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The aim of the study was to evaluate the performance of the HPV-HR test to detect high-risk human papillomavirus (HPV) in urine samples in comparison with a commercial molecular HPV test., The aim of the study was to evaluate the performance of the HPV-HR test to detect high-risk human papillomavirus (HPV) in urine samples in comparison with a commercial molecular HPV test. Materials and Methods This is a prospective study, in which 350 patients diagnosed previously with cervical intraepithelial neoplasia (CIN) grade 2 or higher were enrolled. Urine and cervical specimens were collected. Urine was tested with the HPV-HR test and cervical specimens were tested with the Cobas. Results Of the 336 evaluable patients, there were 271 cases of CIN 2+, of which 202 were CIN 3+ and the remaining 65 patients were less than CIN 2. Positivity was 77.1% (95% confidence interval [CI] = 72.5–81.5) for the urine samples and 83.6% (95% CI = 79.6–87.6) for the cervical samples. Agreement between cervical and urine samples for HPV detection was 79.8% (κ = 0.363; 95% CI = 0.243–0.484). Sensitivity for CIN 2+ was 83.4% (95% CI = 78.4–87.6) for urine and 90.8% (95% CI = 86.7–92.9) for cervical samples. The sensitivity for CIN 3+ was 85.6% (95% CI = 80.0–90.2) for urine and 92.6% (95% CI = 88.0–95.8) for cervical samples. Specificity for worse than CIN 2 was 50.8% (95% CI = 33.7–59.0) and 46.2% (95% CI = 33.7–59.0) for urine and cervical samples, respectively. Conclusions Although these results demonstrated slightly higher detection rates for HR-HPV and clinical sensitivity in cervical samples than in urine, when compared with histological diagnoses, urine sampling is a viable alternative to access women who do not participate in routine screening programs., (undefined), info:eu-repo/semantics/publishedVersion

The present study aimed to ascertain the significance of topoisomerase II alpha (TOP2A) and minichromosome maintenance protein (MCM) 2 expression in anal carcinoma. A total of 75 anal lesions were retrieved from the files of the Department of Pathology of Barretos Cancer Hospital (Barretos, Brazil) in order to verify the human papillomavirus (HPV) statuses of these lesions and characterize the immunohistochemical expression levels of TOP2A and MCM2 in anal carcinoma, as these are important markers for cervical HPV-induced lesions; their expression was also compared with respect to p16 and Ki-67. The vast majority of the cases tested positive for HPV16 (84%); 1 case tested positive for both HPV16 and HPV18. Positive HPV16 status was more frequent in early stages than in advanced stages (P=0.008). Positive immunohistochemical reactivity for MCM2 and TOP2A protein was observed in 71.6 and 100% of cases, respectively. Positive reactivity for p16 was significantly associated (P=0.001) with histological grade, and was more commonly expressed in squamous cell carcinoma than adenocarcinomas. HPV16 was strongly associated with positive p16 protein expression (76.6%). However, the high expression of Ki-67 combined with the high expression of p16 was predominantly observed in Stage III-IV cases. MCM2, TOP2A, p16 and Ki-67 exhibited intense positive staining in the anal lesions, indicating that these markers were significantly and constantly expressed in anal carcinoma., info:eu-repo/semantics/publishedVersion

Persistent human papillomavirus (HPV) infection is a key event in HPV-induced carcinogenesis. As part of the prospective Finnish Family HPV Study, we analysed the physical state and viral copy numbers of HPV16 in asymptomatic oral infections that either persisted or cleared during the 6-year follow-up. The persister group comprised 14 women and 7 men with 51 and 21 HPV16-positive brush samples. The clearance group included 41 women and 13 men, with 64 and 24 samples, respectively. Physical state and viral DNA load were assessed by using quantitative PCR for HPV16 E2 and E6 genes. E2/E6 ratio was calculated and HPV16 was classified as episomal, mixed or integrated with values of 0.93-1.08, The Finnish Family HPV Study has been supported by the Academy of Finland (no. 116438/2006, no. 130204/2008), Finnish Cancer Foundation, Solberg Foundation, Finnish Dental Society Apollonia and the Government Special Foundation (EVO) to Turku University Hospital. The funders had no role in the study design and data analysis, decision to publish or preparation of the manuscript

Adriana T. Lorenzi was supported (via a scholarship) by CNPq process no. 573799/2008-3 and FAPESP process no. 2008/57889-1 through the National HPV Research and Technology Institute of Sao Paulo, Brazil (INCT-HPV)

