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Synthesis of GPI anchor analogues to support the discovery of new molecular targets of Trypanosoma cruzi

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Ana Luisa Malaco Morotti
Total Authors: 1
Document type: Doctoral Thesis
Press: Ribeirão Preto.
Institution: Universidade de São Paulo (USP). Faculdade de Ciências Farmacêuticas de Ribeirão Preto (PCARP/BC)
Defense date:
Examining board members:
Ivone Carvalho; Marcelo Dias Baruffi; Lídia Moreira Lima; Kleber Thiago de Oliveira
Advisor: Ivone Carvalho; Robert Andrew Field

Glycosylphosphatidylinositol (GPI) anchors are essential molecules to attach glycoconjugates and proteins in protozoan\'s cell surface. Trypanosoma cruzi produces a range of unique GPI structures that anchor mucins and trans-sialidases which participate in important processes involved in the interaction between parasite and host. As an effort to study T. cruzi GPI anchor biosynthesis and possibly use it as a potential target for an antichagasic drug, this work aims to synthesize GPI anchor analogs (labelled or not) and analyze the potential of these molecules as substrates in the GPI biosynthetic pathway. In this context, a pseudo-disaccharide 31 was synthesized by O-glycosylation reaction between azide glycosyl donors (32 or 33a-d) and myo-inositol acceptor (34), prepared from glucosamine (35) hydrochloride and methyl ?-D-glucopyranoside (36), respectively, using orthogonal protection/ deprotection. Five different glycosyl donors (32 and 33a-d) were prepared to investigate the influence of their protective groups on the stereoselectivity of the O-glycosylation reaction in the presence of different solvents to afford the required GPI ?-linkage. In addition, the synthesis of the myo-inositol acceptor 34 was achieved using several protection/deprotection steps, besides the Ferrier rearrangement, to form a functionalized cyclitol derivative that enables the regioselective introduction of the azide glycoside unit and phospholipid moiety on its C-1 and C-6 positions, respectively. Then, O-glycosylation of acceptor 34 with donor 33c was accomplished in diethyl ether, using TMSOTf as promoter to give exclusively ?-anomer 31c in high yield. After deallylation of 31c, the phosphodiester moiety bearing an octyl chain (87), prepared by the H-phosphonate approach, was appended to the pseudo-disaccharide to yield, after deprotection, target compounds 30a. The same synthetic strategy was applied to the preparation of 30c, even though in the protective form, compound 91 bearing an alkyl-naphthyl side chain (90). Currently, compound 30a is being tested as substrates of GPI anchor biosynthesis in Euglena gracilis cell membranes, a non-pathogenic unicellular algae, which may potentially be used as a model for phylogenetically related human parasites. After incubation of the potential GPI substrate 30a with E. gracilis microsomal membranes for generation of metabolites, the analysis by LC-MS and, eventually, isolation of the products will be performed for further characterization. Products that show any substrate or inhibitory activities will be also assayed in T. cruzi microsomal membrane. (AU)

FAPESP's process: 14/21200-0 - Synthesis and evaluation of the potential inhibitory activity of carbohydrates that mimic GPI anchor of T. cruzi
Grantee:Ana Luísa Malaco Morotti
Support Opportunities: Scholarships in Brazil - Doctorate