title: Azithromycin suppresses CD4(+) T-cell activation by direct modulation of mTOR activity.
abstract: Advanced macrolides, such as azithromycin (AZM) or clarithromycin (CLM), are antibiotics with immunomodulatory properties. Here we have sought to evaluate their in vitro influence on the activation of CD4(+) T-cells. Isolated CD4(+) T-cells were stimulated with agonistic anti-CD3/anti-CD28 monoclonal antibodies in the presence of 0.6 mg/L, 2.5 mg/L, 10 mg/L or 40 mg/L AZM or CLM. Cell proliferation, cytokine level in supernatants and cell viability was assessed. Intracellular signaling pathways were evaluated using reporter cell lines, FACS analysis, immunoblotting and in vitro kinase assays. AZM inhibited cell proliferation rate and cytokine secretion of CD4(+) T-cells in a dose-dependent manner. Similarly, high concentrations of CLM (40 mg/L) also suppressed these T-cell functions. Analysis of molecular signaling pathways revealed that exposure to AZM reduced the phosphorylation of the S6 ribosomal protein, a downstream target of mTOR. This effect was also observed at 40 mg/L CLM. In vitro kinase studies using recombinant mTOR showed that AZM inhibited mTOR activity. In contrast to rapamycin, this inhibition was independent of FKBP12. We show for the first time that AZM and to a lesser extent CLM act as immunosuppressive agents on CD4(+) T-cells by inhibiting mTOR activity. Our results might have implications for the clinical use of macrolides.
authors.affiliation: Department of Laboratory Medicine, Medical University of Vienna, Austria.
authors.forename: F
authors.initials: F
authors.lastname: Ratzinger
authors.affiliation: Department of Laboratory Medicine, Medical University of Vienna, Austria.
authors.forename: H
authors.initials: H
authors.lastname: Haslacher
authors.affiliation: Division of Infectious Diseases and Tropical Medicine, Department of Medicine I, Medical University of Vienna, Austria.
authors.forename: W
authors.initials: W
authors.lastname: Poeppl
authors.affiliation: Department of Laboratory Medicine, Medical University of Vienna, Austria.
authors.forename: G
authors.initials: G
authors.lastname: Hoermann
authors.affiliation: Clinical Division of Nephrology and Dialysis, Department of Internal Medicine III, Medical University of Vienna, Austria.
authors.forename: J J
authors.initials: JJ
authors.lastname: Kovarik
authors.affiliation: Institute of Immunology, Medical University of Vienna, Austria.
authors.forename: S
authors.initials: S
authors.lastname: Jutz
authors.affiliation: Institute of Immunology, Medical University of Vienna, Austria.
authors.forename: P
authors.initials: P
authors.lastname: Steinberger
authors.affiliation: Division of Infectious Diseases and Tropical Medicine, Department of Medicine I, Medical University of Vienna, Austria.
authors.forename: H
authors.initials: H
authors.lastname: Burgmann
authors.affiliation: Institute of Immunology, Medical University of Vienna, Austria.
authors.forename: W F
authors.initials: WF
authors.lastname: Pickl
authors.affiliation: Department of Laboratory Medicine, Medical University of Vienna, Austria.
authors.forename: K G
authors.initials: KG
authors.lastname: Schmetterer
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D017963:Azithromycin / Q000494:pharmacology*; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects* / Q000502:physiology; D049109:Cell Proliferation; D017291:Clarithromycin / Q000494:pharmacology; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D008213:Lymphocyte Activation; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D038601:Ribosomal Protein S6 / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D017963:Azithromycin; D017291:Clarithromycin; D016207:Cytokines; D007166:Immunosuppressive Agents; D019869:Phosphatidylinositol 3-Kinases; D038601:Ribosomal Protein S6; D058570:TOR Serine-Threonine Kinases; C546842:MTOR protein, human