There has been an increase in the incidence of anal cancer in the past two decades, with squamous cell carcinoma (SCC) being the most frequent histological type identified. Among the risk factors, high-risk human papillomavirus (HPV) infection is the most pervasive. Raf kinase inhibitor protein (RKIP) is expressed in a number of normal human tissues and previous studies have demonstrated the prognostic value of the loss of RKIP expression in several gastrointestinal tumors. Therefore, the present study aimed to evaluate the clinical implications of RKIP expression in a series of neoplastic lesions of the anal canal. The resected tumors of 48 patients [8 high-grade intraepithelial lesions (HSILs), 14 adenocarcinomas and 26 squamous cell carcinomas (SCCs)] were immunohistochemically evaluated for RKIP expression, and the results were correlated with clinicopathological data. The results identified a decreased 5-year overall survival rate in patients with adenocarcinoma (40.8%) compared with patients with SCC (76.7%), and a decreased 5-year disease-free survival rate in patients at clinical stages III/IV (37.3 vs. 62.5 and 82.6% for clinical stages 0 and I/II, respectively). Low RKIP expression was revealed in 62.5% of HSILs, 88.5% of SCCs and 100.0% of the adenocarcinomas. High RKIP expression was associated with patient ethnicity (37.5% in non-Caucasians vs. 7.5% in Caucasians) and patient age (33.3% in younger patients vs. 0.0% in older patients). Finally, high RKIP expression was correlated with HPV16 infection status (40% in HPV- vs. 5.3% in HPV+ patients). A correlation was identified between high RKIP expression and lesions with a generally improved prognosis, such as those diagnosed in younger patients, in situ lesions and lesions of lower clinical grades; there was also a negative correlation between high RKIP expression and HPV16 positivity in patients., São Paulo Research Foundation (grant nos. 2010/16795-4 and 2011/08523-7) and Ministry of Science, Technology, Innovation and Communication (grant no. MCT/FINEP/CT-INFRA-PROINFRA 01/2011), info:eu-repo/semantics/publishedVersion

The Role of Self-Collection by Vaginal Lavage for the Detection of HPV and High-Grade Intraepithelial Neoplasia, Objective: To compare the results of cervical cytology and high-risk HPV tests using samples obtained using two different collection modalities in a population of Brazilian women: self-collection (vaginal lavage) and cervical Pap testing. Methods: We enrolled 204 women who were aged 18-64 years and had previously obtained abnormal cervical cytology test results; 83.8% of them agreed to participate. The sample was divided into two aliquots: one for the cytological study and one for the molecular analysis of high-risk HPV. Results: Fifty-eight percent of the participants preferred to utilize self-collection as an alternative screening method. However, we noticed that the HPV positivity rate was significantly lower in self-collected samples when compared to those obtained using the conventional collection method (p = 0.035). The cytology tests of the samples obtained via self-collection were sensitive and had a positive predictive value and an area under the curve (AUC) that were significantly lower than those of the Pap test. However, the specificity and negative predictive value of these tests were similar. When compared with the HPV test, the self-collected samples demonstrated lower accuracy in predicting high-grade cervical intraepithelial neoplasia or worse, with a significantly lower sensitivity, positive predictive value, and AUC than the cervical Pap test samples. Conclusion: Self-collection by vaginal lavage is simple and well accepted by women. Due to its limitations, however, self-collection by lavage should be utilized with caution., info:eu-repo/semantics/publishedVersion

Background: Incorporation of HPV tests into cervical cancer screening programs may be advantageous over conventional cytology, especially in developing nations, where the largest burden of cervical cancer is observed. Objectives: To conduct an evaluation of commercially available molecular HPV tests in Brazilian women. Study design: Two groups were recruited: group A was composed of 511 women referred to the clinics because of a previous abnormal Pap test while group B consisted of 2464 subjects under routine screening. Cervical samples were collected using SurePath liquid cytology (LBC) device, and split into aliquots which were submitted to molecular testing by Hybrid Capture and cobas HPV. Colposcopy and biopsies were made according to the standard guidelines, directed by cytological diagnosis. Results: Prevalence of HSIL was 5.97% and 0.7% in Group A and B respectively. High-Risk HPV DNA was found in about 9% of group B women, while in group A this frequency was 24%. Having CIN3+ as the study end-point, the negative predictive values for molecular methods were above 99.8%. All “in-situ” and invasive cervical carcinomas were detected by both HPV nucleic acid assays. Conclusion: Use of HPV DNA testing was feasible and highly sensitive in cancer screening settings of Brazil.

