Glycolysis: An early marker for vancomycin-specific T-cell activation

BackgroundVancomycin, a glycopeptide antibiotic used for Gram-positive bacterial infections, has been linked with drug reaction with eosinophilia and systemic symptoms (DRESS) in HLA-A*32:01-expressing individuals. This is associated with activation of T lymphocytes, for which glycolysis has been isolated as a fuel pathway following antigenic stimulation. However, the metabolic processes that underpin drug-reactive T-cell activation are currently undefined and may shed light on the energetic conditions needed for the elicitation of drug hypersensitivity or tolerogenic pathways. Here, we sought to characterise the immunological and metabolic pathways involved in drug-specific T-cell activation within the context of DRESS pathogenesis using vancomycin as model compound and drug-reactive T-cell clones (TCCs) generated from healthy donors and vancomycin-hypersensitive patients.MethodsCD4+ and CD8+ vancomycin-responsive TCCs were generated by serial dilution. The Seahorse XFe96 Analyzer was used to measure the extracellular acidification rate (ECAR) as an indicator of glycolytic function. Additionally, T-cell proliferation and cytokine release (IFN-gamma) assay were utilised to correlate the bioenergetic characteristics of T-cell activation with in vitro assays.ResultsModel T-cell stimulants induced non-specific T-cell activation, characterised by immediate augmentation of ECAR and rate of ATP production (JATPglyc). There was a dose-dependent and drug-specific glycolytic shift when vancomycin-reactive TCCs were exposed to the drug. Vancomycin-reactive TCCs did not exhibit T-cell cross-reactivity with structurally similar compounds within proliferative and cytokine readouts. However, cross-reactivity was observed when analysing energetic responses; TCCs with prior specificity for vancomycin were also found to exhibit glycolytic switching after exposure to teicoplanin. Glycolytic activation of TCC was HLA restricted, as exposure to HLA blockade attenuated the glycolytic induction.ConclusionThese studies describe the glycolytic shift of CD4+ and CD8+ T cells following vancomycin exposure. Since similar glycolytic switching is observed with teicoplanin, which did not activate T cells, it is possible the master switch for T-cell activation is located upstream of metabolic signalling. Vancomycin has been linked with drug reaction with DRESS in HLA-A*32:01 expressing individuals and is associated with activation of drug-specific T lymphocytes. However, the metabolic processes than underpin drug-reactive T-cell activation are undefined. CD4+ and CD8+ vancomycin-specific T-cell clones (TCCs) generated from healthy donors and hypersensitive patients expressing HLA-A*32:01 undergo dose-dependent, drug-specific and HLA restricted glycolytic switching when activated with vancomycin. TCCs displayed 'energetic' cross-reactivity with teicoplanin, indicating the master switch for T-cell activation may be located upstream of metabolic signalling.image