Identification of women for referral to colposcopy by neural networks: A preliminary study based on LBC and molecular biomarkers

Objective of this study is to investigate the potential of the learning vector quantizer neural network (LVQ-NN) classifier on various diagnostic variables used in the modern cytopathology laboratory and to build an algorithm that may facilitate the classification of individual cases. From all women included in the study, a liquid-based cytology sample was obtained; this was tested via HPV DNA test, E6/E7 HPV mRNA test, and p16 immunostaining. The data were classified by the LVQ-NN into two groups: CIN-2 or worse and CIN-1 or less. Half of the cases were used to train the LVQ-NN; the remaining cases (test set) were used for validation. Out of the 1258 cases, cytology identified correctly 72.90% of the CIN-2 or worst cases and 97.37% of the CIN-1 or less cases, with overall accuracy 94.36%. The application of the LVQ-NN on the test set allowed correct classification for 84.62% of the cases with CIN-2 or worse and 97.64% of the cases with CIN-1 or less, with overall accuracy of 96.03%. The use of the LVQ-NN with cytology and the proposed biomarkers improves significantly the correct classification of cervical precancerous lesions and/or cancer and may facilitate diagnosis and patient management

The influence of sexual behavior and demographic characteristics in the expression of HPV-related biomarkers in a colposcopy population of reproductive age greek women

Despite the significant scientific evolution in primary and secondary cervical cancer prevention in the battle started by George Papanicolaou in the previous century, global cervical cancer mortality rates remain disappointing. The widespread implementation of HPV-related molecular markers has paved the way to tremendous developments in cervical cancer screening, with the transition from cytological approach to the more accurate and cost-effective HPV testing modalities. However, the academic audience and different health systems have not yet adopted a universal approach in screening strategies, and even artificial intelligence modalities have been utilized from the multidisciplinary scientific armamentarium. Combination algorithms, scoring systems as well as artificial intelligent models have been so far proposed for cervical screening and management. The impact of sexual lifestyle inherently possesses a key role in the prevalence of HPV-related biomarkers. This study aimed to investigate any possible influence of sexual behavior and demographic characteristics in the expression of HPV-related biomarkers in a colposcopy population from October 2016 to June 2017, and corroborated the determining role of age at fist intercourse; the older the age, the lower the probability for DNA positivity. Multivariate analysis illustrated additionally that a number of sexual partners exceeding 4.2 was crucial, with women with ≤5 partners being approximately four times less likely to harbor a positive HPV DNA test (p < 0.0001). Similarly, a reported partner change during the last year before HPV DNA assessment contributed to 2.5 times higher odds for DNA positivity (p = 0.0006). From this perspective, the further development and validation of scoring systems quantifying lifestyle factors that could reflect cervical precancer risk seems paramount. © 2021 by the authors. Licensee MDPI, Basel, Switzerland

Effect of Condom Use after CIN Treatment on Cervical HPV Biomarkers Positivity: Prolonged Follow Up Study

Background: Several factors contribute in the cervical healing process following local surgical treatment; in a previous work our group has documented a beneficial mid-term role of regular condom use immediately postoperatively in terms of CIN relapse prevention and expression of active viral biomarkers. Materials and Methods: Aiming to investigate whether the favorable contribution of consistent condom use could be extrapolated in the longer term, we conducted a prospective single center observational study including women scheduled to undergo conservative excisional treatment for CIN (LLETZ procedure). In all women a strong recommendation for consistent use for the first 6 months was given. For 204 women who underwent the procedure and completed successfully the two-year follow up a complete dataset of HPV biomarkers’ results obtained six months and two years postoperatively was available. Patients were asked to complete a questionnaire to assess condom use compliance. A 90% compliance rate represented the threshold for consistent use. An LBC sample was obtained and tested for HPV genotyping, E6 & E7 mRNA by NASBA technique as well as flow cytometry, and p16 at 0 (pre-treatment), 6 and 24 months. HPV DNA and other related biomarkers status at 6 and 24 months, treatment failures at 24 months and condom use compliance rates represented study outcomes. Results: Six months post-operatively we documented a reduction in the rates of HPV DNA positivity, which was detected in only 23.2% of compliant condom users in comparison to 61.9% in the non-compliant group (p < 0.001, OR: 0.19, 95%CI: 0.1–0.36). For the HPV mRNA test, either assessed with the NASBA method or with flow cytometry, reduced positivity percentages were observed in the compliant group, in particular 1.6% vs. 8% for NASBA and 7.1% vs. 16.4% using flow cytometry, although these differences were not statistically significant (p = 0.1039 and 0.0791, respectively). Finally, reduced p16 positivity rates were documented in the compliant group. At the two year follow up, a more pronounced difference in HPV DNA positivity rates was observed, specifically only 13% positivity among the compliant women compared with 71% of the non-compliant (p < 0.0001); this illustrates a further decreasing trend compared with the 6th month in the compliant group as opposed to an increasing tendency in the non-compliant group, respectively (difference: 9.0%, 95% CI: 0% to 20.6%, p = 0.1523). At that time, 80% of the failed treatments were HPV mRNA positive compared to 10% positivity for the cases treated successfully (OR: 34, 95%CI: 6.8–173, p < 0.0001), a finding indicative that HPV mRNA E6 & E7 positivity accurately predicts treatment failure; p16 positivity was also observed at higher rates in cases with treatment failure. Conclusions: Consistent condom use following conservative excisional CIN treatment appears to significantly reduce rates of CIN recurrence and biomarkers of HPV expression. Additional HPV vaccination at the time of treatment could further enhance the positive effect of consistent condom use. © 2022 by the authors

