Hemoglobin (Hb) is known to stimulate erythropoiesis, a process that may be mediated by CD163. CD163 is a receptor for the hemoglobin-haptoglobin (Hb-Hp) complex expressed on monocyte/macrophages as well as a subpopulation of human CD34+ hematopoietic progenitor cells (HPCs). We have demonstrated that administration of ligands to the CD163 receptor can measurably stimulate erythropoiesis in human CD34+ cell-engrafted severe-combined immunodeficiency (SCID) mice. To better elucidate the role of CD163 in hematopoiesis, we investigated the effects of the natural ligand to CD163 (Hb-Hp) as well as a stimulatory antibody, TBI 304H, on HPCs in vivo. SCID mice engrafted with human CD34+ cells were used as a model system were used to investigate the effect of Hb and anti-CD163 monoclonal antibodies (TBI 304 and TBI 304H) on human hematopoiesis in vivo. In an initial study, NOD-SCID IL2R gammanull (NSG) mice were engrafted with human CD34+ cells and animals with < 30% human CD45+ cells in the peripheral blood were administered 2 mg Hb/mouse, or 100 or 500 µg/mouse TBI 304 every 4 days for a total of four doses. At study termination on day 14, bone marrow cells (BMCs) were examined by flow cytometry and CD34+ cells were recovered from the BMCs for enumeration in colony-forming assays. Hemoglobin administration resulted in an increase of human CD34+ cells ranging from 4% to 7% of BMCs and a corresponding 57% increase in colony-forming cells (CFCs) over control animals. In contrast, the monoclonal antibody (mAb), TBI 304, produced a dose-dependent decrease in CD34+ and colonies, possibly reflecting a depletion of CD34+/CD163+ cells as a result of overstimulation due to the much longer circulating half-life of the mAb compared to Hb.. To confirm this hypothesis, human CD34+ cell engrafted animals were given only a single dose of 10 or 100 µg/mouse of TBI 304 and BMCs were examined earlier on day 7. TBI 304 provided a 3.5-fold increase in human CD34+ cells as well as a 1.8 to 6.7-fold increase in bone marrow erythroid lineage engraftment (huGlyA+, huCD36+ and huCD71+) and a 2-fold increase in colony-forming cells. The ability of TBI 304 to stimulate erythropoiesis in preclinical models led to the creation of an anti-CD163 mAb suitable for human clinical use. TBI 304H was generated by grafting the complementarity-determining regions derived from TBI 304 onto a humanized IgG4 framework without altering antigen specificity. An IgG4 framework, as an antibody without Fc effector function, was deemed the most suitable for an agonistic mAb. In the single dose, 7 day Hu-SCID model human CD34+ cells were mobilized from the mouse bone marrow by TBI 304H, as reflected by dose dependent decreases in huCD34+, huCD71+, and huGlyA+ cells in the mouse marrow. At the highest dose tested (500 µg/mouse) the decrease in human HPCs was similar to that found in animals administered Hb (2 mg/mouse). In this model, human hematopoiesis derived from the engrafted human CD34+ cells is not sustained and these date may reflect a mobilization of human HPCs through stimulation by an anti-CD163 antibody. Therapure has received U.S. FDA approval to conduct a Phase I trial of the novel therapeutic antibody TBI 304H. The Phase I clinical trial is a single-center, open-label, intra-subject escalating dose study, which will evaluate the safety, tolerability and pharmacokinetics of TBI 304H following administration to subjects experiencing chemotherapy-induced anemia.
Matthews:Therapure Biopharma: Employment. Eberle-Ayres:Therapure Biopharma: Employment. Lu:Therapure Biopharma: Employment. Singh:Therapure Biopharma: Employment. Cutler:Therapure Biopharma: Employment. Bell:Therapure Biopharma: Employment.
Asterisk with author names denotes non-ASH members.
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