title: Halofuginone suppresses T cell proliferation by blocking proline uptake and inducing cell apoptosis.
abstract: Inactivation of T cells is a widely used strategy for immunosuppression. Halofuginone (HF) is an antiprotozoal agent for treating parasites in veterinary medicine, and has been demonstrated to inhibit collagen type 1 synthesis, T helper 17 cell differentiation and cytokine production in activated T cells. The present study was designed to examine the biological effects of HF against T cell receptor and interleukin (IL)-2 stimulated T cell proliferation. T cell proliferation in cultured murine splenocytes was determined by methylthiazol tetrazolium assay. Cell apoptosis was mainly determined by fluorescence-activated cell sorting with Annexin-V and 7-aminoactinomycin D staining. Here, we showed that HF significantly suppressed T cell proliferation in naïve splenocyte cultures in response to alloantigen or anti-CD3 antibody (IC₅₀, 2-2.5 nM; P<0.0001), or in activated T cell cultures in response to IL-2 (IC₅₀, 16 nM; P<0.0001) in a dose-dependent manner. HF did neither attenuate IL-2 production in anti-CD3 antibody activated T cells nor disrupt STAT5 signaling in IL-2-stimulated T cells, but its anti-T cell proliferation was correlated with an increase in cell apoptosis and a decrease in proline uptake in culture medium. Further experiments showed that proline supplement in cell culture medium significantly prevented HF-mediated suppression of T cell proliferation and cell apoptosis. In conclusion, these data suggest that HF interferes with proline incorporation or uptake, resulting in apoptosis via amino acid starvation response in T cells in the response to antigen/mitogen or IL-2 stimulation.
authors.affiliation: Department of Urologic Sciences, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Tony L H
authors.initials: TL
authors.lastname: Chu
authors.forename: Qiunong
authors.initials: Q
authors.lastname: Guan
authors.forename: Christopher Y C
authors.initials: CY
authors.lastname: Nguan
authors.forename: Caigan
authors.initials: C
authors.lastname: Du
mesh_terms: D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D003470:Culture Media; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D020287:In Situ Nick-End Labeling; D007376:Interleukin-2 / Q000276:immunology / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects; D051379:Mice; D008807:Mice, Inbred BALB C; D010880:Piperidines / Q000494:pharmacology*; D011392:Proline / Q000378:metabolism*; D052999:Quinazolinones / Q000494:pharmacology*; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000378:metabolism; D013154:Spleen / Q000166:cytology / Q000276:immunology; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism / Q000473:pathology
chemical_list: D000981:Antiprotozoal Agents; D003470:Culture Media; D007376:Interleukin-2; D010880:Piperidines; D052999:Quinazolinones; D011948:Receptors, Antigen, T-Cell; D011392:Proline; C010176:halofuginone