Synthesis of glycopeptides from type II collagen-incorporating galactosylated hydroxylysine mimetics and their use in studying the fine specificity of arthritogenic T cells

Five analogues of the bovine type II collagen (bcII) immunodominant glycopeptide [beta-D-Gal-(5R)-5-Hyl264]CII(256-270) (1) carrying diverse modifications at the critical hydroxylysine (Hyl) 264 side chain were designed and synthesised, to explore the fine specificity of bCII-reactive T cells involved in the initiation and/or regulation of collagen-induced arthritis (CIA), a mouse model for rheumatoid arthritis (RA). beta-D-Galactosyl-(5R)-5-hydroxy-L-lysine (79) and corresponding mimetics (22-25), conveniently protected for solid-phase synthesis, were all obtained by a divergent, route involving enantiopure 5-hydroxylated 6-oxo-1,2-piperidinedicarboxylates as the key intermediates. All three bcII-specific T hybridomas used, as well as a recurrent pathogenic CD4(+) T-cell clone isolated from bCII-immunised DBA/1 mice, recognised the galactosylated form I of the immunodominant bCII (256-270) epitope. These cells were extremely sensitive to changes at the epsilon-amino group of Hyl264, but differed in their pattern of recognition of analogues with a Hyl264 side chain modified at C-5 (i.e. inversion of stereochemistry, methylation). These data further document the importance of collagen post-translational modifications in autoimmunity and in the CIA model in particular, and provide a new insight into the molecular interaction between glycopeptide I and the TCR of pathogenic T cells.