Triage by PAX1 and ZNF582 methylation in women with cervical intraepithelial neoplasia grade 3: a multicenter case-control study

Background The colposcopy-conization inconsistency is common in women with cervical intraepithelial neoplasia grade 3 (CIN3). No adequate method has been reported to identify the final pathology of conization. In this study, we explored the ability of PAX1 and ZNF582 methylation to predict the pathological outcome of conization in advance. Methods This was a multicenter study and included 277 histologically confirmed CIN3 women who underwent cold knife conization (CKC) from January 2019 to December 2020. The methylation levels of PAX1 (PAX1m) and ZNF582 (ZNF582 m) were determined by quantitative methylation specific PCR (qMSP) and expressed in ΔCp. Receiver-operating characteristic (ROC) curves were used to evaluate predictive accuracy. Results The final pathological results in 48 (17.33%) patients were inflammation or low-grade squamous intraepithelial lesion (LSIL), 190 (68.59%) were high grade squamous intraepithelial lesion (HSIL) and 39 (14.08%) were squamous cervical cancer (SCC). PAX1 m and ZNF582 m increased as lesions progressed from inflammation/LSIL, HSIL to SCC. PAX1 and ZNF582 methylation yielded better prediction performance compared to common screening strategies, whether individually or combined. ΔCpZNF582 ≥19.18). A 6.53-fold increase in SCC risk was observed in patients with elevated ZNF582 methylation (ΔCpZNF582 < 7.09). Conclusion DNA Methylation would be an alternative screening method to triage and predict the final outcome of conization of the CIN3 cases.

Diagnostic markers of human papillomavirus persistence and progression of cervical intraepithelial neoplasia

Cervical intraepithelial neoplasia is widespread among patients of the reproductive period, the peak of the incidence occurs at a young age, often when the reproductive function has not yet been performed. The oncological alertness of the medical community is justified: the incidence of cervical cancer has increased by 24.6% over the past ten years, and in the structure of cancer mortality in the age group of 30–39 years cervical cancer occupies the 1st place accounting for 23%. Тhe same time aggressive management of the disease in patients who have not yet performed their reproductive function is associated with the loss of significant volume of cervical tissues and an increase in the frequency of premature birth. The most acute issue is the tactics of managing cervical neoplasia of the second degree of severity, when there is still hope for the regression of the disease, but at the same time there is a high risk of transition to dysplasia of the third degree of severity and subsequently to invasive cancer. The search for diagnostic markers that allow predicting the probability of regression and, conversely, the unfavorable course of the disease is the most important task solution of which will allow optimizing tactics in controversial clinical cases and avoiding unnecessary surgical aggression, improving the perinatal outcomes of patients undergoing surgical treatment. The search for new economically feasible and diagnostically valuable markers allowing us to predict the course of the disease and determine the amount of necessary surgical treatment continues, but the most significant at this moment is still the determination of the coexpression of p16 and Ki-67 in immunocytochemical studies and p16INK4a in immunohistochemical studies. The diagnostic sensitivity of determining the co-expression of p16INK4a/Ki67 is 98.5%, and the expression of p16INK4a in CIN II indicates that in 81% of cases there is a tendency to a progressive course of the disease.

High-Grade Cervical Intraepithelial Neoplasia (CIN) Associates with Increased Proliferation and Attenuated Immune Signaling

Implementation of high-risk human papilloma virus (HPV) screening and the increasing proportion of HPV vaccinated women in the screening program will reduce the percentage of HPV positive women with oncogenic potential. In search of more specific markers to identify women with high risk of cancer development, we used RNA sequencing to compare the transcriptomic immune-profile of 13 lesions with cervical intraepithelial neoplasia grade 3 (CIN3) or adenocarcinoma in situ (AIS) and 14 normal biopsies from women with detected HPV infections. In CIN3/AIS lesions as compared to normal tissue, 27 differential expressed genes were identified. Transcriptomic analysis revealed significantly higher expression of a number of genes related to proliferation, (CDKN2A, MELK, CDK1, MKI67, CCNB2, BUB1, FOXM1, CDKN3), but significantly lower expression of genes related to a favorable immune response (NCAM1, ARG1, CD160, IL18, CX3CL1). Compared to the RNA sequencing results, good correlation was achieved with relative quantitative PCR analysis for NCAM1 and CDKN2A. Quantification of NCAM1 positive cells with immunohistochemistry showed epithelial reduction of NCAM1 in CIN3/AIS lesions. In conclusion, NCAM1 and CDKN2A are two promising candidates to distinguish whether women are at high risk of developing cervical cancer and in need of frequent follow-up.

