T cell costimulation with anti-CD137 monoclonal antibodies is mediated by K63-polyubiquitin-dependent signals from endosomes

Agonist anti-CD137 (4-1BB) mAbs enhance CD8-mediated antitumor immunity. Agonist anti-human CD137 mAbs binding to four distinct epitopes on the CD137 glycoprotein costimulated T cell activation irrespective of the engaged epitope or its interference with CD137L binding. CD137 perturbation with all these agonist mAbs resulted in Ag and Ab internalization toward an endosomal vesicular compartment. Internalization was observed in activated T lymphocytes from humans and mice, not only in culture but also in Ab-injected living animals. These in vivo experiments were carried out upon systemic i.v. injections with anti-CD137 mAbs and showed CD137 internalization in tumor-infiltrating lymphocytes and in activated human T cells transferred to immunodeficient mice. Efficient CD137 internalization required K63 polyubiquitination and endocytosed CD137-containing vesicles recruited TNFR-associated factor (TRAF) 2 and were decorated with K63 polyubiquitins. CD137 stimulation activates NF-κB through a K63-linked polyubiquitination-dependent route, and CD137-associated TRAF2 becomes K63 polyubiquitinated. Consistent with a role for TRAF2 in CD137 signaling, transgenic mice functionally deficient in TRAF2 showed delayed immunotherapeutic activity of anti-CD137 mAbs. As a whole, these findings advance our knowledge of the mechanisms of action of anti-CD137 immunostimulatory mAbs such as those currently undergoing clinical trials in cancer patients.This work was supported by the Ministerio de Economía y Competitividad (SAF2008-03294 and SAF2011-22831 to I.M. and SAF2011-23846 and PI12/01135 to J.M.Z.). I.M. was also supported by the Departamento de Educacion del Gobierno de Navarra and the Departamento de Salud del Gobierno de Navarra, Redes Tematicas de Investigacion Cooperativa (RD06/0020/0065), the European Commission VII Framework Programme, the Programme of Territorial Cooperation of the European Southwest Area-Immunotherapy Network, and “Union Temporal de Empresas for project Fundacion de Investigacion Médica Aplicada.” I.M.-F. is supported by an Alejandro Lopez Fellowship from the World Bank-Departamento Administrativo de Ciencia, Tecnología e Innovación Colciencias. A.P. is a recipient of a Fondo de Investigaciones Sanitarias predoctoral scholarship. A.T. and A.M.-K. are recipients of predoctoral scholarships from the Ministerio de Economia. G.P.-C. is the recipient of a Junta de Ampliacion de Estudios contract from the Consejo Superior de Investigaciones Científicas.Peer Reviewe

T cell costimulation with anti-CD137 monoclonal antibodies is mediated by K63-polyubiquitin-dependent signals from endosomes

Agonist anti-CD137 (4-1BB) mAbs enhance CD8-mediated antitumor immunity. Agonist anti-human CD137 mAbs binding to four distinct epitopes on the CD137 glycoprotein costimulated T cell activation irrespective of the engaged epitope or its interference with CD137L binding. CD137 perturbation with all these agonist mAbs resulted in Ag and Ab internalization toward an endosomal vesicular compartment. Internalization was observed in activated T lymphocytes from humans and mice, not only in culture but also in Ab-injected living animals. These in vivo experiments were carried out upon systemic i.v. injections with anti-CD137 mAbs and showed CD137 internalization in tumor-infiltrating lymphocytes and in activated human T cells transferred to immunodeficient mice. Efficient CD137 internalization required K63 polyubiquitination and endocytosed CD137-containing vesicles recruited TNFR-associated factor (TRAF) 2 and were decorated with K63 polyubiquitins. CD137 stimulation activates NF-κB through a K63-linked polyubiquitination-dependent route, and CD137-associated TRAF2 becomes K63 polyubiquitinated. Consistent with a role for TRAF2 in CD137 signaling, transgenic mice functionally deficient in TRAF2 showed delayed immunotherapeutic activity of anti-CD137 mAbs. As a whole, these findings advance our knowledge of the mechanisms of action of anti-CD137 immunostimulatory mAbs such as those currently undergoing clinical trials in cancer patients.This work was supported by the Ministerio de Economía y Competitividad (SAF2008-03294 and SAF2011-22831 to I.M. and SAF2011-23846 and PI12/01135 to J.M.Z.). I.M. was also supported by the Departamento de Educacion del Gobierno de Navarra and the Departamento de Salud del Gobierno de Navarra, Redes Tematicas de Investigacion Cooperativa (RD06/0020/0065), the European Commission VII Framework Programme, the Programme of Territorial Cooperation of the European Southwest Area-Immunotherapy Network, and “Union Temporal de Empresas for project Fundacion de Investigacion Médica Aplicada.” I.M.-F. is supported by an Alejandro Lopez Fellowship from the World Bank-Departamento Administrativo de Ciencia, Tecnología e Innovación Colciencias. A.P. is a recipient of a Fondo de Investigaciones Sanitarias predoctoral scholarship. A.T. and A.M.-K. are recipients of predoctoral scholarships from the Ministerio de Economia. G.P.-C. is the recipient of a Junta de Ampliacion de Estudios contract from the Consejo Superior de Investigaciones Científicas.Peer Reviewe