Monoclonal antibodies are being considered as
biopharmaceuticals for the in vivo targeting of
acute myeloid leukemia. Here we describe the generation and characterization of a fully-human
monoclonal antibody specific to CD123, a surface marker which is overexpressed in a variety of hematological disorders, including
acute myeloid leukemia. The cloning and expression of the extracellular portion of CD123 as recombinant Fc fusion allowed the selection and affinity maturation of a human antibody, called H9, which specifically recognized the cognate
antigen in biochemical assays and on leukemic cells. The H9 antibody and a previously-described anti-CD123 antibody (CSL362) were reformatted into full
immunoglobulin human
IgG1 formats, including a variant bearing S293D and I332E mutations to enhance antibody-dependent cell-mediated cytotoxicity (ADCC). The two
antibodies recognized different
epitopes on the surface of the N-terminal domain of CD123, as revealed by crystallography and SPOT analysis. Both H9 and CSL362 in full
immunoglobulin format were able to selectively kill leukemic cells in in vitro ADCC assays, performed both with cell lines and with patient-derived AML blasts. Further, the two
antibodies, when reformatted as bispecific BiTE™
reagents by fusion with the anti-CD3 scFv(
OKT3)
antibody fragment, induced selective killing of AML blasts by patient-derived, autologous T-cells in an in vitro setting, but
BiTE(CSL362/
OKT3) exhibited a 10-fold higher potency compared to
BiTE(H9/
OKT3). The availability of two classes of CD123-specific
biopharmaceuticals, capable of redirecting the cytolytic activity of NK cells and T cells against AML blasts, may enable novel interventional strategies and combination opportunities for the treatment of AML.