Alterations of HPV-related biomarkers after prophylactic HPV vaccination. A prospective pilot observational study in Greek women

The objective of this study was to investigate the hypothesis that HPV vaccination administered in patients with low-grade (LG) cytology shortly after an initial colposcopic assessment could prospectively alter HPV-related biomarkers. This was a prospective pilot observational study involving women attending a colposcopy clinic for evaluation of abnormal LG cytology that were advised to undergo HPV vaccination and proceeded accordingly. These women were compared with a matched unvaccinated group. Women requiring cervical biopsies or CIN treatment were excluded. Intervention: A full three-dose HPV vaccination was undertaken with either the 2-valent or the 4-valent anti-HPV VLP vaccine. LBC samples were obtained prior and after the completion of the vaccination regimen and tested for HPV DNA genotyping (CLART-2 HPV test) and E6 and E7 mRNA (NASBA technique). Results: Alterations of HPV-related biomarkers at a colposcopy reassessment appointment 12 months later. Analysis: The p-values, relative risk (RR), absolute relative risk (ARR), number needed to treat (NNT) and 95% confidence intervals for each biomarker in each group were assessed. Results: A total of 309 women were included in the analysis. One hundred fifty-two women received the vaccine. HPV vaccination reduced in a statistically significant manner (p < 0.05) HPV DNA positivity rates for genotypes 16, 18, and 31, RR = 1.6 (95% CI: 1.1 to 2.3), RR = 1.7 (95% CI: 1.1 to 2.8), and RR = 1.8 (95% CI: 1.0 to 2.9), in women who only tested DNA-positive for HPV16, 18, and 31 genotypes, respectively, prior to vaccination. A less pronounced, statistically insignificant reduction was shown for women who tested positive for both HPV DNA and mRNA E6 and E7 expression for HPV16, 18, and 33 subtypes. Statistically significant reduction in HPV mRNA positivity was solely documented for genotype 31 (p = 0.0411). Conclusions: HPV vaccination appears to significantly affect the rates of HPV16, 18, and 31 DNA-positive infections in the population testing HPV DNA-positive for the aforementioned genotypes. The above findings deserve verification in larger cohorts. © 2020 by the authors. Licensee MDPI, Basel, Switzerland

High-risk human papillomavirus DNA test and p16INK4a in the triage of LSIL: A prospective diagnostic study

Objective. The detection of high-grade cervical intraepithelial neoplasia (CIN) amongst patients with low-grade cytology (LSIL) is challenging. This study evaluated the role of high-risk HPV (HR-HPV) DNA test and p16INK4a immunostaining in identifying women with LSIL cytology at risk of harboring CIN2 or worse (CIN2+) and the role of p16INK4a in the triage of a population of HR-HPV positive LSIL. Methods. We conducted a prospective study including women with LSIL cytology. Detection of HR-HPV was carried out by means of a polymerase chain reaction based assay. p16INK4a immunostaining was performed using the Dako CINtec cytology kit. All patients had colposcopically directed punch biopsies or large loop excision of the transformation zone of the cervix. The endpoint was detection of a biopsy-confirmed CIN2+. Results. A series of 126 women with LSIL cytology were included. HR-HPV test had sensitivity 75% and specificity 64% for an endpoint of CIN2+. p16INK4a had significantly higher specificity of 89% (p = 0.0000) but low sensitivity of 42%. The role of p16INK4a immunostaining in the triage of LSIL positive for HR-HPV was also evaluated. p16INK4a triage had 70% positive predictive value (PPV); however, this was not significantly higher than the PPV (56%) of HR-HPV test alone (p = 0.4). Conclusions. The results indicate that HR-HPV or p16INK4a cannot be used as solitary markers for the assessment of LSIL. The addition of p16INK4a immunostaining led to an increase in HR-HPV specificity; however, the biomarker needs to be assessed further to establish its role as an adjunct test in the triage of LSIL. © 2010 Elsevier Inc

