Non-glycosidic analogues of alpha-galactosylceramide: Design, synthesis and biological activity
[eng] The Immune System (IS) is a complex collection of tissues, organs, cells and molecules to defense the body against threatens. IS is organized into two lines of defense: innate immunity which acts in a non-specific manner and adaptive immunity, able to release a specific response. Natural Kille...
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2018 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:diposit.ub.edu:2445/123925 |
| Acceso en línea: | https://hdl.handle.net/2445/123925 http://hdl.handle.net/10803/587193 |
| Access Level: | acceso abierto |
| Palabra clave: | Sistema immunològic Cèl·lules immunocompetents Immune system Immunocompetent cells |
| Sumario: | [eng] The Immune System (IS) is a complex collection of tissues, organs, cells and molecules to defense the body against threatens. IS is organized into two lines of defense: innate immunity which acts in a non-specific manner and adaptive immunity, able to release a specific response. Natural Killer T cells (NKT) are a IS cell class that combine features of Natural Killer cells (innate immunity) and T cells (adaptive immunity), and are able to trigger IS to combat a threat (pro-inflammatory cytokines - Th1 response) as well as regulate it (anti-inflammatory cytokines – Th2 response). This versatility prompts NKT cells to be studied into different disease models, from type 1 diabetes to cancer. NKT cells present a characteristic T cell receptor (TCR), typically found in T lymphocytes, which recognize in a restricted manner glycolipid antigens (Ag) bound to CD1d protein in presenting cells. Although the endogenous ligand has not been yet identified, a glycolipid isolated from a marine sponge was identified is a powerful NKT cell activator. A synthetic analogue of this antigen is alpha-galactosylceramide (αGalCer) a glycosphingolipid with alpha- galactose O- linked to a phytoceramide skeleton with a C26 N-acyl chain. This ligand is the standard antigen and has a pivotal importance in NKT biology, but presents several drawbacks: NKT cell overstimulation leading to an unresponsive state, a metabolically unstable glycosidic bond and unselective NKT cell response (both Th1 and Th2 cytokines are simultaneously released after αGalCer NKT cell stimulation). During the present doctoral Thesis, new NKT stimulators with unprecedented structures were studied. Our idea consisted in the replacement of the αGalCer sugar by aromatic rings in the search of ground-breaking analogues. Computational tools were used to screen a virtual library of ligands into the ternary complex CD1d-Ag-TCR to design the new compounds. Although the protein system complexity and the large number of degrees of freedom of these molecules precluded to establish a precise computational model, Molecular Dynamics studies added to other chemical criteria and the precedent of one compound activity led to propose a small family of 20 compounds. The chemical synthesis of 16 of the proposed compounds was accomplished through different strategies involving aziridine ring opening reactions and aromatic nucleophilic substitution as key reactions. The desired ceramide-like compounds, having an aromatic ring in place of the sugar present in the glycolipid antigens, were obtained in moderate to good yields and high purity. A representative set of compounds was tested as NKT cell activators in both mice and human cell in vitro assays. In mice splenocytes, the aromatic compounds induced a total polarization towards Th1 responses, as no Th2 cytokines were detected. In this experiment, the aromatic analogues were tested 10 times more concentrated than standard glycolipid αGalCer and similar amounts of IFN- were released. When compounds were tested in purified human NKT cells, the aromatic compound were also very potent stimulators, giving responses polarized to Th1 profile although a slight production of Th2 cytokines was observed, in all cases notoriously lower than that generated by αGalCer. In human NKT cells, the new compounds tested at same dose than αGalCer gave a comparable amount of IFN-g (around 80%) but only around 20% of IL-4. From these experiments it was evident that this family of structurally new NKT stimulators has a high potency in vitro, being able to promote NKT cell cytokine responses in comparable levels to that induced by the standard ligand αGalCer with a much better selectivity towards Th1 response. |
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