Genotyping HPV from samples tested positive to careHPV™ assay in rural and remote areas of Brazilian territory. Objectives Genotyping HPV from samples tested positive to careHPV™ assay in rural and remote areas of Brazilian territory. Methods A total of 5079 women were enrolled in an opportunistic screening from the Barretos Cancer Hospital, through mobile units or ambulatory unit. All careHPV™ hr-HPV positive samples were tested by a Luminex-based protocol in order to evaluate the HPV infecting types. Results Positive hr-HPV results were obtained in 10.6% (536/5068) of women. Among these cases, HPV-56 and HPV-51 were the most common types detected in 32.3% and 31.4%, respectively. HPV-53 (20.5%), HPV-18 (18.5%), HPV-58 (17.6%), HPV-52 (16.0%) and HPV-16.6%) were the other most frequent types detected. These frequencies represent prevalences of 2.35%, 2.12%, 2.02%, 1.84% and 1.80% respectively, within the population studied. Regarding low-risk HPVs, HPV-6 was detected in 12.9% of the samples. The less frequent types (, National Institute of Research and Technology in HPV (INCT - HPV) of São Paulo, through the CNPq process nº 573799/2008-3, FAPESP process nº 2008/57889-1, through and CNPq process nº 203348/2014-1.

Anal cancer is a rare type of digestive tract disease, which has had a crescent incidence in a number of regions. Carcinomas are most frequently found, with squamous cell carcinoma (SCC) comprising similar to 95% of all anal tumors. The major risk factor for development of this type of tumor is human papillomavirus (HPV) infection. However, previous studies have identified patients with anal cancer that are HPV-/p16-and observed that they have a poorer outcome compared with HPV+/p16+ patients. This suggests that molecular profile may drive anal cancer progression. The aim of the present study was to evaluate the mutational status of two important oncogenes, KRAS and BRAF, in a series of anal cancer lesions. Resected tumors of the anal canal (n=43) were evaluated, nine of these were high-grade squamous intra-epithelial lesion cases (HSIL), 11 were adenocarcinomas, and 23 SCCs. Direct sequencing of KRAS proto-oncogene, GTPase (KRAS; codons 12 and 13) and B-Raf proto-oncogene, serine/threonine kinase (BRAF; codon 600) was performed and associated with patient clinicopathological and molecular features. There was a trend of poorer prognosis of adenocarcinoma compared with HSIL and SCC. Analysis indicated one SCC patient (2.3%) exhibited a KRAS p.G13D mutation, and one adenocarcinoma patient (2.3%) exhibited a BRAF p.V600E mutation. It was observed that, these mutations are rare in anal tumors, and certain patients may be at a disadvantage using targeted therapies based on KRAS and BRAF mutational status. As there is a low mutation percentage in SCCs, adenocarcinomas and HSIL, there may exist other underlying molecular alterations that result in anal cancer development, which require further elucidation., The present study was partially supported by the São Paulo Research Foundation (grant no. 2010/16795-4 to Dr Adhemar Longatto-Filho) and the Ministério da Ciência e Tecnologia/Financiadora de Estudos e Projetos (grant no. CT-INFRA-PROINFRA 01/2011). Dr Lucas Tadeu Bidinotto received a São Paulo Research Foundation fellow-ship (grant no. 2011/08523-7)., info:eu-repo/semantics/publishedVersion

Objective To evaluate the reproducibility and accuracy of the HPV16/18-E6 test. Methods The study population was comprised of 448 women with a previously abnormal Pap who were referred to the Barretos Cancer Hospital (Brazil) for diagnosis and treatment. Two cervical samples were collected immediately before colposcopy, one for the hr-HPV-DNA test and cytology and the other for the HPV16/18-E6 test using high-affinity monoclonal antibodies (mAb). Women with a histologic diagnosis of cervical intraepithelial neoplasia grade 2 or 3 were considered to be positive cases. Different strategies using a combination of screening methods (HPV-DNA) and triage tests (cytology and HPV16/18-E6) were also examined and compared. Results The HPV16/18-E6 test exhibited a lower positivity rate compared with the HPV-DNA test (19.0% vs. 29.3%, p, Cancer Prevention Department, Center for the Researcher Support and Pathology Department of the Barretos Cancer Hospital. This study was supported by CNPq 573799/2008-3 and FAPESP 2008/57889-1, info:eu-repo/semantics/publishedVersion

This review tackles the issues related to disease burden caused by cervical cancer (CC) and its precursor (CIN) lesions in Brazil. A special focus is given to new technologies with potential to interfere with the development of CC by reducing the high-risk human papillomavirus (hr-HPV)-induced lesions that remain a major public health burden in all developing countries where organized screening programs do not exist. Globally, 85 % of all incident CC and 50 % of CC deaths occur in the developing countries. Unfortunately, most regions of Brazil still demonstrate high mortality rates, ranking CC as the second most common cancer among Brazilian women. Recently, CC screening programs have been tailored in the country to enable early detection of CC precursor lesions and thereby reduce cancer mortality. A combination of HPV testing with liquid-based cytology (LBC) seems to be a promising new approach in CC screening, with high expectation to offer an adequate control of CC burden in this country.