Effect of Condom Use after CIN Treatment on Cervical HPV Biomarkers Positivity: Prolonged Follow Up Study

Background: Several factors contribute in the cervical healing process following local surgical treatment; in a previous work our group has documented a beneficial mid-term role of regular condom use immediately postoperatively in terms of CIN relapse prevention and expression of active viral biomarkers. Materials and Methods: Aiming to investigate whether the favorable contribution of consistent condom use could be extrapolated in the longer term, we conducted a prospective single center observational study including women scheduled to undergo conservative excisional treatment for CIN (LLETZ procedure). In all women a strong recommendation for consistent use for the first 6 months was given. For 204 women who underwent the procedure and completed successfully the two-year follow up a complete dataset of HPV biomarkers’ results obtained six months and two years postoperatively was available. Patients were asked to complete a questionnaire to assess condom use compliance. A 90% compliance rate represented the threshold for consistent use. An LBC sample was obtained and tested for HPV genotyping, E6 & E7 mRNA by NASBA technique as well as flow cytometry, and p16 at 0 (pre-treatment), 6 and 24 months. HPV DNA and other related biomarkers status at 6 and 24 months, treatment failures at 24 months and condom use compliance rates represented study outcomes. Results: Six months post-operatively we documented a reduction in the rates of HPV DNA positivity, which was detected in only 23.2% of compliant condom users in comparison to 61.9% in the non-compliant group (p < 0.001, OR: 0.19, 95%CI: 0.1–0.36). For the HPV mRNA test, either assessed with the NASBA method or with flow cytometry, reduced positivity percentages were observed in the compliant group, in particular 1.6% vs. 8% for NASBA and 7.1% vs. 16.4% using flow cytometry, although these differences were not statistically significant (p = 0.1039 and 0.0791, respectively). Finally, reduced p16 positivity rates were documented in the compliant group. At the two year follow up, a more pronounced difference in HPV DNA positivity rates was observed, specifically only 13% positivity among the compliant women compared with 71% of the non-compliant (p < 0.0001); this illustrates a further decreasing trend compared with the 6th month in the compliant group as opposed to an increasing tendency in the non-compliant group, respectively (difference: 9.0%, 95% CI: 0% to 20.6%, p = 0.1523). At that time, 80% of the failed treatments were HPV mRNA positive compared to 10% positivity for the cases treated successfully (OR: 34, 95%CI: 6.8–173, p < 0.0001), a finding indicative that HPV mRNA E6 & E7 positivity accurately predicts treatment failure; p16 positivity was also observed at higher rates in cases with treatment failure. Conclusions: Consistent condom use following conservative excisional CIN treatment appears to significantly reduce rates of CIN recurrence and biomarkers of HPV expression. Additional HPV vaccination at the time of treatment could further enhance the positive effect of consistent condom use

Alterations in human papillomavirus-related biomarkers after treatment of cervical intraepithelial neoplasia

Objective: This study aims to assess the alterations in various HPV-related biomarkers 6 months post-treatment and how these relate to various risk factors and individual characteristics; their role for the prediction of treatment failure was also evaluated. Material and methods: Design: Prospective observational study. Population: Women planning to undergo treatment for cervical intraepithelial neoplasia. Intervention: A liquid-based cytology sample was taken pre-operatively. This was tested for HPV genotyping, Nucleic Acid Sequence Based Amplification, flow cytometric evaluation and p16 immunostaining. A repeat LBC sample was obtained 6 months post-treatment and was tested for the same biomarkers. Outcomes: The alterations of the biomarkers 6 months post-treatment were recorded. Their relation to individual characteristics and risk factors (age, smoking, sexual history, use of condom, CIN grade, excision margin status, crypt involvement) as well as their role for the prediction of residual/recurrent disease were assessed. Analysis: The accuracy parameters (sensitivity, specificity, positive and negative predictive value and the likelihood ratios) of each biomarker for the prediction of recurrent/residual CIN were calculated. Results: A total of 190 women were recruited. All biomarkers had significantly higher negativity rates post-treatment compared to pre-treatment ones. Multivariate analysis demonstrated that consistent condom use post-treatment significantly reduces the high-risk HPV positivity rates in comparison to no use (OR = 0.18; 95% CI: 0.09-0.38). Sensitivity and specificity for all high risk HPV DNA testing were 0.5/0.62, respectively; the relevant values for only type 16 or 18 DNA typing were 0.5/0.92, for NASBA 0.5/0.94, for flow 0.5/0.85 and for p16 0.25/0.93. Conclusion: CIN treatment reduces positivity for all HPV-related biomarkers. Consistent condom use significantly reduces high-risk HPV positivity rates. More cases of treatment failures are required in order to specify whether different combinations of HPV-related biomarkers could enhance the accuracy of follow up, possibly in the form of a Scoring System that could allow tailored post-treatment surveillance

