RESUMO
The human fibroblast growth factor-2 (FGF-2) highly expressed in tumours is an important factor to promote tumour angiogenesis and lymphangiogenesis. A disulphide-stabilized diabody (ds-Diabody) could specifically target FGF-2 and show its advantages in inhibition of tumour angiogenesis and growth. It is very important for antibody drugs to confirm the fine epitope. Here, theoretical structure models of FGF-2 and antibody were built by homology modelling. The amino acid residues in the interaction interface of antigen and antibody were analysed by molecular docking. The potential epitope was predicted by homology modelling and molecular docking of antigen-antibody and site-directed mutation assays of alanine scanning. The predicted epitope was verified by antigen mutagenesis and enzyme-linked immunosorbent assay (ELISA). The epitope mapping assay showed that the epitope of ds-Diabody against FGF-2 was defined by the discontinuous sites including six amino acid residues (P23, Q65, R69, G70, Y82 and R118). The results showed that the epitope was localized in the interaction interface of FGF-2 and ds-Diabody. The fine epitope mapping provided the important information for understanding the inhibition activity of ds-Diabody against FGF-2 and helping in the further development of ds-Diabody against FGF-2 as a potentially promising antibody drug for future cancer therapy.
Assuntos
Anticorpos Biespecíficos/imunologia , Anticorpos Biespecíficos/farmacologia , Dissulfetos/química , Mapeamento de Epitopos , Fator 2 de Crescimento de Fibroblastos/antagonistas & inibidores , Fator 2 de Crescimento de Fibroblastos/imunologia , Neoplasias/tratamento farmacológico , Anticorpos Biespecíficos/química , Anticorpos Biespecíficos/uso terapêutico , Reações Antígeno-Anticorpo , Proliferação de Células/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Fator 2 de Crescimento de Fibroblastos/genética , Células HEK293 , Humanos , Modelos Moleculares , Mutagênese Sítio-Dirigida , Neoplasias/imunologia , Neoplasias/patologiaRESUMO
To automatically infer the patterns of vessel structure such as the distal ends, segments, bifurvessel structures, and crossing of two vessels in X-ray angiographic images, a novel method is presented based on Gabor filter and circle detector. The method can cope with varying vessel curvature and intensity feature occur along the longitudinal vessel direction. The present study can facilitate 2-D quantitative description of vessel tree and 3-D vessel reconstruction, and provide an elementary clue for the diagnostics. The proposed method has been successively applied to both synthetic images for validation purposes and the actual angiographic images, which yielded encouraging results.
