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Background:  MHC alleles carried on the 57.1 ancestral haplotype (AH), including HLA-B*5701 and heat shock protein (Hsp70) genes, have been shown to be highly predictive of abacavir (ABC) hypersensitivity suggesting a strong immunogenetic basis for this syndrome. We wanted to characterize the specific antigen processing pathways involved in eliciting ABC-specific immune responses.

Methods:  The effect of ABC on intracellular localisation of HLA-B57, Hsp70 in peripheral blood mononuclear cells (PBMC), monocyte-derived dendritic cells (MDDC) was examined by immunofluorescence confocal microscopy from ABC hypersensitivity (n = 9), ABC-tolerant controls (n = 6), 57.1 AH-positive ABC-naïve individuals (n = 8), and B-cell lines homozygous for MHC haplotypes (n = 13). Hsp70 redistribution was quantified using Image J. The influence of 4-MP-mediated inhibition of abacavir pro-drug metabolising enzyme, type I alcohol dehydrogenase levels on Hsp70 redistribution and interferon-γ (IFN-γ) was also studied.

Results:  Co-localized distribution of Hsp70 and HLA-B57 was observed in CD14⁺ and CD56⁺ cells and was located within peripheral vesicles staining positively for early, late endosomal, or phagosomal markers. Increase in intracellular Hsp70 redistribution occurred after 3-hour ABC stimulation in PBMC of ABC-hypersensitive (n = 8) and ABC-naïve (n = 3) compared with controls (n = 5) with 21.19, 47.0 vs 1.50 average arbitrary units (p = 0.023). Following a 40-hour 4-MP exposure in PBMC of ABC-hypersensitive patients (n = 6), ABC-specific Hsp70 redistribution decreased from 11.2 to 2.9 mean arbitrary units (p = 0.024). Hsp70 redistribution levels correlated with extracellular IFN-γ (r = 0.64, p = 0.0335). In B-cell lines, tumor necrosis factor (TNF)–238A allele was associated with increased Hsp70 redistribution after ABC exposure. Biotinylated ABC retained immunogenicity compared with unlabelled ABC, co-localizing with Hsp70 and HLA-B57 within similar compartments and stimulating similar levels of IFN-γ in PBMC cultures.

Conclusions:  These data suggest that Hsp70 redistribution represents an early component of the ABC-specific immune response which is sensitive to inhibition of type I alcohol dehydrogenase, and which influences IFN-γ expression. Regulation of Hsp70 redistribution may also be associated or linked with TNF-238A allele. The development of HSR thus represents a multi-step process likely to involve the generation of an aldehyde metabolite by alcohol dehydrogenase followed by intracellular Hsp70 redistribution and finally development of an HLA-B*5701-restricted immune response mediated by inflammatory cytokines.

Keywords: abacavir hypersensitivity; Hsp70; alcohol dehydrogenase