Recombinant chimeric OKT3 scFv IgM antibodies mediate immune suppression while reducing T cell activation in vitro

OKT3, a mouse anti-human CD3 monoclonal antibody (mAb), is a potent immunos uppressive agent used in clinical transplantation to treat allograft reject ion. Two major drawbacks of this therapy are the systemic release of severa l cytokines due to cross-linking mediated by the mAb between T cells and Fc gammaR-bearing cells and the human anti-mouse antibody (HAMA) response. To overcome these side effects, three chimeric OKT3 single chain variable fra gment (scFv) IgM antibodies, scOKT3-gamma Delta IgM wt, scOKT3-gamma Delta IgM C575S and scOKT3-gamma Delta IgM VAEVD, were generated. They consist of the light and heavy variable binding domains of OKT3 mAb as well as the CH 3 and CH4 domains of different human IgM variants linked with a human IgG3 hinge region to provide more flexibility and stability Like the native IgM, scOKT3-gamma Delta IgM antibodies are able to form polymeric structures, w hich lead to an increase in binding affinity and immunosuppressive potentia l compared with the parental OKT3 mAb. However, independently of their poly merization, all scOKT3-gamma Delta IgM constructs do not induce any signifi cant T cell proliferation or cytokine release (IL-2, TNF-alpha and IFN-gamm a) in in vitro assays, while their CD3-modulating properties are retained. These results suggest that the use of scOKT3-gamma Delta IgM antibodies may offer significant advantages over the OKT3 mAb in improving clinical immun osuppressive treatment.