Rational design of a bispecific antibody for rapid and prolonged neutrophil mobilization and proliferation

Human granulocyte colony stimulating factor (G-CSF) and the G-protein C-X-C chemokine receptor 4 (CXCR4)/stromal cell-derived factor-1 (SDF-1) axis play pivotal roles in neutrophil mobilization, differentiation and proliferation. Bispecific antibodies represent a viable therapeutic strategy by simultaneously targeting these two granulocyte-associated receptors while providing an extended half-life. In this study, we engineered a bispecific antibody by grafting human G-CSF and a synthetic anti-CXCR4 peptide into the heavy chain complementarity-determining region 3 (CDR3) of the humanized antibody Herceptin. This bispecific antibody targets both G-CSF receptor (G-CSFR) and CXCR4, demonstrating rapid and long-last activities in increasing neutrophil counts, surpassing the efficacy of combined G-CSF and CXCR4-targeted therapies. Its ability to synergize the effects of G-CSF stimulation and CXCR4 inhibition underscores the potential as a new treatment approach for neutropenia. Moreover, this CDR grafting strategy provides a versatile platform for designing bispecific antibodies that combine agonistic and antagonistic activities.