Evaluation of CIN2/3 Lesion Regression in GynTect® DNA Methylation-Marker-Negative Patients in a Longitudinal Study.

Cervical intraepithelial neoplasia (CIN) grade 2/3 has a high spontaneous regression rate, especially among women ≤29 years of age. To reduce overtreatment, reliable prognostic biomarkers would be helpful. The main aim of this study was to analyze the negative predictive value of the methylation marker panel GynTect® for lesion regression. In this prospective, multicenter, longitudinal observational proof-of-concept study, women aged ≤29 years with histologically confirmed CIN2 (n = 24) or CIN3 (n = 36) were closely monitored without treatment for up to 24 or 12 months, respectively. The outcome was either regression, persistence, or progression of the lesion. For each patient, a single baseline sample (V0) for cytology, hrHPV detection and methylation analysis was taken. In a primary analysis, the negative predictive value (NPV) of a GynTect®-negative test result at V0 for regression was determined. We tested the null hypothesis NPV ≤ 70% against the alternative hypothesis NPV ≥ 90%. Twelve of the eighteen GynTect®-negative CIN2 patients showed regression (NPV = 67%, 90% CI 44-85%, p = 0.53). Of the 27 GynTect®-negative CIN3 lesions, 15 regressed (NPV = 56%, 90% CI 38-72%, p = 0.92). Although the majority of GynTect®-negative lesions regressed, the postulated NPV of ≥90% was not observed. Thus, the clinical relevance for an implementation of the GynTect® assay for patients undergoing watchful waiting remains questionable. Further studies with longer observation periods should be undertaken.

A Therapeutic Antigen-Presenting Cell-Targeting DNA Vaccine VB10.16 in HPV16-Positive High-Grade Cervical Intraepithelial Neoplasia: Results from a Phase I/IIa Trial.

PURPOSE: To evaluate the safety, immunogenicity and efficacy of a therapeutic DNA vaccine VB10.16, using a unique modular vaccine technology that is based on linking antigens to CCL3L1 targeting module, in women with HPV16-positive high-grade cervical intraepithelial neoplasia (CIN). PATIENTS AND METHODS: We conducted a first-in-human, open-label, phase I/IIa clinical trial of VB10.16 in subjects with confirmed HPV16-positive CIN 2/3. The primary endpoint was the proportion of participants with adverse events, including dose-limiting toxicities. Secondary outcome measures included measuring the E6/E7-specific cellular immune response. In the Expansion cohort HPV16 clearance, regression of CIN lesion size and grading were assessed during a 12-month follow-up period. RESULTS: A total of 34 women were enrolled: 16 in two dose cohorts and 18 in the expansion cohort. No serious adverse events or dose-limiting toxicities were observed, and none of the subjects discontinued treatment with VB10.16 due to an adverse event. Mild to moderate injection site reactions were the most commonly reported adverse event (79%). HPV16-specific T-cell responses were observed after vaccination in the majority of the subjects. In the expansion cohort, HPV16 clearance was seen in 8 of 17 evaluable subjects (47%). Reductions in lesion size were seen in 16 subjects (94%) and 10 subjects (59%) had regression to CIN 0/1. Correlation between strong IFNγ T-cell responses and lesion size reduction was statistically significant (P < 0.001). CONCLUSIONS: The novel therapeutic DNA vaccine VB10.16 was well tolerated and showed promising evidence of efficacy and strong HPV16-specific T-cell responses in subjects with high-grade CIN.

Evaluation of FAM19A4/miR124-2 methylation performance in the management of CIN3 diagnosed pregnant women.

Pregnant women diagnosed with CIN3 have high regression rates after delivery. Biomarkers are needed to only identify pregnant women with progressive CIN requiring treatment to reduce overreferral and overtreatment. In our study we evaluated the performance of the FAM19A4/miR124-2 methylation test for molecular triage on FFPE samples of CIN3+-diagnosed pregnant women with known clinical course over time as well in a cross-sectional setting. In this German multicenter retrospective study biopsy material was collected from pregnant women diagnosed with cervical cancer (n = 16), with CIN3 that progressed to cancer during pregnancy (n = 7), with CIN3 that regressed to CIN1 or less within 6 months after delivery (n = 41), without CIN (n = 16), CIN3 covering 3-4 quadrants (n = 14) and randomly selected CIN3 (n = 41). FAM19A4/miR124-2 methylation analysis was performed blinded on first diagnosis. All pregnant women with cervical cancer and with CIN3 progressing to cancer tested positive for FAM19A4/miR124-2 methylation (100%, 22/22). In the regressing CIN3 group 47.5% and in the group without CIN 21.6% tested methylation positive. High-volume CIN3 and random selected CIN3 were methylation-positive in 91.7% and 82.1%, respectively. Methylation levels were significantly higher in progressive CIN3 and cancer compared to the controls (P < .0005). The likelihood ratio of a negative methylation test (LR-) for progressive CIN3+ was 0 (95% CI: 0-0.208). A negative FAM19A4/miR124-2 methylation test can rule out progressive CIN disease in pregnant women diagnosed with CIN3. This can help the clinician by managing these pregnant women with conservative follow-up until after delivery.