High-risk human papillomavirus (hrHPV) is an essential cause of cervical carcinoma and is also strongly related to anal cancer development. The hrHPV E6 oncoprotein plays a major role in carcinogenesis. We aimed to evaluate the frequency of hrHPV DNA and E6 oncoprotein in the anuses of women with cervical carcinoma. We analyzed 117 women with cervical cancer and 103 controls for hrHPV and the E6 oncogene. Positive test results for a cervical carcinoma included 66.7 % with hrHPV-16 and 7.7 % with hrHPV-18. One case tested positive for both HPV variants (0.9 %). The samples from the anal canal were positive for HPV-16 in 59.8 % of the cases. Simultaneous presence of HPV in the cervix and anal canal was found in 53.8 % of the cases. Regarding expression of E6 RNA, positivity for HPV-16 in the anal canal was found in 21.2 % of the cases, positivity for HPV-16 in the cervix was found in 75.0 %, and positivity for HPV-18 in the cervix was found in 1.9 %. E6 expression in both the cervix and anal canal was found in 19.2 % of the cases. In the controls, 1 % tested positive for HPV-16 and 0 % for HPV-18. Anal samples from the controls showed a hrHPV frequency of 4.9 % (only HPV16). The presence of hrHPV in the anal canal of women with cervical cancer was detected at a high frequency. We also detected E6 RNA expression in the anal canal of women with cervical cancer, suggesting that these women are at risk for anal hrHPV infection., We acknowledge the Research Support Fund of the State of São Paulo (FAPESP), process number 2010/16795–4), for the financial support grants. FAPESP had no other type of influence on this manuscript beyond the financial support

Cervical cancer is the second most common cancer among Brazilian women. High-risk human papillomavirus (hr-HPV) persistence is the primary cause of cervical neoplasia. Early detection of hr-HPV is important for identifying women at risk for developing cervical lesions. Approximately 85% of new cases of cervical cancer worldwide and 50% of the total cervical cancer deaths occurred in developing countries. Here, a new methodology to support a cervical cancer screening program was evaluated in women from various Brazilian regions. METHODS: Two thousand women aged 18-77years were enrolled in an opportunistic cervical cancer screening program and were randomized into self-vaginal or health professional-guided cervical sampling groups. The Qiagen careHPV™ test was performed on all samples. Pap tests were performed on all women using liquid-based cytology. RESULTS: Positive hr-HPV results were obtained in 12.3% (245/2000) of women; similar rates were observed in self- or health professional-collected samples. Eighty-nine percent (1719/2000) of cervical cytologies classified as normal were negative to hr-HPV. Among the cytological samples, 36.6% classified as ASC-US+ were positive to hr-HPV, 78.8% were LSIL and 75.0% were HSIL. CONCLUSIONS: Self-sampled and health professional-sampled vaginal/cervical specimens did not differ in their rates of detection of hr-HPV. Therefore, HPV DNA testing in self-sampled vaginal cells is an alternative to primary screening in low-resource settings., The authors thank the following: Cancer Prevention Department Team, Cleyton Zanardo de Oliveira and Allini Mafra of the Researcher Support Team and the Pathology Depai Intent of the Barretos Cancer Hospital. Rui Manuel Reis and Andre Lopes Carvalho from the Molecular Oncology Center; Jose Eduardo Levi from Sao Paulo University; Cintia B. Oliveira, Raphael Haikel junior and Edmundo Mauad from Barretos Cancer Hospital. Luisa Lina Villa was supported by a grant from CNPq and FAPESP (INCT-HPV). Study Supported by CNPq - Process no 573799/2008-3 and FAPESP no 2008/57889-1. They also thank all volunteer women who participated of this study.