Classification of cervical cytology for human papilloma virus (HPV) infection using biospectroscopy and variable selection techniques

Cervical cancer is the second most common cancer in women worldwide. We set out to determine whether attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy combined with principal component analysis-linear discriminant analysis (PCA-LDA) or, variable selection techniques employing successive projection algorithm or genetic algorithm (GA) could classify cervical cytology according to human papilloma virus (HPV) infection [high-risk (hr) vs. low-risk (lr)]. Histopathological categories for squamous intraepithelial lesion (SIL) were segregated into grades (low-grade vs. high-grade) of cervical intraepithelial neoplasia (CIN) expressing different HPV infection (16/18, 31/35 or HPV Others). Risk assessment for HPV infection was investigated using age (≤29 years vs. >30 years) as the distinguishing factor. Liquid-based cytology (LBC) samples (n = 350) were collected and interrogated employing ATR-FTIR spectroscopy. Accuracy test results including sensitivity and specificity were determined. Sensitivity in hrHPV category was high (≈87%) using a GA-LDA model with 28 wavenumbers. Sensitivity and specificity results for >30 years for HPV, using 28 wavenumbers by GA-LDA, were 70% and 67%, respectively. For normal cervical cytology, accuracy results for ≤29 years and >30 years were high (up to 81%) using a GA-LDA model with 27 variables. For the low-grade cervical cytology dataset, 83% specificity for ≤29 years was achieved using a GA-LDA model with 33 wavenumbers. HPV16/18 vs. HPV31/35 vs. HPV Others were segregated with 85% sensitivity employing a GA-LDA model with 33 wavenumbers. We show that ATR-FTIR spectroscopy of cervical cytology combined with variable selection techniques is a powerful tool for HPV classification, which would have important implications for the triaging of patients. © The Royal Society of Chemistry 2014

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. © 2015 Abraham Pouliakis et al

The effect of a structured individualized educational intervention on breastfeeding rates in Greek women

Breastfeeding rates remain extremely low in Greece and women with gestational diabetes mellitus and hypothyroidism may experience additional difficulties. The aim of the study was to investigate the effect of a structured individualized lactation educational intervention by a midwife on increasing breastfeeding rates in women with endocrine disorders and low‐risk women compared to women receiving standard care, 24 months after delivery. Two‐hundred women made up the study population. Half of them were experiencing endocrine pregnancy disorders and 100 women constituted the low‐risk pregnancy standard care control group. Women who were breast-feeding exclusively were significantly higher in the midwifery intervention group with endocrine disorders, namely breastfeeding continued at four months (breastfeeding: 20% vs. 12%, exclusive breastfeeding: 50% vs. 26%, p = 0.0228), and at six months after childbirth (breastfeeding: 54% vs. 28%, exclusive breastfeeding: 32% vs. 12%, p = 0.0011), compared to the standard care control group with endocrine disorder. The low‐risk midwifery intervention group breastfed at four months (22% vs. 14%, p = 0.0428) and at six months (52% vs. 26%, p = 0.0018) at higher rates compared to the standard care control group. In addition, exclusive breastfeeding was significantly higher in the low‐risk midwifery intervention group at four months (46% vs. 20%, p = 0.0102) and six months (38% vs. 4%, p < 0.0001) compared to the standard care control group. This study was the first at-tempt of a structured midwifery breastfeeding education in Greece and its major contribution reflects a significant positive impact on breastfeeding rates in terms of duration and exclusivity in women with gestational endocrine disorders as well as in low‐risk women, and could possibly be applied and instituted in everyday clinical practice to increase the low breastfeeding rates in Greece. © 2021 by the authors. Licensee MDPI, Basel, Switzerland