Risk for Pelvic Metastasis and Role of Pelvic Lymphadenectomy in Node-Positive Vulvar Cancer-Results from the AGO-VOP.2 QS Vulva Study.

The need for pelvic treatment in patients with node-positive vulvar cancer (VSCC) and the value of pelvic lymphadenectomy (LAE) as a staging procedure to plan adjuvant radiotherapy (RT) is controversial. In this retrospective, multicenter analysis, 306 patients with primary node-positive VSCC treated at 33 gynecologic oncology centers in Germany between 2017 and 2019 were analyzed. All patients received surgical staging of the groins; nodal status was as follows: 23.9% (73/306) pN1a, 23.5% (72/306) pN1b, 20.4% (62/306) pN2a/b, and 31.9% (97/306) pN2c/pN3. A total of 35.6% (109/306) received pelvic LAE; pelvic nodal involvement was observed in 18.5%. None of the patients with nodal status pN1a or pN1b and pelvic LAE showed pelvic nodal involvement. Taking only patients with nodal status ≥pN2a into account, the rate of pelvic involvement was 25%. In total, adjuvant RT was applied in 64.4% (197/306). Only half of the pelvic node-positive (N+) patients received adjuvant RT to the pelvis (50%, 10/20 patients); 41.9% (122/291 patients) experienced recurrent disease or died. In patients with histologically-confirmed pelvic metastases after LAE, distant recurrences were most frequently observed (7/20 recurrences). Conclusions: A relevant risk regarding pelvic nodal involvement was observed from nodal status pN2a and higher. Our data support the omission of pelvic treatment in patients with nodal status pN1a and pN1b.

Evaluation of Integrated HPV DNA as Individualized Biomarkers for the Detection of Recurrent CIN2/3 during Post-Treatment Surveillance.

PURPOSE: Post-treatment follow-up in women with cervical pre-cancers (CIN3) is mandatory due to relapse in up to 10% of patients. Standard follow-up based on hrHPV-DNA/cytology co-testing has high sensitivity but limited specificity. The aim of our prospective, multicenter, observational study was to test the hypothesis that an individualized viral-cellular-junction test (vcj-PCR) combined with cytology has a lower false positive rate for the prediction of recurrence compared to standard co-testing. METHODS: Pre-surgical cervical swabs served for the identification of HPV16/18 DNA integration sites by next-generation-sequencing (NGS). Samples taken at 6, 12 and 24 months post-surgery were evaluated by cytology, hrHPV-DNA and the patients' individual HPV-integration sites (vcj-PCR on the basis of NGS). RESULTS: Integration sites were detected in 48 of 445 patients (10.8%), 39 of them had valid follow-up data. The false positive rate was 18.2% (95% CI 8.6-34.4%) for standard hrHPV/cytology at six months compared to 12.1% (95% CI 4.8-27.3%) for vcj-PCR/cytology, respectively (McNemar p = 0.50). Six patients developed recurrences (1 CIN2, 5 CIN3) during follow-up. Standard co-testing detected all, whereas vcj-PCR/cytology detected only five patients with recurrences. Data of 269 patients without evidence of HPV16/18 integration were subject to post-hoc analyses. Standard co-testing revealed a false positive rate of 15.7% (95% CI 11.7-20.7%) and predicted ten of fourteen recurrences at six months. CONCLUSIONS: Although highly specific on its own vcj-PCR could not detect all recurrent CIN2/3. Possible reasons for this unexpected result may be multifocal lesions, intratumoral heterogeneity with respect to HPV integration and/or incident CIN.

Significant decline of HPV 6 infection and genital warts despite low HPV vaccination coverage in young women in Germany: a long-term prospective, cohort data analysis.

