Lipid and glycolipid antigens of CD1d-restricted natural killer T cells

Abstract

In spite of their relatively limited antigen receptor repertoire, CD1d-restricted NKT cells recognize a surprisingly diverse range of lipid and glycolipid antigens. Recent studies of natural and synthetic CD1d-presented antigens provide an increasingly detailed picture of how the specific structural features of these lipids and glycolipids influence their ability to be presented to NKT cells and stimulate their diverse immunologic functions. Particularly for synthetic analogues of alpha-galactosylceramides which have been the focus of intense recent investigation, it is becoming clear that the design of glycolipid antigens with the ability to precisely control the specific immunologic activities of NKT cells is likely to be feasible. The emerging details of the mechanisms underlying the structure-activity relationship of NKT cell antigens will assist greatly in the design and production of immunomodulatory agents for the precise manipulation of NKT cells and the many other components of the immune system that they influence.

Fig 1

Structure of the prototypical synthetic iNKT cell antigen KRN7000, which is an αGalCer with a C26:0 amide linked acyl chain and a C18 saturated phytosphingosine base. Key structural components that have been systematically modified to create analogues of this potent iNKT cell antigen are indicated by the boxed areas.

Fig 2

Four examples of sphingosine chain analogues of KRN7000 with iNKT cell activating properties. OCH is a Th2-cytokine biasing iNKT cell activator, which has a markedly truncated C9 phytosphingosine and a slightly shortened N-acyl chain (C24:0) compared to KRN7000. C13 is an example of a truncated sphingosine derivative in which incorporation of an aromatic ring led to increased iNKT cell activating potency. Triazole 8 has a 24 carbon alkyl chain linked to the sphingoid base by triazole ring, which is a highly stable rigid linking unit that mimics the topological and electronic features of an amide bond. RCAI-18 demonstrates how a trans-2,4-dialkylated azetidine ring can be incorporated in the ceramide portion to restrict flexibility while retaining the ability to stimulate iNKT cell responses. See text for references.

Fig 3

Structures of representative N-acyl chain analogues of KRN7000. (A) Five examples are shown of glycolipids that induce Th2-biased iNKT cell responses. Although the structures vary widely, they all contain acyl chain modifications that introduce increased polarity such as unsaturations, chain truncation or incorporation of oxygen atoms. (B) An example of a highly active analogue in which the acyl chain is terminated by an aromatic ring. Such compounds have been reported to be potent iNKT cell activators and have been suggested to favor a more Th1-biased cytokine response. See text for references.

Fig 4

Structures of representative carbohydrate and glycosidic bond derivatives of KRN7000. (A) Glycosidic bond substitution based derivatives including the original C-glycoside of KRN7000 (α-C-GalCer with a methylene unit (-CH₂-) in place of oxygen in the glycosidic linkage, and GCK109 which is a highly active E-alkene linked analogue of α-C-GalCer. (B) Carbohydrate modification based derivatives with replacement of galactose by α-linked glucose, di-galactose (Gal(α1→6)αGalCer) or 1-naphthyl group linked to the 6'-ureido-6'deoxy-αGalCer. (C) An active non-glycosidic derivative represented by threitolceramide. See text for references.

Fig 5

Model for mechanism leading to stimulation of biased cytokine responses by analogues of αGalCer. The αGalCer analogues such as KRN7000 (shown in red) that generate mixed cytokine responses comprised of both Th1 and Th2 type cytokines are endocytosed after binding to extracellular lipid carrier proteins such as ApoE and loaded into CD1d intracellularly in endosomes where this process is facilitated by low pH and the presence of lipid transfer proteins such as the GM2 activator protein (GM2A) and saposins. These intracellularly generated CD1d-lipid complexes are transported to lipid raft microdomains on the plasma membrane and activate strong and prolonged iNKT cell responses that are characterized by a mixture of Th1 and Th2 type cytokines with prominent and sustained IFNγ production. Note that the majority of IFNγ generated in this situation is most likely not a direct product of the activated iNKT cells, but is generated by the secondary activation of NK cells. In contrast, glycolipids that are rapidly loaded into surface CD1d (shown in green) activate iNKT cells to induce responses that are dominated by Th2 type cytokines without stimulating prolonged secretion of IFNγ or other proinflammatory Th1 type cytokines. Glycolipids of this type are unloaded from CD1d in the late endosome and lysosome, so that intracellular complexes containing these are inhibited from forming and therefore do not undergo transport into plasma membrane lipid rafts. TGN, trans Golgi network; ER, endoplasmic reticulum. See text for more detailed explanation and references.