Yukihiko Saeki1,2, Yasutaka Okita3,4, Eri Igashira-Oguro3,4, Chikako Udagawa5,6, Atsuko Murata5, Takashi Tanaka6, Jyunji Mukai7, Keiji Miyazawa8, Yoshihiko Hoshida9, Shiro Ohshima3,5. 1. Rheumatology & Allergology, NHO Osaka Minami Medical Center, Kidohigashi-machi, Kawachinagano, Osaka, 586-8521, Japan. saekiy@ommc-hp.jp. 2. Department of Clinical Research, NHO Osaka Minami Medical Center, 2-1 Kidohigashi-machi, Kawachinagano, Osaka, 586-8521, Japan. saekiy@ommc-hp.jp. 3. Rheumatology & Allergology, NHO Osaka Minami Medical Center, Kidohigashi-machi, Kawachinagano, Osaka, 586-8521, Japan. 4. Respiratory Medicine and Clinical Immunology, Graduate School of Medicine, Osaka University, Yamada-oka, Suita, Osaka, 565-0871, Japan. 5. Department of Clinical Research, NHO Osaka Minami Medical Center, 2-1 Kidohigashi-machi, Kawachinagano, Osaka, 586-8521, Japan. 6. Molecular Chemistry, Faculty of Pharmacology, Osaka Ohtani University, Nishikiori-kita, Tondabayashi, Osaka, 584-8540, Japan. 7. Department of Pharmacy, Izumi-City General Hospital, Wake-Cho, Izumi City, Osaka, 594-0072, Japan. 8. KISSEI Pharmaceutical CO., L.T.D, Yoshino, Matsumoto City, Nagano Prefecture, 399-8710, Japan. 9. Pathology, NHO Osaka Minami Medical Center, Kidohigashi-machi, Kawachinagano, Osaka, 586-8521, Japan.
Abstract
OBJECTIVES: To evaluate the ability of geldanamycin to modulate two opposing TNFα/TNFR1-triggered signals for inflammation and cell death. METHODS: The effects of geldanamycin on TNFα-induced proinflammatory cytokine production, apoptosis, NF-κB activation, caspase activation, and necroptosis in a human rheumatoid synovial cell line (MH7A) were evaluated via ELISA/qPCR, flow cytometry, dual-luciferase reporter assay, and western blotting assay, respectively. In addition, therapeutic effects on murine collagen-induced arthritis (CIA) were also evaluated. RESULTS: Geldanamycin disrupted RIPK1 in MH7A, thereby inhibiting TNFα-induced proinflammatory cytokine production and enhancing apoptosis. TNFα-induced NF-κB and MLKL activation was inhibited, whereas caspase 8 activation was enhanced. Recombinant RIPK1 restored the geldanamycin-mediated inhibition of TNFα-induced NF-κB activation. In addition, GM showed more clinical effectiveness than a conventional biologic TNF inhibitor, etanercept, in murine CIA and significantly attenuated synovial hyperplasia, a histopathological hallmark of RA. CONCLUSIONS: GM disrupts RIPK1 and selectively inhibits the TNFR1-triggered NF-κB activation signaling pathway, while enhancing the apoptosis signaling pathway upon TNFα stimulation, thereby redressing the balance between these two opposing signals in a human rheumatoid synovial cell line. Therapeutic targeting RIPK1 may be a novel concept which involves TNF inhibitor acting as a TNFR1-signal modulator and have great potential for a more fundamental, effective, and safer TNF inhibitor. Key Points • Geldanamycin (GM) disrupts RIPK1 and selectively inhibits the TNFR1-triggered NF-κB activation signaling pathway while enhancing the apoptosis signaling pathway upon TNFα stimulation, thereby redressing the balance between these two opposing signals in a human rheumatoid synovial cell line, MH7A. • GM showed more clinical effectiveness than a conventional biologic TNF-inhibitor, etanercept, in murine collagen-induced arthritis (CIA), and significantly attenuated synovial hyperplasia, a histopathological hallmark of RA. • Therapeutic targeting RIPK1 may be a novel concept which involves TNF inhibitor acting as a TNFR1-signal modulator and have great potential for a more fundamental, effective, and safer TNF-inhibitor.
OBJECTIVES: To evaluate the ability of geldanamycin to modulate two opposing TNFα/TNFR1-triggered signals for inflammation and cell death. METHODS: The effects of geldanamycin on TNFα-induced proinflammatory cytokine production, apoptosis, NF-κB activation, caspase activation, and necroptosis in a humanrheumatoid synovial cell line (MH7A) were evaluated via ELISA/qPCR, flow cytometry, dual-luciferase reporter assay, and western blotting assay, respectively. In addition, therapeutic effects on murine collagen-induced arthritis (CIA) were also evaluated. RESULTS:Geldanamycin disrupted RIPK1 in MH7A, thereby inhibiting TNFα-induced proinflammatory cytokine production and enhancing apoptosis. TNFα-induced NF-κB and MLKL activation was inhibited, whereas caspase 8 activation was enhanced. Recombinant RIPK1 restored the geldanamycin-mediated inhibition of TNFα-induced NF-κB activation. In addition, GM showed more clinical effectiveness than a conventional biologic TNF inhibitor, etanercept, in murine CIA and significantly attenuated synovial hyperplasia, a histopathological hallmark of RA. CONCLUSIONS:GM disrupts RIPK1 and selectively inhibits the TNFR1-triggered NF-κB activation signaling pathway, while enhancing the apoptosis signaling pathway upon TNFα stimulation, thereby redressing the balance between these two opposing signals in a humanrheumatoid synovial cell line. Therapeutic targeting RIPK1 may be a novel concept which involves TNF inhibitor acting as a TNFR1-signal modulator and have great potential for a more fundamental, effective, and safer TNF inhibitor. Key Points • Geldanamycin (GM) disrupts RIPK1 and selectively inhibits the TNFR1-triggered NF-κB activation signaling pathway while enhancing the apoptosis signaling pathway upon TNFα stimulation, thereby redressing the balance between these two opposing signals in a humanrheumatoid synovial cell line, MH7A. • GM showed more clinical effectiveness than a conventional biologic TNF-inhibitor, etanercept, in murine collagen-induced arthritis (CIA), and significantly attenuated synovial hyperplasia, a histopathological hallmark of RA. • Therapeutic targeting RIPK1 may be a novel concept which involves TNF inhibitor acting as a TNFR1-signal modulator and have great potential for a more fundamental, effective, and safer TNF-inhibitor.