Ena Pešut

Application of mass cytometry in multiparametric characterization of precancerous cervical lesions

AbstractCervical cancer (CC) is the fourth most common malignant tumor in women worldwide. Detecting different biomarkers together on single cells by novel method mass cytometry could contribute to more precise screening. Liquid‐based cytology (LBC) cervical samples were collected (N = 53) from women categorized as normal and precancerous lesions. Human papillomavirus was genotyped by polymerase chain reaction, while simultaneous examination of the expression of 29 proteins was done by mass cytometry (CyTOF). Differences in cluster abundances were assessed with Spearman's rank correlation as well as high dimensional data analysis (t‐SNE, FlowSOM). Cytokeratin (ITGA6, Ck5, Ck10/13, Ck14, Ck7) expression patterns allowed determining the presence of different cells in the cervical epithelium. FlowSOM analysis enabled to phenotype cervical cells in five different metaclusters and find new markers that could be important in CC screening. The markers Ck18, Ck18, and CD63 (Metacluster 3) showed significantly increasing associated with severity of the precancerous lesions (Spearman rank correlation rho 0.304, p = 0.0271), while CD71, KLF4, LRIG1, E‐cadherin, Nanog and p53 (Metacluster 1) decreased with severity of the precancerous lesions (Spearman rank correlation rho −0.401, p = 0.0029). Other metaclusters did not show significant correlation, but metacluster 2 (Ck17, MCM, MMP7, CD29, E‐cadherin, Nanog, p53) showed higher abundance in low‐ and high‐grade intraepithelial lesion cases. CyTOF appears feasible and should be considered when examining novel biomarkers on cervical LBC samples. This study enabled us to characterize different cells in the cervical epithelium and find markers and populations that could distinguish precancerous lesions.

Monitoring HPV Prevalence and Risk Cofactors for Abnormal Cytology in the Post-Vaccination Period among Croatian Women

The incidence and mortality rate of cervical cancer in Croatia remains a health challenge despite screening efforts. Besides the persistent infection with HPV, the development of cancer is also associated with some cofactors. The goal of this study was to assess circulating HPV genotypes and risk factors for the development of cervical precancer after almost 16 years from the onset of HPV vaccination in Croatia. In this study, a total of 321 women attending gynecological care were evaluated. Relevant medical and demographic information, including cytology, were collected. HPV genotyping was performed by PCR. Comparing the HPV types found in circulation in the pre-vaccination (1999–2015) and post-vaccination periods (2020–2023), a statistically significant reduction in HPV 31 was noted, while the overall prevalence increased in the post-vaccination period. Besides the expected HPV positivity as a risk factor, the history of smoking was associated with LSIL or worse cytology at enrollment. For the first time, this population study revealed a statistically significant shift in the HPV genotype in the post-vaccination period, as well as the confirmation of risk factors for the development of abnormal cytology among Croatian women.