Ultrasonic Elastography Combined with Human Papilloma Virus Detection Based on Intelligent Denoising Algorithm in Diagnosis of Cervical Intraepithelial Neoplasia

The aim of this research was to study the application of ultrasonic elastography combined with human papilloma virus (HPV) detection based on bilateral filter intelligent denoising algorithm in the diagnosis of cervical intraepithelial neoplasia (CIN) and provide a theoretical basis for clinical diagnosis and treatment of CIN. In this study, 100 patients with cervical lesions were selected as research objects and randomly divided into control group and experimental group, with 50 cases in each group. Patients in control group and experimental group were diagnosed by ultrasonic elastography combined with HPV detection. The experimental group used the optimized image map of bilateral filter intelligent denoising algorithm for denoising and optimization, while the control group did not use optimization, and the differences between them were analyzed and compared. The diagnostic effects of the two groups were compared. As a result, the three accuracy rates of the experimental group were 95%, 95%, and 98%, respectively; the three sensitivity rates were 96%, 92%, and 94%, respectively; and the three specificity rates were 99%, 97%, and 98%, respectively. In the control group, the three accuracy rates were 84%, 86%, and 84%, respectively; the three sensitivity rates were 88%, 84%, and 86%, respectively; and the three specificity rates were 81%, 83%, and 88%, respectively. The accuracy, sensitivity, and specificity of experiment group were significantly higher than those of control group, and the difference was statistically significant ( P < 0.05 ). In summary, the bilateral filter intelligent denoising algorithm has a good denoising effect on the ultrasonic elastography. The ultrasonic image processed by the algorithm combined with HPV detection has a better diagnosis of CIN.

Increased Expression of TIGIT And KLRG1 Correlates With Impaired CD56Bright NK Cell Immunity In HPV16-Rrelated Cervical Intraepithelial Neoplasia

Background: The onset and progression of cervical intraepithelial neoplasia (CIN) are closely associated with the persistent infection of high-risk HPV (especially type16), which is mainly caused by immune escape. Natural killer (NK) cells play an important role against virally infected cells and tumor cells through a fine balance of signals from multiple surface receptors. Overexpression of non-MHC-I specific inhibitory receptors TIGIT, KLRG1, Siglec-7, LAIR-1, and CD300a on NK cells correlates with cellular exhaustion and immune evasion, but these receptors have not been investigated in CIN. The aim of the present study was to examine the potential role of NK cell non-MHC-I specific inhibitory receptors expression in immune escape from HPV16-related CIN patients .Methods: the subset distribution, IFN-γ and TNF-α expression levels and immunophenotype of TIGIT, KLRG1, Siglec-7, LAIR-1, and CD300a of NK cells were investigated in peripheral blood mononuclear cell samples by flow cytometry from 82 women, including HPV16(+) subjects with CIN 0, CIN I, and CIN II-III, and HPV subjects with CIN 0 (control). immunohistochemistry was applied to detect the expression of ligands for NK receptors in the cervical tissues. HPV types were identified by PCR assays.Results: The HPV16(+) subjects with high-grade lesions had an increased number of circulating peripheral blood CD56bright NK cells with reduced functionality and IFN-γ secretion. The expression levels of the inhibitory molecules TIGIT and KLRG1 on CD56bright NK cells increased in parallel with increasing CIN grade. In addition, TIGIT and KLRG1 related ligands, PVR, N-Cadherin and E-Cadherin expression level was also elevated with increasing CIN grade. Conclusions: Our results suggested that the reduced viability of CD56bright NK cells and increased TIGIT and KLRG1 expression represent an escape mechanism associated to persistent HPV16 infection and CIN progression. These results may facilitate the development of early-warning immune predictors and therapeutic strategies for HPV16-associated CIN based on the TIGIT and KLRG1 inhibitory pathways of NK cells.