4"-O-Alkylated α-Galactosylceramide Analogues as iNKT-Cell Antigens: Synthetic, Biological, and Structural Studies.

Invariant natural killer T-cells (iNKT) are a glycolipid-responsive subset of T-lymphocytes that fulfill a pivotal role in the immune system. The archetypical synthetic glycolipid, alpha-galactosylceramide (alpha-GalCer), whose molecular framework is inspired by a group of amphiphilic natural products, remains the most studied antigen for iNKT-cells. Nonetheless, the potential of alpha-GalCer as an immunostimulating agent is compromised by the fact that this glycolipid elicits simultaneous secretion of Th1- and Th2-cytokines. This has incited medicinal chemistry efforts to identify analogues that are able to perturb the Th1/Th2 balance. In this work, we present the synthesis of an extensive set of 4"-O-alkylated alpha-GalCer analogues, which were evaluated in vivo for their cytokine induction. We have found that conversion of the 4"-OH group to ether moieties decreases the immunogenic potential in mice relative to alpha-GalCer. Yet, the benzyl-modified glycolipids are able to produce a distinct pro-inflammatory immune response. The crystal structures suggest an extra hydrophobic interaction between the benzyl moiety and the alpha 2-helix of CD1d.