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Authors
Abstract(s)
AVALIAÇÃO DA EXPRESSÃO IMUNOHISTOQUÍMICA DE EGFR E KI-67 EM CARCINOMA DE CÉLULAS ESCAMOSAS ORAIS EM GATO - O carcinoma de células escamosas oral (CCEO) é uma neoplasia maligna de origem epitelial e é o tumor mais prevalente na cavidade oral do gato. Apesar da cirurgia ser o tratamento nuclear, nem sempre é curativo, existindo a necessidade de terapias neoadjuvantes e adjuvantes. O presente estudo teve como objetivo principal avaliar por técnica de imunohistoquímica (IHC) a expressão do Recetor do Fator de Crescimento Epidérmico (EGFR) e do marcador de proliferação Ki-67 em CCEO e averiguar a presença de associações entre variáveis clínicas, patológicas e imunohistoquímicas com o prognóstico. Foram incluídos 30 casos de CCEO cujos tumores se encontravam conservados em blocos de parafina. Para cada caso foi consultado o registo de variáveis clínicas e de prognóstico, nomeadamente a idade (<10 anos ou 10 anos), o sexo, a localização primária, o estadiamento TNM e o tempo de sobrevivência global (SG). As amostras tumorais foram processadas da seguinte forma: cortes de 3μm para coloração em hematoxilina-eosina (H&E) e para o estudo imunohistoquímico. Este incluiu uma recuperação antigénica (RA) (digestão enzimática com pepsina para o EGFR e citrato ph=6.0 para o Ki-67); incubação com o anticorpo primário (EGFR – Policlonal, Abcam; Ki-67 – Monoclonal, Dako) e utilização de polímero como sistema de deteção. A expressão de EGFR foi avaliada de forma cega por um patologista, através do sistema de graduação Herceptest. A expressão de Ki-67 foi avaliada de forma automática, definindo o índice deste marcador de forma percentual calculada a partir da contagem de pelo menos 1000 células. A mediana foi o valor de cut-off utilizado para diferenciar os tumores com baixa ou alta expressão de Ki-67. Foi utilizado um software comercial de análise estatística, onde foram calculados os testes de Qui-quadrado/Fisher para variáveis categóricas e o teste de Log-rank para comparação de curvas de sobrevivência de Kaplan-Meyer. Em relação à expressão de EGFR, 13,3% foram classificados com pontuação 0, todos os restantes casos mostraram positividade para o EGFR: 36,7% com pontuação 1+, 30% com pontuação 2+ e 20% com pontuação 3+. A marcação de EGFR não mostrou associação com as variáveis clínicas, o grau de diferenciação tumoral, ocorrência de recidiva, marcação de Ki-67 ou SG. Quanto aos valores do índice de Ki-67 variaram entre 3,85% e 71,62%, com uma média de 33,62% e mediana de 36,08%. A marcação de Ki-67 foi significativamente associada com o sexo. Não foi encontrada qualquer associação com as restantes variáveis estudadas. A análise de sobrevivência mostrou que os animais intervencionados viveram significativamente mais tempo (p=0.007), assim como os animais do sexo masculino (p=0.029). Foi possível observar uma tendência para uma sobrevivência maior em tumores com localização mandibular e baixa expressão de EGFR pontuação (0 e 1). Os resultados obtidos neste estudo são preliminares, mas clinicamente relevantes. A maioria dos tumores apresentava algum grau de expressão para o marcador EGFR, sendo que pontuações mais elevadas de imunomarcação se encontravam associados a um pior prognostico.
