We aimed to characterize miR-125b and miR-34a expression in 114 women with different cervical lesions: normal epithelium with (n= 20) and without (n= 29) HPV infection; LSIL (n= 28); HSIL (n= 29); and ICC (n= 8). miRNA expression analysis was performed by comparing the distinct groups with the reference group (women with normal epithelium without HPV). For miR-125b, we observed a twofold (2-ΔΔCt= 2.11;P= 0.038) increased expression among women with normal epithelium with HPV infection and a trend of downregulation in different cervical lesions including an 80% reduction (2-ΔΔCt= 0.21;P= 0.004) in ICC. Similarly, miR-34a expression analysis revealed an increased expression (2-ΔΔCt= 1.69;P= 0.049) among women with normal cervix and HPV infection, and despite no significant correlation with cervical lesions, its expression was increased by twofold (2-ΔΔCt= 2.08;P= 0.042) in ICC. Moreover, miR-125b levels were able to predict invasive cancers with 88% sensitivity and 69% specificity. Results showed that while miR-34a expression seems to be correlated with invasive cervical cancer, miR-125b expression is significantly changed within the different cervical lesions and their levels should be further investigated as possible predictive/prognostic biomarkers using a noninvasive approach.

Cervical cancer prevention is based on primary prevention with vaccines against Human Papillomavirus (HPV) and secondary prevention by screening with High‐Risk‐HPV (Hr‐HPV) detection. Since 2017, cervical cancer screening in women aged 25−60 years has been performed in Portugal using Hr‐HPV detection, followed by cytology in Hr‐HPV‐positive cases. Herein we report the prevalence of Hr‐HPV genotypes and cytological abnormalities among 462 401 women (mean age: 43.73 ± 10.79; median age: 45; range: 24−66 years) that participated in the Regional Cervical Cancer Screening Program of the Northern Region of Portugal, performed between August 2016 and December 2021. Overall, we describe a prevalence rate of 12.50% for Hr‐HPV varying from 20.76% at age 25% to 8.32% at age 64. The five most common Hr‐HPV genotypes identified were HPV‐68 (16.09%), HPV‐31 (15.30%), HPV‐51 (12.96%), HPV‐16 (11.06%), and HPV‐39 (11.01%). The prevalence of Hr‐HPV included in the nonavalent vaccine (HPV‐9valent) was 55.00% ranging from 47.78% to 59.18% across different age groups. Considering positive Hr‐HPV cases, 65.68% had a Negative for Intraepithelial Lesion or Malignancy (NILM) cytology, 20.83% atypical squamous cells of undetermined significance (ASC‐US), 8.85% Low‐Grade Squamous Intraepithelial Lesion (LSIL), 1.65% High‐Grade Squamous Intraepithelial Lesion (HSIL), 2.85% ASC‐H, 0.09% Atypical Glandular Cells, 0.02% Adenocarcinomas, and 0.02% Squamous Cell Carcinoma (SCC). Our analysis revealed that HPV‐9val genotypes were responsible for 52.13% NILM, 59.21% ASC‐US, 55.06% LSIL, 90.14% HSIL, 83.50% ASC‐H, and 100.00% SCC. Furthermore, multiple Hr‐HPV infections (risk ratio [RR] = 1.46; 95% confidence interval [CI] 1.34−1.58), HPV‐16/18 (RR = 5.16; 95% CI 4.75−5.93), or HPV‐9val genotypes (RR = 5.23; 95% CI 4.68−5.85) were associated with a significant risk of developing > HSIL ( p < 0.001). To date, this is the largest study on Hr‐HPV genotyping in cervical cancer screening that includes data from a complete cycle of the screening program. Our findings suggest a high prevalence of HPV‐9valent genotypes and a significant association with an increased risk of developing > HSIL. This constitutes important data for health authorities, which may help define the future of vaccination and cervical cancer screening strategies.