BACKGROUND: The introduction of human papillomavirus (HPV) vaccination has resulted in a remarkable decline of genital warts in women and men, but in Germany historical rates of vaccination are relatively low. We report long-term surveillance data on changes in HPV 6 and HPV 11 infection and the prevalence of genital warts in young women in the Wolfsburg HPV epidemiological study (WOLVES). METHODS: Women born in 1983/84, 1988/89, and 1993/94 participated in four cohorts between 2009/10 and 2014/15. Quadrivalent vaccination coverage and prevalence of HPV 6/11 infection and genital warts are reported for participants aged 19-22 years and 24-27 years at the time of sample collection. Statistical analyses were done to compare similarly aged participants using 2 × 2 contingency tables (Röhmel-Mansmann unconditional exact test; two-side alpha of 0.05). RESULTS: A total of 2456 women were recruited. Between 2010 and 2015, there was a statistically significant decrease in the prevalence of HPV 6 infection among women aged 24-27 years (2.1% versus 0.0%; P < 0.0001) and women aged 19-22 years (2.0% versus 0.0%; P = 0.0056). There was no significant decline in HPV 11 infection. In total, 52 of 2341 participants were diagnosed with genital warts. There was a statistically significant drop in the risk of developing genital warts in women aged 24-27 years between 2010 and 2015 (4.7% versus 1.7%, respectively; P = 0.0018). The overall risk of developing genital warts in women aged 19-27 years decreased from 3.1% in 2010 to 1.2% in 2015 (P = 0.0022). CONCLUSIONS: An increase in vaccination coverage was associated with a decreased prevalence of genital warts in young women. A protective effect greater than herd immunity alone was seen despite low vaccination rates. Quadrivalent vaccine had a protective effect on genital HPV 6 infection and an almost fully protective effect on the development of genital warts in the youngest population.

Determinants of Human Papillomavirus Vaccine Uptake by Adult Women Attending Cervical Cancer Screening in 9 European Countries.

INTRODUCTION: Human papillomavirus-vaccinated cohorts, irrespective of age, will likely reduce their subsequent screening requirements, thus opening opportunities for global cost reduction and program sustainability. The determinants of uptake and completion of a 3-dose human papillomavirus vaccination program by adult women in a European context were estimated. STUDY DESIGN: This was an intervention study. SETTING/PARTICIPANTS: Study participants were women aged 25-45 years, attending opportunistic or population-based cervical cancer screening in Belgium, Denmark, Finland, France, Germany, Slovenia, Spain, Sweden, and the United Kingdom between April 2016 and May 2018. INTERVENTION: Study participants completed a questionnaire on awareness and attitudes on adult female human papillomavirus vaccination and were invited to receive free human papillomavirus vaccination. MAIN OUTCOME MEASURES: Main outcome measures were acceptance, uptake, and completion of vaccination schedule. Determinants of vaccine uptake were explored using multilevel logistic models in 2019. RESULTS: Among 3,646 participants, 2,748 (range by country=50%-96%) accepted vaccination, and 2,151 (range=30%-93%) received the full vaccination course. The factors associated with higher vaccine acceptance were previous awareness of adult female (OR=1.22, 95% CI=1.00, 1.48) and male (OR=1.59, 95% CI=1.28, 1.97) vaccination. Women in stable relationships (OR=0.56, 95% CI=0.45, 0.69) or with higher educational level (OR=0.76, 95% CI=0.63, 0.93) were more likely to refuse vaccination. Recruitment by postal invitation versus personal invitation from a healthcare professional resulted in lower vaccine acceptance (OR=0.13, 95% CI=0.02, 0.76). Vaccination coverage of >70% of adolescent girls in national public programs was of borderline significance in predicting human papillomavirus vaccine uptake (OR=3.23, 95% CI=0.95, 10.97). The main reasons for vaccine refusal were vaccine safety concerns (range=30%-59%) and the need for more information on human papillomavirus vaccines (range=1%-72%). No safety issues were experienced by vaccinated women. CONCLUSIONS: Acceptance and schedule completion were largely dependent on recruitment method, achieved coverage of national vaccination programs, and personal relationship status. Knowledge of benefits and safety reassurance may be critical to expanding vaccination target ages. Study results suggest that there are no major opinion barriers in adult women to human papillomavirus vaccination, especially when vaccination is offered face to face in healthcare settings. TRIAL REGISTRATION: EudraCT Number 2014-003177-42.

Reduction of cervical cancer incidence within a primary HPV screening pilot project (WOLPHSCREEN) in Wolfsburg, Germany.

BACKGROUND: Randomised controlled trials showed human papillomavirus (HPV)-based screening leads to a significant reduction in cervical cancer incidence compared with cytology-based screening only. METHODS: Non-hysterectomised participants ≥30 years underwent co-testing with Papanicolaou (Pap) smear and HR-HPV testing (Hybrid Capture 2; HC2). Women with normal findings had their next screening round after 5 years, and HC2+ and Pap abnormal cases were immediately referred for colposcopy, while cases with discordant findings had repeat testing after 12 months with referral to colposcopy in cases with persistent positive findings. RESULTS: Twenty-six thousand six hundred and twenty-four women were recruited between February 2006 and December 2016. Two hundred and seventy-four CIN3+ cases were diagnosed (270 HPV+, 4 HPV-), including 31 invasive cervical cancers (29 HPV+, 2 HPV-). No CIN3+ was detected in HPV- women with abnormal cytology. We observed a significant decline in the 5-year incidence of CIN3+ (from 0.96% [95% CI 0.85-1.09%] to 0.16% [95% CI 0.10-0.25%]; p < 0.0001) and cervical cancer (from 0.10% [95% CI 0.07%-0.15%] to 0.025% [95% CI 0.01-0.08%]; p = 0.01) between the first and subsequent rounds. Approximately 90% (246/274) of CIN3+ cases were diagnosed at first colposcopy. CONCLUSIONS: The decline in disease rates with 5-yearly co-testing seems mainly attributable to HPV testing since no CIN3+ occurred in HPV-/Pap+ women.

