SummarySelf-peptides bound to HLA-DR4 (DRA-DRBl*0405 complex) were eluted from the purified DR4 complex, ffactionated on reverse-phase HPLC, and subjected to NH2-terminal sequencing. Seven independent sequences were obtained, and all putative peptides synthesized bound to DRBI*0405 as well as DRBI*0406 complex, which differ only at DRfl residues 37, 57, 74, and 86. Binding assay using analogue peptides of a DR4 binder GSTVFDNLPNPE revealed that FxxLxN is an important anchor motif necessary for binding (where x is any amino acid), which was common to DRBI*0405 and 0406. Determination of the binding affinity of 60 synthetic AAFAALANAA-based analogue peptides showed that substituting F to W or C; L to F, W, or Y; and N to Q or S on AAFAALANAA changed the affinity substantially between DRBI*0405 and DRBI*0406. It is noteworthy that all patients with methimazole-induced insulin autoimmune syndrome are positive for DRBI*0406 and negative for DRBI*0405. Interestingly, the quantitative structural motif identified in this study predicted that 8TSICSLYQLE 17 of human insulin cx chain may bind specifically to DRBI*0406 using its l~ motif. Indeed, DRBI*0406 complex bound sTSICSLYQLE17 with a high affinity, and in striking contrast, DRBI*0405 complex did not. Furthermore, a short-term T cell line specific to human insulin established from a DRBI*0406-bearing individual did show reactivity with a peptide fragment containing the l~ motif. Although this fragment probably exists at a very low level under normal physiological conditions due to the disulfide bond between flanking cysteine residues (6Cys-llCys), a reducing compound such as methimazole may cleave the disulfide bond in vivo and allow DRcr-DRBI*0406 complex on antigen-presenting cells to bind much of the linear fragment of insulin c~ chain, which may lead to the activation of self-insulin-specific T-helper cells.

Infection with Shiga toxin (Stx)-producing Escherichia coli (STEC) causes hemorrhagic colitis and hemolytic uremic syndrome. TMA-15 is a humanized monoclonal antibody against Stx2, a major pathogenic factor. In a mouse infection model that used B2F1, a virulent STEC strain, the efficacy of TMA-15 was assessed when it was administered after bacterial and toxin exposure. In this model, a time-course analysis of the serum Stx2 level showed that the toxin was detectable from 24 h after infection. In an evaluation of the time-dependent efficacy, treatment with TMA-15 up to 24 h after infection ameliorated the lethal challenge, although treatment at 48 h showed no efficacy. To determine the effective dose, escalating doses were administered at 24 h after infection. The number of mice that survived after doses of 0, 0.25, 0.5, 1.0, and 2.0 mg/kg were 0/20, 11/20, 17/20, 20/20, and 20/20, respectively. These findings suggest that TMA-15 shows potential for prevention of severe complications associated with STEC infection.

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