Foram estudados os efeitos da adição de plantas medicinais de princípios cicatrizantes (Centelha asiática, Papaína e Confrei) na ração controle de caracóis terrestres, para se avaliar a interferência destas plantas na composição do muco glicoprotéico. Foram utilizados 80 caracóis terrestres Achatina sp, baseados em um peso homogêneo (49 e 40 g e idade média de 10 e 19 meses para Achatina fulica e Achatina monochromatica, respectivamente). Os animais foram distribuídos aleatoriamente em oito grupos experimentais: controle Achatina fulica (FC) e Achatina monochromatica (MC), centelha asiática Achatina fulica (FCe) e Achatina monochromatica (MCe), papaína Achatina fulica (FPa) e Achatina monochromatica (MPa) e confrei Achatina fulica (FCo) e Achatina monochromatica (MCo). Água e ração foram fornecidos ad libitum. Ao final de 150 dias de tratamento, os animais foram submetidos à técnica de extração do muco glicoprotéico, por meio do estímulo manual da glândula podal, responsável pela secreção deste muco. Esta metodologia considerou o bem-estar dos animais, uma vez que os mesmos não foram sacrificados e retornaram ao seu sistema de criação. Os mucos foram analisados por meio de testes colorimétricos e espectroscópicos, que constataram alterações semelhantes, porém apresentaram variação significativa em sua composição glicoprotéica. The effects of adding medicinal plants with healing properties (Centelha asiatica, Papaína and Confrei) in the control diet of land snails were studied to evaluate the effect of these plants on the mucus glicoproteic composition. Eighty Achatina sp snails, based on a homogeneous weight (49 and 40 g and averaging 10 and 19 months of age for Achatina fulica and Achatina monochromatica, respectively). The animals were randomly allotted to eight experimental groups: Achatina fulica (FC) and Achatina monochromatica (MC) control, Asian Achatina fulica (FCe) and Achatina monochromatica (MCe) centelha, Achatina fulica (FPa) and Achatina monochromatica (MPa) papaína and Achatina fulica (FCo) and Achatina monochromatica (MCo) confrei. Ration and water were fed ad libitum. At the end of 150 days of treatment, the animals were submitted to the technique for extraction of mucus glicoproteic by manual stimulation of podal gland, responsible for the secretion of this mucus. This methodology considered the well-being of the animals since they were not slaughtered but returned to their creation system. The mucus was analyzed by means of color and spectroscopic tests, that indicated similar changes, but showed significant variation in their glicoproteic composition.

Os caracóis terrestres pertencem às famílias Achatinidae (África) e Helicidae (Europa). A espécie mais conhecida é a Achatina fulica (Gigante africano), sendo a mais recomendada para as regiões tropicais e subtropicais devido a sua capacidade de adaptação a estes climas. O muco de caracóis terrestres Achatina sp tem sido pesquisado devido sua atividade cicatrizante, além da atividade antibacteriana. A suplementação (plantas com finalidades cicatrizantes definidas como: confrei, papaína e centelha asiática nas rações destes animais) teve o propósito de caracterizar a composição glicoproteica do muco, considerando o bem-estar do animal, haja vista que os mesmos não foram sacrificados. Fez-se uso de uma metodologia envolvendo a coleta através de estímulo manual da glândula podal, responsável pela secreção do muco. Metodologia esta divergente de alguns autores que fizeram uso de estímulo elétrico com corrente elétrica nos animais para a coleta do muco, método este que vai contra os propósitos de bem-estar animal. Este estudo, além de utilizar uma metodologia menos drástica e prejudicial aos animais, permitiu que os mesmos retornassem ao seu ambiente de criação. As análises realizadas por testes colorimétricos e espectroscópicos, constataram que as alterações apresentadas nos testes foram muito semelhantes; no entanto, mostraram uma variação significativa na composição glicoproteica dos mucos analisados. The terrestrial snails belong to the families Achatinidae (Africa) and Helicidae (Europe). The most known species is the Achatina fulica (Giant African), being the most recommended for the tropical and subtropical regions due its capacity of adaptation to these climates. Mucus of terrestrial snails Achatina sp has been searched had its wound healing activity, beyond the antibacterial activity. The suplementation (plants with defined wound healing purposes as comfrey, papain and centella asiatic in the rations of these animals) had the intention to characterize the glycoprotein composition of mucus, considering well-being of the animal, since the same ones had not been sacrificed. Using a methodology holding the collection through manual stimulation of the podal, responsible gland for the secretion of mucus. This methodology is divergent of some authors who had used electric stimulation with electric current on the animals for the collection of mucus, method contrary to the intentions of animal well-being. This study, beyond using a less drastic and harmful methodology to the animals, allowing that the same ones returned to its own creation environment. The analyses carried through for color and spectroscopy tests, evidenced that the alterations presented in the tests had been very similar; however, showed a significant variation in the glycoproteic composition of the analysed mucus.