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O microambiente tumoral como alvo terapêutico: avaliação do efeito de antagonistas do receptor de bradicinina tipo 1 em melanoma murino / Tumor microenvironment as therapeutic target: evaluation of the effect of bradkinin receptor 1 antagonists in murine melanoma
São Paulo; s.n; 2009. 182 p. ilus, tab.
Thesis in Portuguese | LILACS, Inca | ID: lil-553370
RESUMO
O microambiente tumoral tem sido considerado um importante alvo para terapias anticâncer. Recentemente, o receptor de bradicinina tipo 1 (BKR1), o qual é expresso em leucócitos e células endoteliais, foi implicado na progressão de diversos cânceres, incluindo o melanoma... Dois diferentes métodos foram utilizados na seleção de genes diferencialmente expressos (differentially expressed genes -DEGs) o test “t de Student” e o teste Limma. Um total de 87 DEGs foram selecionadas com fold change >1,5 e valor de p<0,05. Alguns desses genes, envolvidos em importantes processos como angiogênese, invasão, metástase e quimioresistência, têm sido validados por PCR em tempo real, como transglutaminase-2, HSP-1b, neuropilina-2, serpine 1, fator de crescimento do tecido conjuntivo (Ctgf) e serglicina. Em um segundo conjunto de experimentos foi avaliado o efeito de antagonistas de BKR1 em alterações de permeabilidade vascular e distribuição de droga fluorescente (doxorrubicina) no microambiente do tumor. Os antagonistas de BKR1, R-954 e R-715, causaram redução da penetração de doxorrubicina no microambiente tumoral após tratamento por 2 horas, precedido ou não de tratamento crônico (diário), entretanto após 6 horas de tratamento foi observado um aumento da quantidade de doxorrubicina, comparado ao controle, seguido de redução após 24 horas de tratamento. Já em tumores EMT6 (sarcoma mamário murino) após 2 horas de tratamento com esses antagonistas foi possível observar um aumento da quantidade de doxorrubicina no microambiente tumoral. Esses resultados sugerem mecanismos pelos quais as complexas interações entre as células tumorais e as células do hospedeiro podem resultar na resistência tumoral aos quimioterápicos e a consequente falha dos tratamentos convencionais.
ABSTRACT
Tumor microenvironment has been considered an important target for anticancer therapies. Recently, the bradykinin receptor type 1 (BKR1), which is expressed on leukocytes and endothelial cells, has been implicated in the progression of several cancers, including melanoma. In this work we have shown that there is a delay in BKR1 knockout mice melanoma engraftment and we also observed higher survival rates in these animals compared to wild type mice. We have investigated the effects of selectives BKR1 antagonists evaluating its combination with the chemotherapeutic agent dacarbazine (DTIC). B16-F10 melanoma cells were injected subcutaneously in C57bl/6 mice, which were treated with DTIC (2mg/kg) every 3 days, R-954 (1mg/kg) daily, DTIC plus R-954 or PBS, as control. Mice were sacrificed at day 12 of treatment; tumors were excised and processed for RNA extraction. Treatments with R-954, DTIC or R-954 plus DTIC did not reduced tumor mass. However, the combined treatment increased the global survival of mice bearing tumor. Microarray experiments were performed using GeneChip Mouse 430 (Affymetrix). Data was normalized using the RMA method. Two different methods were employed for selection of differentially expressed genes (DEGs)the Student's T-test and the Limma package. A total of 87 DEGs were identified with fold change>1,5 and p value<0,05. Some of these genes, involved in important process as angiogenesis, invasion, metastasis and chemoresistance, had been validated by real time PCR as; transglutaminase-2, HSP-1b, neuropilin-2, serpine 1, connective tissue growth factor and serglycin. In a second set of experiments we evaluated the effect of antagonists BKR1 in vascular permeability alterations and distribution of fluorescent drug (doxorubicin) in tumor microenvironment. BKR1 antagonists, R-954 and R-715, caused a reduction of the penetration of doxorubicin in the tumor microenvironment after treatment for 2 hours, followed or not by chronic treatment (daily), but after 6 hours of treatment was observed an increase in the amount of doxorubicin, compared to the control, followed by reduction after 24 hours of treatment. However, in EMT6 tumors (murine mammary sarcoma) after 2 hours of treatment with these antagonists we observed an increased amount of doxorubicin in tumor microenvironment. These results suggest mechanisms by which the complex interactions between tumor and BKR1 positive host cells may result in tumor resistance to chemotherapeutic agents and the consequent failure of regular treatments.
Subject(s)
Full text: Available Index: LILACS (Americas) Main subject: Receptors, Bradykinin / Dacarbazine / Drug Therapy / Melanoma / Mice Type of study: Prognostic study Limits: Animals Language: Portuguese Year: 2009 Type: Thesis

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Full text: Available Index: LILACS (Americas) Main subject: Receptors, Bradykinin / Dacarbazine / Drug Therapy / Melanoma / Mice Type of study: Prognostic study Limits: Animals Language: Portuguese Year: 2009 Type: Thesis