| Autor: |
Kang TH; Department of Chemical Engineering, University of Texas at Austin, Austin, TX, United States., Lee CH; Department of Chemical Engineering, University of Texas at Austin, Austin, TX, United States., Delidakis G; Department of Chemical Engineering, University of Texas at Austin, Austin, TX, United States., Jung J; Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, United States., Richard-Le Goff O; Unit of Antibodies in Therapy and Pathology, Department of Immunology, Institut Pasteur, Paris, France., Lee J; Department of Chemical Engineering, University of Texas at Austin, Austin, TX, United States., Kim JE; Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, United States., Charab W; Department of Chemical Engineering, University of Texas at Austin, Austin, TX, United States., Bruhns P; Unit of Antibodies in Therapy and Pathology, Department of Immunology, Institut Pasteur, Paris, France.; INSERM, U1222, Paris, France., Georgiou G; Department of Chemical Engineering, University of Texas at Austin, Austin, TX, United States.; Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, United States.; Center for Systems and Synthetic Biology, University of Texas at Austin, Austin, TX, United States.; Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, United States. |
| Abstrakt: |
IgG antibodies mediate the clearance of target cells via the engagement of Fc gamma receptors (FcγRs) on effector cells by eliciting antibody-dependent cellular cytotoxicity and phagocytosis (ADCC and ADCP, respectively). Because (i) the IgG Fc domain binds to multiple FcγRs with varying affinities; (ii) even low Fc:FcγRs affinity interactions can play a significant role when antibodies are engaged in high avidity immune complexes and (iii) most effector cells express multiple FcγRs, the clearance mechanisms that can be mediated by individual FcγR are not well-understood. Human FcγRIIIa (hFcγRIIIa; CD16a), which exists as two polymorphic variants at position 158, hFcγRIIIa V158 and hFcγRIIIa F158 , is widely considered to only trigger ADCC, especially with natural killer (NK) cells as effectors. To evaluate the role of hFcγRIIIa ligation in myeloid-derived effector cells, and in particular on macrophages and monocytes which express multiple FcγRs, we engineered an aglycosylated engineered human Fc (hFc) variant, Fc3aV, which binds exclusively to hFcγRIIIa V158 . Antibodies formatted with the Fc3aV variant bind to the hFcγRIIIa V158 allotype with a somewhat lower K D than their wild type IgG1 counterparts, but not to any other hFcγR. The exceptional selectivity for hFcγRIIIa V158 was demonstrated by SPR using increased avidity, dimerized GST-fused versions of the ectodomains of hFcγRs and from the absence of binding of large immune complex (IC) to CHO cells expressing each of the hFcγRs, including notably, the FcγRIIIa F158 variant or the highly homologous FcγRIIIb. We show that even though monocyte-derived GM-CSF differentiated macrophages express hFcγRIIIa at substantially lower levels than the other two major activating receptors, namely hFcγRI or hFcγRIIa, Fc3aV-formatted Rituximab and Herceptin perform ADCP toward CD20- and Her2-expressing cancer cells, respectively, at a level comparable to that of the respective wild-type antibodies. We further show that hFcγRIIIa activation plays a significant role on ADCC by human peripheral monocytes. Our data highlight the utility of Fc3aV and other similarly engineered exquisitely selective, aglycosylated Fc variants toward other hFcγRs as tools for the detailed molecular understanding of hFcγR biology. |
