Estudo farmacológico pré-clínico de análogos de LASSBio-1586 candidatos a protótipos de fármacos anticâncer

• Autor(a) principal: Carneiro, Teiliane Rodrigues
• Data de Publicação: 2016
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da Universidade Federal do Ceará (UFC)
• Texto Completo: http://www.repositorio.ufc.br/handle/riufc/42639
• topic: Combretum caffrum; Combrestatatina A4; LASSBio-1586; Antineoplásicos

In Brazil, the estimate for the biennium 2016-2017 indicates the occurrence of 600.000 new cases of cancer. Combretastatin A4 (CA-4), found in the bark of Combretum caffrum, is the compound with simpler chemical structure known to exert potent cytotoxic activity, through reversible interaction with the colchicine site of β-tubulin. In recent years, it has been studied and pointed with great potential anticancer. Thus, this study aims to evaluate the cytotoxic effect of analogous compounds of LASSBio-1586, a promising anticancer prototype, CA-4 analogue, study possible mechanisms involved and to evaluate some pharmacokinetic parameters in cancer cell lines with and without overexpression of tyrosine protein kinases. Using as a selection criterion the selectivity for tumor type and/or cytotoxic potency was selected compounds LASSBio-1918 and LASSBio-1920 and cell lines of colorectal cancer (HCT-116) without overexpression of tyrosine kinases and of non-small cell lung cancer overexpressing EGFR L858R mutation to (PC-9). The results of cytotoxicity obtained by the MTT technique showed that, in incubation time of 48 hours, LASSBio-1918 had IC50 values of 15.81μM and 15.5μM while LASSBio-1920 had IC50 values of 2.7μM and 1.0μM, in HCT- 116 and PC-9, respectively. LASSBio-1918 showed selectivity index of 1.15 and 10.61, while LASSBio-1920 showed 2.3 and 2.1 index in HCT-116 and PC-9, respectively. In these cell lines, analysis of the cell cycle profile, membrane integrity, externalization of phosphatidylserine and expression of PARP protein, suggested that the cytotoxic effect of these compounds is associated with death by apoptosis, with sale cycle stop cell in G2 / M profile similar to the CA-4. The evaluation of the profile of inhibition of tubulin polymerization showed that LASSBio-1920, the compound with higher power cytotoxic, at a concentration of 10μM, inhibit the polymerization of tubulin, showing inhibition profile similar to LASSBio-1586 at 30μM, while that LASSBio-1918 presented profile to cause inhibition only in the concentration of 30μM. Importantly LASSBio-1920 did not show multitargeted profile, because it had no second target protein kinase EGFR (wild type). Regarding studies of permeability and stability, LASSBio-1920 was absorbed into bloodbrain artificial barrier (Permeability Coefficient = 11:48) and artificial barrier of the gastrointestinal tract (Absorbed Fraction = 99.7%). This compound was also stable in medium buffered to pH7.4 and rat plasma and was significantly metabolized only in the microsomal fraction in the presence of NADPH-generating system (cofactor), indicating the action of FMO (flavin-containing monooxygenase) and / or cytochromes P450. The studies of molecular anchoring suggested that LASSBio-1586 and LASSBio-1920 have interaction modes similar to colchicine site on the β-tubulin protein and as expected, the non-polar interactions in the lipophilic pocket of colchicine binding site are important, appear to be essential to explain the higher potency of cytotoxic LASSBio-1920 compared to LASSBio- 1586. Thus, we can conclude that the LASSBio-1920 compound is a lead candidate to be optimized and other tests should be performed for a determination of their antitumor activity in vivo model as well as for the determination of toxicological parameters and evaluation of additional mechanism of action , targeting EGFR mutated.