Atividade antic??ncer da Biflorina em c??lulas tumorais g??stricas

• Autor(es): Barbosa, Gleyce dos Santos
• Data: 2016
• Origem: Oasisbr
• Assunto(s): C??ncer g??strico; Plantas medicinais; Biflorina; Gastric cancer; CI??NCIAS DA SA??DE: FARM??CIA
• Tipo de Documento: Dissertação de mestrado
• Idioma: Português

Descrição

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CAPES - Coordena????o de Aperfei??oamento de Pessoal de N??vel Superior

In the northern Brazil, especially in the states of Amazonas and Par??, there is a high incidence of gastric cancer which available treatments are ineffective in most cases. The biflorin, a prenylated ortanaftoquinona obtained from the roots of Capraria biflora L., showed, in previous studies, an inhibition of the tumor growth in vivo and in vitro in several cell lines, without inducing mutagenicity. The ACP02, main cell line used in this work, was established from a primary diffuse gastric adenocarcinoma, removed from the stomach cardia region of a 66-year-old pacient born in Par??. Among the main karyotypes and genetic changes of this lineage are the trisomy of chromosome 8, with amplification of the MYC oncogene, and the deletion of the chromosome 17???s short arm, where the tumor suppressor gene TP53 is located. These characteristics observed in the established line correspond to those observed in the original tumor, what indicates that this line is a good alternative to the study of the human gastric cancer pathophysiology and the drug screening. Considering biflorin???s promising potencial and the necessity to regionalize the study of new anticancer drugs to answer genotype particularities of each group of patients, we evaluated the activity of biflorin on ACP02 through the morphological cell analysis, the cell viability assays, the clonogenic assay, scratch assay, the cell differentiation test by NBT, the evaluation of the MYC status and telomere length by FISH, in 0, 1.0, 2.5 and 5.0 ???M concentrations, after 24 and 72 hours of treatment. The morphological changes indicate cell death by necrosis, and suggest the occurrence of the differentiation process, due to change in the format of the attached cell. The biflorin showed a cytotoxic activity (IC50 1.92 ???M) and cytostatic, anticlonogenic and antimotility activities, statistically significant (p<0,05) since 1 ???M. There was a significant reduction in MYC amplification rate in cells treated with 2.5 e 5 ???M of the drug (p<0,05), which can explain the occurrence of cellular differentiation process, in view of the physiological role of this gene. This differentiation was confirmed by the results obtained during the NBT test, in which the cells showed the ability to metabolize salt from a treatment of 1.0 ???M. The telomere length was reduced in cells treated with 5.0 ???M of biflorina (p<0,05). These results showed that biflorin acts on important targets of anticancer therapy, when used on the gastric line ACP02, at concentrations around 2.5 ???M, which makes it a promising substance for the treatment of this tumor type.

No norte do Brasil, sobretudo nos Estados do Amazonas e Par??, h?? alta incid??ncia de c??ncer g??strico, cujos tratamentos dispon??veis s??o ineficazes, na maioria dos casos. A biflorina, uma ortonaftoquinona prenilada obtida das ra??zes de Capraria biflora L., demonstrou, em trabalhos pr??vios, inibir o crescimento tumoral in vivo e in vitro de v??rias linhagens celulares, sem induzir mutagenicidade. A ACP02, principal linhagem celular utilizada nesse trabalho, foi estabelecida a partir de um adenocarcinoma g??strico difuso prim??rio, retirado da regi??o c??rdia estomacal de um paciente de 66 anos oriundo do Par??. Entre as principais altera????es cariot??picas e gen??ticas dessa linhagem est??o a trissomia do cromossomo 8, com amplifica????o do oncogene MYC, e a dele????o do bra??o curto do cromossomo 17, onde estaria localizado o gene supressor de tumor TP53. Essas caracter??sticas observadas na linhagem estabelecida correspondem ??quelas observadas no tumor que a deu origem, evidenciando que tal linhagem ?? uma boa alternativa para o estudo da fisiopatologia do c??ncer g??strico humano e triagem de drogas. Tendo em vista o potencial promissor da biflorina e a necessidade de regionalizar o estudo de novas drogas antic??ncer para atender as particularidades genot??picas de cada grupo de pacientes, avaliamos a atividade da biflorina sobre ACP02 atrav??s da an??lise morfol??gica das c??lulas, dos ensaios de viabilidade celular, do ensaio clonog??nico, do teste de motilidade, do teste de diferencia????o celular pelo NBT e da avalia????o do status de MYC e comprimento dos tel??meros por FISH, nas concentra????es de 0, 1,0, 2,5 e 5 ???M, ap??s 24 e 72h de tratamento. As altera????es morfol??gicas indicaram morte celular por necrose, al??m de sugerirem a ocorr??ncia de processo de diferencia????o, devido ?? altera????o no formato da c??lula aderida. A biflorina apresentou atividade citot??xica (CI50 1,92 ???M) e atividades citost??tica, anticlonog??nica e inibidora da motilidade in vitro, estatisticamente significativas (p<0,05) desde a concentra????o de 1 ???M. Houve redu????o significativa da taxa de amplifica????o de MYC, nas c??lulas tratadas com 2,5 e 5,0 ???M da droga (p<0,05), o que pode justificar a ocorr??ncia de processo de diferencia????o celular, tendo em vista o papel fisiol??gico desse gene. Essa diferencia????o foi confirmada pelos resultados obtidos durante o teste do NBT, no qual as c??lulas apresentaram capacidade de metabolizar o sal a partir do tratamento de 1,0 ???M. O comprimento do tel??mero foi reduzido nas c??lulas tratadas com 5,0 ???M de biflorina (p<0,05). Esses resultados demonstram que a biflorina atua sobre importantes alvos da terapia antic??ncer, quando utilizada sobre a linhagem g??strica ACP02 em concentra????es em torno de 2,5 ???M, o que a torna uma subst??ncia promissora para o tratamento desse tipo de tumor.