Activated T cells are key players in chronic inflammatory diseases, including
atherosclerosis.
Salicylates, like
aspirin, display not only anti-inflammatory, anti-thrombotic, anti-atherosclerotic activities, but also immunomodulatory effects in T cells at high dosages. Here, we aimed to identify potent
immunomodulators for T cells through cell-based screening from a mini-library of 300
salicylate-based small molecules, and elucidate the mechanisms. Human peripheral blood T cells were isolated from buffy coat.
Phorbol 12-myristate 13-acetate plus
ionomycin (P/I) was used to stimulate T cells.
Cytokine production was measured by
enzyme-linked
immunosorbent assays. T cell activation markers were determined by flow cytometry. The activation of
transcription factors and
kinases was analyzed by western blotting, electrophoretic mobility shift assay, or
kinase assay. Through library screening, we identified a small molecule named HS-Cm [C13H9ClFNO2; N-(4-chloro-2-fluorophenyl)-2-hydroxybenzamide] that exhibited potent immunomodulatory effects on T cells with low cytotoxicity. In P/I-stimulated T cells, HS-Cm inhibited the production of
interleukin-2,
tumor necrosis factor-alpha, and
interferon-gamma and suppressed the expression of surface activation markers CD25, CD69, and CD71, but not CD45RO. HS-Cm down-regulated
DNA-binding activities of
activator protein-1 and
nuclear factor-kappa B, but not nuclear factor of activated T-cells, through inhibiting
c-Jun N-terminal kinase/p38 and inhibitor of kappaB alpha (IκBα)
kinase (IKK)/IκBα pathways, respectively. On the basis of structure-activity relationship, HS-Cm exerted considerable inhibition of
dipeptidyl-peptidase IV/CD26 activity in T cells. Our results suggested that the small molecule HS-Cm exhibiting immunomodulatory effects on T cells may be useful for
therapeutics in chronic inflammatory diseases, like
atherosclerosis, diabetes and autoimmune
arthritis.