2016 Fiscal Year Annual Research Report
脂質と抗原提示タンパク質の相互作用の解明を目指した結合分子の合成と生物活性評価
| Project/Area Number |
16J01933
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| Research Institution | Osaka University |
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Principal Investigator |
HOSSAIN MD. IMRAN 大阪大学, 理学研究科, 特別研究員(DC2)
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| Project Period (FY) |
2016-04-22 – 2018-03-31
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| Keywords | GalCer, / Glycosylation / KRN7000 / CD1d / Cytokines / Natural productcs |
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| Outline of Annual Research Achievements |
Purpose of this study is determination of the precise binding modes of the acyl chain of KRN7000 at A’ pocket of its binding protein CD1d.
Achievements: (1) Synthesis of a novel series of KRN7000 analogues has completed. Se atom is incorporated at 12C and 18C in place of methylene and Br atom introduced at 26C in place of methyl group in the acyl chain. (2) The binding of CD1d with the synthesized ligands was confirmed from the immunostimulatory activities of iNKT cells using murine spleen cells. All of the compounds induced high level of cytokines, IFN-γ and IL-4, productions. The analogues generated similar amount of IFN-γ compared to that of KRN7000, but showed high selectivity (IFN-γ/IL-4 = 2.9-5) towards this cytokine.
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| Current Status of Research Progress |
Current Status of Research Progress
3: Progress in research has been slightly delayed.
Reason
Synthesis of heavy atom (Se at 12 C and 18C) and halogens (F, Cl, Br at 26C) acyl chain containing KRN7000 analogues has completed. The cytokine production activity of murine spleen cells was observed using the synthesized probes in presence of CD1d. Produced cytokines IFN-γ and IL-4 were measured using ELISA. All of the compounds showed high cytokine production ability and selectivity for IFN- γ compared to KRN7000. These results suggest their effective binding with CD1d and following recognition by the T cell receptor (TCR) of iNKT cells. The high potency and selectivity of the ligands also suggests that they can form stable complex with CD1d and TCR. Recently we established surface plasmon resonance technique to determine binding kinetics of CD1d and its ligands.
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| Strategy for Future Research Activity |
So far, we cannot conclude the cytokine stimulation by the synthetic compounds is through CD1d pathway. Therefore, we will study the detail of the binding between CD1d and the synthetic ligands using in vitro experimental techniques, such as surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC). Co-crystallization and crystallographic data accumulation of the hCD1d-synthetic ligands is underway.
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