Angiopoietin-2 in adults with congenital heart disease and heart failure.

BACKGROUND: Chronic heart failure is an important cause for morbidity and mortality in adults with congenital heart disease (ACHD). While NT-proBNP is an established biomarker for heart failure of non-congenital origin, its application in ACHD has limitations. The angiogenic factors Angiopoietin-1 and -2 (Ang-1, Ang-2), vascular endothelial growth factor (VEGF), and soluble receptor tyrosine kinase of the Tie family (sTie2) correlate with disease severity in heart failure of non-congenital origin. Their role in ACHD has not been studied. METHODS: In 91 patients Ang-2 and NT-proBNP were measured and related to New York Heart Association class, systemic ventricular function and parameters of cardiopulmonary exercise testing. Ang-1, VEGF, and sTie2 were also measured. RESULTS: Ang-2 correlates with NYHA class and ventricular dysfunction comparable to NT-proBNP. Further, Ang-2 showed a good correlation with parameters of cardiopulmonary exercise testing. Both, Ang-2 and NT-proBNP identified patients with severely limited cardiopulmonary exercise capacity. Additionally, Ang-2 is elevated in patients with a single ventricle physiology in contrast to NT-proBNP. VEGF, Ang-1, and sTie2 were not correlated with any clinical parameter. CONCLUSION: The performance of Ang-2 as a biomarker for heart failure in ACHD is comparable to NT-proBNP. Its significant elevation in patients with single ventricle physiology indicates potential in this patient group and warrants further studies.

Asymmetrical dimethylarginine--more sensitive than NT-proBNP to diagnose heart failure in adults with congenital heart disease.

BACKGROUND: Chronic heart failure is an important cause for morbidity and mortality in adults with congenital heart disease (ACHD). While NT-proBNP is an established biomarker for heart failure of non-congenital origin, its value in ACHD has limitations. Asymmetrical dimethylarginine (ADMA) correlates with disease severity and independently predicts adverse clinical events in heart failure of non-congenital origin. Its role in ACHD has not been investigated. METHODS: In 102 patients ADMA and NT-proBNP were measured and related to NYHA class, systemic ventricular function and parameters of cardiopulmonary exercise testing. RESULTS: In contrast to NT-proBNP ADMA differentiated between NYHA classes I-III. Both, ADMA and NT-proBNP showed a good correlation with parameters of cardiopulmonary exercise testing with comparable receiver-operating characteristic curves for identifying patients with severely limited cardiopulmonary exercise capacity. CONCLUSION: ADMA seems to be a better biomarker than NT-proBNP for the assessment of NYHA class and as a good as NT-proBNP for the estimation of maximum exercise capacity in adults with congenital heart disease. Its use in clinical routine should be evaluated.

Symmetrical dimethylarginine outperforms CKD-EPI and MDRD-derived eGFR for the assessment of renal function in patients with adult congenital heart disease.

BACKGROUND/AIMS: Adults with congenital heart disease exhibit a 3-fold higher mortality in the presence of chronic kidney disease, hence assessment of renal function is crucial in this patient population. Formulas for the estimation of glomerular filtration rate (GFR) have not been evaluated in this patient population. Therefore, this study compares different markers and equations for the estimation of renal function in adults with congenital heart disease. METHODS: Renal function was assessed in 102 patients using the Modification of Diet in Renal Disease (MDRD) equation, the simplified MDRD equation, the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation and the Cockcroft-Gault formula. Additionally, symmetrical dimethylarginine (SDMA) was measured. Those parameters were compared to cystatin C-derived GFR using the Larsson equation. RESULTS: GFR estimates using the original MDRD (r = 0.465, p < 0.001) and the CKD-EPI equation (r = 0.462, p < 0.001) showed a similar strong correlation with the cystatin C-based eGFR equation, while eGFR using the simplified MDRD equation showed a slightly weaker correlation (r = 0.439, p < 0.001). The Cockcroft-Gault formula showed no correlation at all to the cystatin C-based eGFR (r = 0.144, p = 0.17). The strongest correlation was observed for SDMA and cystatin C-based eGFR (r = -0.552, p < 0.001). CONCLUSION: GFR in adults with congenital heart disease should be estimated using the original MDRD or the CKD-EPI formula. SDMA seems to be a promising marker of renal function for this patient group.