Cytotoxicity of sulfonamide reactive metabolites: apoptosis and selective toxicity of CD8 super(+) cells by the hydroxylamine of sulfamethoxazole

Treatment with sulfonamide antibiotics in HIV-infected patients is associated with a high incidence (> 40%) of adverse drug events, including severe hypersensitivity reactions. Sulfonamide reactive metabolites have been implicated in the pathogenesis of these adverse reactions. Sulfamethoxazole hydroxylamine (SMX-HA) induces lymphocyte toxicity and suppression of proliferation in vitro; the mechanism(s) of these immunomodulatory effects remain unknown. We investigated the cytotoxicity of SMX-HA via apoptosis on human peripheral blood mononuclear cells and purified cell subpopulations in vitro. CD19 super(+), CD4 super(+), and CD8 super(+) cells were isolated from human peripheral blood by positive selection of cell surface molecules by magnetic bead separation. SMX-HA induced significant CD8 super(+) cell death (67 plus or minus 7%) at 100 mu M SMX-HA, with only minimal CD4 super(+) cell death (8 plus or minus 4%). No significant subpopulation toxicity was shown when incubated with parent drug (SMX). Flow cytometry measuring phosphatidylserine externalization 24 h after treatment with 100 mu M and 400 mu M SMX-HA revealed 14.1 plus or minus 0.7% and 25.6 plus or minus 4.2% annexin-positive cells, respectively, compared to 3.7 plus or minus 1.2% in control PBMCs treated with 400 mu M SMX. Internucleosomal DNA fragmentation was observed in quiescent and stimulated PBMCs 48 h after incubation with SMX-HA. Our data show that CD8 super(+) cells are highly susceptible to the toxic effects of SMX-HA through enhanced cell death by apoptosis.