A salicylate-based small molecule HS-Cm exhibits immunomodulatory effects and inhibits dipeptidyl peptidase-IV activity in human T cells.
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.
Jun-Ting
JT
Liou
Hsu-Shan
HS
Huang
Meng-Lin
ML
Chiang
Chin-Sheng
CS
Lin
Shih-Ping
SP
Yang
Ling-Jun
LJ
Ho
Jenn-Haung
JH
Lai
D001549:Benzamides / Q000737:chemistry / Q000494:pharmacology*; D015415:Biomarkers / Q000378:metabolism; D016207:Cytokines / Q000096:biosynthesis; D004247:DNA / Q000378:metabolism; D018819:Dipeptidyl Peptidase 4 / Q000378:metabolism*; D054873:Dipeptidyl-Peptidase IV Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007155:Immunologic Factors / Q000737:chemistry / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects; D016328:NF-kappa B / Q000378:metabolism; D020156:Salicylic Acid / Q000737:chemistry*; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology*; D018808:Transcription Factor AP-1 / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D001549:Benzamides; D015415:Biomarkers; D016207:Cytokines; D054873:Dipeptidyl-Peptidase IV Inhibitors; D007155:Immunologic Factors; C577570:N-(4-chloro-2-fluorophenyl)-2-hydroxybenzamide; D016328:NF-kappa B; D018808:Transcription Factor AP-1; D004247:DNA; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D018819:Dipeptidyl Peptidase 4; D020156:Salicylic Acid