ASC-US cytology and positive hrHPV by Aptima assay during the period 06/ 2015-02/2017 were retrieved from archived pathology reports. Immediate histological follow-up results were analyzed within 6 months interval after cotesting. Among 4196 women with ASC-US Pap tests and positive hrHPV, 51.1% of them had the immediate histological follow-up within 6 months. With positive Aptima hrHPV as the adjunct, 46.5% (95%CI 46.2-46.8) of ASC-US women were found to have cervical intraepithelial neoplasia type 1 (CIN1); 8.8% (95%CI 8.1-9.5) women were detected CIN2+ lesion including eight adenocarcinoma in-situ (AIS)s. CIN2+ detection rates were highest in women under 25 (15.4%, n = 65), when comparison with different age cutoffs, younger age women had higher CIN2+ lesion detection rate than that in older group (P <.01). This is by far one of the largest retrospective studies to analyze the histological follow-up results of ASC-US women with positive hrHPV tested by Aptima hrHPV mRNA assay. The results indicated that younger women with ASC-US and positive hrHPV testing have highest risk of developing high grade CIN lesions as compared to the older women. Lastly, with positive HPV as the adjunct, 55.3% (1186/2145) of ASC-US women will result in the positive finding on histological follow-up.

The role of EP-2 receptor expression in cervical intraepithelial neoplasia

Prostaglandin induced signalling is involved in different cancers. As previously described, the EP3 receptor expression decreases with increasing stage of cervical intraepithelial lesions (CIN). In addition, in cervical cancer EP3 is an independent prognosticator for overall survival and correlates with FIGO stages. Currently the role of Prostaglandin 2 receptor 2 (EP2) in CIN is unknown. The aim of this study was to analyse the expression of EP2 for potential prognostic value for patients with cervical dysplasia. EP2 expression was analysed by immunohistochemistry in 33 patient samples (CIN1–3) using the immune-reactivity scoring system (IRS). Expression levels were correlated with clinical outcome to analyse prognostic relevance in patients with CIN2. Data analysis was performed using non parametric Kruskal–Wallis and Spearman rank sum test. Cytoplasmic expression levels of EP2 correlated significantly (p < 0.001) with different grades of cervical dysplasia. Median EP2-IRS in CIN1 was 2 (n = 8), 3 in CIN2 (n = 9) and 6 in CIN3 (n = 16). Comparing regressive (n = 3, median IRS = 2) to progressive (n = 6, median IRS = 4) CIN2 cases the median IRS differed significantly (p = 0.017). Staining intensity (p = 0.009) and IRS (p = 0.005) of EP2 and EP3 correlate inversely. EP2 expression level significantly increases with higher grade of CIN and could qualify as a potential prognostic marker for the regressive or progressive course in CIN2 lesions. These findings emphasize the significant role of PGE2 signalling in CIN and could help to identify targets for future therapies.