Ovarian cancer (OC) patients develop ascites, an accumulation of ascitic fluid in the peritoneal cavity anda sign of tumour dissemination within the peritoneal cavity. This body fluid is under-researched, mainly regarding the ascites formed during tumour progression that have no diagnostic value and, therefore, are discarded. We performed a discovery proteomics study to identify new biomarkers in the ascites supernatant of OC patients. In this preliminary study, we analyzed a small amount of OC ascites to highlight the importance of not discarding such biological material during treatment, which could be valuable for OC management. Our findings reveal that OC malignant ascitic fluid (MAF) displays a proliferative environment that promotes the growth of OC cells that shift the metabolic pathway using alternative sources of nutrients, such as the cholesterol pathway. Also, OC ascites drained from patients during treatment showed an immunosuppressive environment, with up-regulation of proteins from the signaling pathways of IL-4 and IL-13 and down-regulation from the MHC-II. This preliminary study pinpointed a new protein (Transmembrane Protein 132A) in the OC context that deserves to be better explored in a more extensive cohort of patients’ samples. The proteomic profile of MAF from OC patients provides a unique insight into the metabolic kinetics of cancer cells during disease progression, and this information can be used to develop more effective treatment strategies.

Bladder cancer (BlCa) is a common malignancy with significant morbidity and mortality. Current diagnostic methods are invasive and costly, showing the need for newer biomarkers. Although several epigenetic-based biomarkers have been proposed, their ability to discriminate BlCa from common benign conditions of the urinary tract, especially inflammatory diseases, has not been adequately explored. Herein, we sought to determine whether VIM_me and miR663a_me might accurately discriminate those two conditions, using a multiplex test. Performance of VIM_me and miR663a_me in tissue samples and urines in testing set confirmed previous results (96.3% sensitivity, 88.2% specificity, area under de curve (AUC) 0.98 and 92.6% sensitivity, 75% specificity, AUC 0.83, respectively). In the validation sets, VIM_me-miR663a_me multiplex test in urine discriminated BlCa patients from healthy donors or patients with inflammatory conditions, with 87% sensitivity, 86% specificity and 80% sensitivity, 75% specificity, respectively. Furthermore, positive likelihood ratio (LR) of 2.41 and negative LR of 0.21 were also disclosed. Compared to urinary cytology, VIM_me-miR663a_me multiplex panel correctly detected 87% of the analysed cases, whereas cytology only forecasted 41%. Furthermore, high miR663a_me independently predicted worse clinical outcome, especially in patients with invasive BlCa. We concluded that the implementation of this panel might better stratify patients for confirmatory, invasive examinations, ultimately improving the cost-effectiveness of BlCa diagnosis and management. Moreover, miR663a_me analysis might provide relevant information for patient monitoring, identifying patients at higher risk for cancer progression.

Cervical cancer screening remains an essential preventive tool worldwide. First line high-risk Human Papillomavirus (HrHPV) genotyping became gold standard for cervical cancer screening, and has been adopted by several countries, including Portugal. Herein, we aimed to assess the early outcomes of the regional Cervical Cancer Screening Program of Northern Portugal. The analysis of a representative set of cases evaluated during a one-month period (January 2020), with adequate follow-up was performed. Descriptive analysis was performed. Overall, 7278 samples were received, of which 15.2% were HrHPV positive, most of these disclosing a negative result in subsequent liquid-based cytology. Nearly half of the HrHPV-positive women were referred to colposcopy. Within this group, HPV16/18⁺ cases depicted the higher frequency of high-grade squamous intraepithelial lesion (HSIL) or worse, compared with abnormal cytology or persistent HrHPV infection. Among women with non-HPV16/18 HrHPV infection and negative cytology, which are eligible for repeat sampling in one year, 65% were re-tested. Importantly, nearly half of these cleared HrHPV infection. Furthermore, referral to colposcopy due to HPV16/18 infection and/or abnormal cytology results were associated with > 40% risk for HSIL or worse lesion. Our study confirmed the reliability and effectiveness of first line HrHPV genotyping in the Cervical Cancer Screening Program of Northern Portugal. Nonetheless, it also raised concerns about excessive referral to colposcopy, with the inherent human and financial costs. Thus, further improvement and optimization are key to ensure the sustainability of the program.