Stratifin em Tumores de Cabeça e Pescoço: Um alvo molecular identificado por Phage Display / Stratifin in head and neck tumors: a molecular target identified by Phage Display

Nos últimos anos, houve um aumento na frequência dos casos de tumores de cabeça e pescoço apesar da diminuição do consumo do tabaco e álcool, e isso tem sido atribuído, em parte, à infecção pelo Papilomavírus Humano HPV. Por apresentar baixa sobrevida em 5 anos e ter alta morbidade, tem se buscado novos alvos moleculares para terapias combinadas. Nesse contexto nosso grupo identificou, através da tecnologia de Phage Display, uma sequência peptídica com interação preferencial por células tumorais com relação à células não transformadas, e ensaios adicionais identificaram seu alvo como sendo a proteína Stratifin. Stratifin tem sido reportado como um oncogene em diversos modelos tumorais, entretanto seu papel em carcinoma de células escamosas de cabeça e pescoço (CCECP) permanece desconhecido e poucos trabalhos na literatura reportam sua atividade em CCECP e/ou outro tumores relacionados ao HPV. Dessa forma, o objetivo desse trabalho foi explorar o potencial valor clínico e o papel biológico da Stratifin em CCECP. Dados do perfil de expressão e de metilação assim como dados clínicos foram extraídos em base de dados do The Cancer Genoma Atlas TCGA. Paralelamente, o perfil de expressão de Stratifin foi verificado através de ensaios de RT/qPCR e Western Blot em um painel de linhagens celulares de CCECP que contempla as principais características moleculares para esses tipos tumorais. A partir da observação de que todas as linhagens expressam Stratifin, utilizou-se a tecnologia de CRISPR/Cas9 para modular sua expressão (nocauteando ou superexpressando o gene) de modo a se observar parâmetros relacionados ao processo tumorigênico. Dessa forma, foi possivel verificar os efeitos da Stratifin em ensaios de proliferação, viabilidade após tratamentos com quimioterápicos, irradiação, crescimento livre de ancoragem e clonogenicidade. Como resultados, observamos que expressão aumentada de Stratifin no tecido tumoral quando comparado ao tecido normal, foi positivamente relacionada com o grau histológico, negatividade para HPV, mutação em TP53 e CDKN2A. Biologicamente, o nocaute de Stratifin foi relacionado com maior sensibilidade à quimioterápicos, menor capacidade de formação de colônias, e reduzida capacidade de crescimento livre de ancoragem. Esses resultados sugerem que Stratifin atue como um oncogene em CCECP, entretanto ensaios adicionais devem ser realizados para corroborar esse achados

Over recent years, there has been an increase of head and neck tumors frequency despite the decrease in tobacco and alcohol consumption, and this has been attributed, in part, to Human Papillomavirus infection. Due to its low 5-year survival and high morbidity, new molecular targets for combined therapies have been sought. In this context, our group identified, through Phage Display technology, a peptide sequence with preferential interaction by tumor cells in relation to non-transformed cells, and further assays identified its target as the Stratifin protein. Stratifin has been reported as an oncogene in several tumor models, however its role in head and neck squamous cell carcinoma (HNSCC) remains unknown and few works in the literature report its activity in HNSCC and/or other HPV-related tumors. Therefore, the aim of this study was to explore the potential clinical value and biological role of Stratifin in HNSCC. Expression profile data as well as clinical data were extracted from The Cancer Genome Atlas - TCGA database. In parallel, the expression profile of Stratifin was verified through RT/qPCR and Western Blot assays in a panel of HNSCC cell lines that address the main molecular characteristics for these tumor types. Since all cell lines express Stratifin, CRISPR/Cas9 technology was used to modulate its expression (gene knocking out or overexpressing) in order to check parameters related to the tumorigenic process. Thus, it was possible to verify the Stratifin effects in proliferation assays, viability after chemotherapy treatments, irradiation, anchorage-free growth and clonogenicity. As a result, we observed an increased expression of Stratifin in tumor tissue when compared to normal tissue, which was positively related to histological grade, HPV negativity, mutation in TP53 and CDKN2A. Biologically, knockout of Stratifin was associated with greater sensitivity to chemotherapy, less colony-forming capacity, and reduced anchorage-free growth capacity. These results suggest that Stratifin acts as an oncogene in HNSCC, however additional assays should be performed to corroborate these findings

HPV-related external genital lesions among men residing in Brazil

Abstract The aims of this study were to determine the incidence of external genital lesions (EGLs), specifically histologically confirmed condyloma (genital warts) and Penile Intraepithelial Neoplasia (PeIN), and genital HPV infection progression to EGLs among healthy men aged 18-73 residing in Brazil. Subjects included 1118 men enrolled in the HPV Infection in Men (HIM) study between July 2005 and June 2009. At each visit, EGLs were biopsied and subjected to pathological evaluation. HPV status in genital swabs and biopsies was determined by Linear Array and INNO-LiPA, respectively. Age-specific EGLs incidence and the proportion and median time to EGL development were estimated. Kaplan-Meier cumulative incidence rates at 6, 12, and 24 months were determined. During follow-up, 73 men developed an incident EGL. Men could develop multiple EGLs and there were 36 men with condyloma, 27 men with lesions suggestive of condyloma, six men with PeIN, and 20 men with non-HPV lesions. HPV-positive men who developed EGLs were younger (p = 0.002) than men that did not develop lesions. Among the 815 men with HPV infection, 4% progressed to EGL with the same HPV detected in the swab. During follow up, 15.7% of genital HPV-6 and HPV-11 infections progressed to condyloma (median progression time of nine months for HPV-6 versus 6.8 months for HPV-11). Approximately 1% of HPV-16 infections progressed to PeIN with a median progression time of 25 months. HPV types covered by the 4-valent HPV vaccine were detected in 82.3% and 83.3% of condyloma and PeIN, respectively. The high burden of HPV and high frequency of progression to disease underscores the need to offer HPV prophylactic vaccination to men to reduce the overall burden of infection and diseases caused by HPV.