ABSTRACT - EVALUATION OF THE IMUNOHISTOCHEMISTRY EXPRESSION OF EGFR AND KI-67 IN FELINE ORAL SQUAMOUS CELL CARCINOMA - Oral squamous cell carcinoma (OSCC) is a malignant neoplasm of epithelial origin and is the most prevalent tumor in the oral cavity of the cat. Although surgery is considered curative treatment, associated neoadjuvant and adjuvant therapies are often required and there is a need to better understand the behavior and prognosis of this tumor. The main purpose of the present study was to evaluate by immunohistochemistry (IHC) the expression of the Epidermal Growth Factor Receptor (EGFR) and the proliferation marker Ki-67 in OSCC and to assess the presence of associations with prognosis. We included 30 cases of OSCC tumors that were preserved in paraffin blocks. Clinical and prognostic variables were recorded for each case, including age (<10 years or 10 years), gender, primary location, TNM staging, and overall survival time (OS). Tumor samples were processed as follows: 3μm sections for hematoxylin-eosin (H&E) staining and for immunohistochemical study. This included antigen retrieval (AR) (enzymatic digestion with pepsin for EGFR and citrate ph=6.0 for Ki-67); incubation with the primary antibody (EGFR - Polyclonal, Abcam; Ki-67 - Monoclonal, Dako) and use of the polymer as detection system. EGFR expression was evaluated blindly by a pathologist using the Herceptest grading system. Ki-67 expression was assessed automatically, defining the index of this marker in a percentage form calculated from the count of at least 1000 cells. The median was the cut-off value used to differentiate tumors with low or high Ki-67 expression. A commercial statistical analysis software was used, where the Chi-square/Fisher test for categorical variables and the Log-rank test for Kaplan-Meyer survival curve comparison were calculated. Regarding EGFR expression, 13.3% were classified with score 0, all remaining cases showed positivity for EGFR: 36.7% with score 1+, 30% with score 2+ and 20% with score 3+. EGFR staining showed no association with the clinical variables, the degree of tumor differentiation, occurrence of recurrence, Ki-67 staining and SG. As for the Ki-67 index values ranged from 3.85% to 71.62%, with a mean of 33.62% and median of 36.08%. Ki-67 labeling was significantly associated with gender. No association was found with the remaining clinical variables, tumor differentiation, the occurrence of recurrence, EGFR staining and SG. Survival analysis showed that the intervened animals lived significantly longer (p=0.007), and male animals as well (p=0.029). A trend towards higher survival could be observed in tumors with mandibular location and low EGFR score expression (0 and 1). The results obtained in this study are preliminary but clinically relevant. Most tumors had some degree of expression for EGFR, and higher score of immunolabeling were associated with a worse prognosis.
ABSTRACT - EVALUATION OF THE IMUNOHISTOCHEMISTRY EXPRESSION OF EGFR AND KI-67 IN FELINE ORAL SQUAMOUS CELL CARCINOMA - Oral squamous cell carcinoma (OSCC) is a malignant neoplasm of epithelial origin and is the most prevalent tumor in the oral cavity of the cat. Although surgery is considered curative treatment, associated neoadjuvant and adjuvant therapies are often required and there is a need to better understand the behavior and prognosis of this tumor. The main purpose of the present study was to evaluate by immunohistochemistry (IHC) the expression of the Epidermal Growth Factor Receptor (EGFR) and the proliferation marker Ki-67 in OSCC and to assess the presence of associations with prognosis. We included 30 cases of OSCC tumors that were preserved in paraffin blocks. Clinical and prognostic variables were recorded for each case, including age (<10 years or 10 years), gender, primary location, TNM staging, and overall survival time (OS). Tumor samples were processed as follows: 3μm sections for hematoxylin-eosin (H&E) staining and for immunohistochemical study. This included antigen retrieval (AR) (enzymatic digestion with pepsin for EGFR and citrate ph=6.0 for Ki-67); incubation with the primary antibody (EGFR - Polyclonal, Abcam; Ki-67 - Monoclonal, Dako) and use of the polymer as detection system. EGFR expression was evaluated blindly by a pathologist using the Herceptest grading system. Ki-67 expression was assessed automatically, defining the index of this marker in a percentage form calculated from the count of at least 1000 cells. The median was the cut-off value used to differentiate tumors with low or high Ki-67 expression. A commercial statistical analysis software was used, where the Chi-square/Fisher test for categorical variables and the Log-rank test for Kaplan-Meyer survival curve comparison were calculated. Regarding EGFR expression, 13.3% were classified with score 0, all remaining cases showed positivity for EGFR: 36.7% with score 1+, 30% with score 2+ and 20% with score 3+. EGFR staining showed no association with the clinical variables, the degree of tumor differentiation, occurrence of recurrence, Ki-67 staining and SG. As for the Ki-67 index values ranged from 3.85% to 71.62%, with a mean of 33.62% and median of 36.08%. Ki-67 labeling was significantly associated with gender. No association was found with the remaining clinical variables, tumor differentiation, the occurrence of recurrence, EGFR staining and SG. Survival analysis showed that the intervened animals lived significantly longer (p=0.007), and male animals as well (p=0.029). A trend towards higher survival could be observed in tumors with mandibular location and low EGFR score expression (0 and 1). The results obtained in this study are preliminary but clinically relevant. Most tumors had some degree of expression for EGFR, and higher score of immunolabeling were associated with a worse prognosis.
Description
Dissertação de Mestrado Integrado em Medicina Veterinária, na área científica de Clínica
Keywords
Carcinoma de células escamosas oral recetores ErbB Ki-67 Imunohistoquímica Cirurgia oncológica Squamous Cell Carcinoma of Head and Neck ErbB Receptors Ki-67 Antigen Immunohistochemistry Surgical Oncology
Pedagogical Context
Citation
Gouveia GPF. 2022. Avaliação da expressão imunohistoquímica de EGFR e KI-67 em carcinoma de células escamosas oral em gato [dissertação]. Lisboa: FMV, Universidade de Lisboa
Publisher
Universidade de Lisboa, Faculdade de Medicina Veterinária