A Chemo-enzymatically Linked Bispecific Antibody Retargets T Cells to a Sialylated Epitope on CD43 in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) is a high-risk disease with a poor prognosis, particularly in elderly patients. Because current AML treatment relies primarily on untargeted therapies with severe side effects that limit patient eligibility, identification of novel therapeutic AML targets is highly desired. We recently described AT1413, an antibody produced by donor B cells of a patient with AML cured after allogeneic hematopoietic stem cell transplantation. AT1413 binds CD43s, a unique sialylated epitope on CD43, which is weakly expressed on normal myeloid cells and overexpressed on AML cells. Because of its selectivity for AML cells, we considered CD43s as a target for a bispecific T-cell-engaging antibody (bTCE) and generated a bTCE by coupling AT1413 to two T-cell-targeting fragments using chemo-enzymatic linkage. In vitro, AT1413 bTCE efficiently induced T-cell-mediated cytotoxicity toward different AML cell lines and patient-derived AML blasts, whereas endothelial cells with low binding capacity for AT1413 remained unaffected. In the presence of AML cells, AT1413 bTCE induced upregulation of T-cell activation markers, cytokine release, and T-cell proliferation. AT1413 bTCE was also effective in vivo. Mice either coinjected with human peripheral blood mononuclear cells or engrafted with human hematopoietic stem cells [human immune system (HIS) mice] were inoculated with an AML cell line or patient-derived primary AML blasts. AT1413 bTCE treatment strongly inhibited tumor growth and, in HIS mice, had minimal effects on normal human hematopoietic cells. Taken together, our results indicate that CD43s is a promising target for T-cell-engaging antibodies and that AT1413 holds therapeutic potential in a bTCE-format. SIGNIFICANCE: These findings offer preclinical evidence for the therapeutic potential of a bTCE antibody that targets a sialylated epitope on CD43 in AML.

AML-specific cytotoxic antibodies in patients with durable graft-versus-leukemia responses.

Most patients with acute myeloid leukemia (AML) can only be cured when allogeneic hematopoietic stem-cell transplantation induces a graft-versus-leukemia immune response (GVL). Although the role of T cells and natural killer cells in tumor immunology has been established, less is known about the contribution of B cells. From B cells of high-risk patients with AML with potent and lasting GVL responses, we isolated monoclonal antibodies directed against antigens expressed on the cell surface of AML cells but not on normal hematopoietic and nonhematopoietic cells. A number of these donor-derived antibodies recognized the U5 snRNP200 complex, a component of the spliceosome that in normal cells is found in the cell. In AML however, the U5 snRNP200 complex is exposed on the cell membrane of leukemic blasts. U5 snRNP200 complex-specific antibodies induced death of AML cells in an Fc receptor-dependent way in the absence of cytotoxic leukocytes or complement. In an AML mouse model, treatment with U5 snRNP200 complex-specific antibodies led to significant tumor growth inhibition. Thus, donor-derived U5 snRNP200 complex-recognizing AML-specific antibodies may contribute to antitumor responses.

Patient-derived antibody recognizes a unique CD43 epitope expressed on all AML and has antileukemia activity in mice.

Immunotherapy has proven beneficial in many hematologic and nonhematologic malignancies, but immunotherapy for acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) is hampered by the lack of tumor-specific targets. We took advantage of the tumor-immunotherapeutic effect of allogeneic hematopoietic stem cell transplantation and searched the B-cell repertoire of a patient with a lasting and potent graft-versus-AML response for the presence of AML-specific antibodies. We identified an antibody, AT1413, that was of donor origin and that specifically recognizes a novel sialylated epitope on CD43 (CD43s). Strikingly, CD43s is expressed on all World Health Organization 2008 types of AML and MDS. AT1413 induced antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity of AML cells in vitro. Of note, AT1413 was highly efficacious against AML cells in a humanized mouse model without affecting nonmalignant human myeloid cells, suggesting AT1413 has potential as a therapeutic antibody.

Simultaneous measurement of RBC velocity, flux, hematocrit and shear rate in vascular networks.

Not all tumor vessels are equal. Tumor-associated vasculature includes immature vessels, regressing vessels, transport vessels undergoing arteriogenesis and peritumor vessels influenced by tumor growth factors. Current techniques for analyzing tumor blood flow do not discriminate between vessel subtypes and only measure average changes from a population of dissimilar vessels. We developed methodologies for simultaneously quantifying blood flow (velocity, flux, hematocrit and shear rate) in extended networks at single-capillary resolution in vivo. Our approach relies on deconvolution of signals produced by labeled red blood cells as they move relative to the scanning laser of a confocal or multiphoton microscope and provides fully resolved three-dimensional flow profiles within vessel networks. Using this methodology, we show that blood velocity profiles are asymmetric near intussusceptive tissue structures in tumors in mice. Furthermore, we show that subpopulations of vessels, classified by functional parameters, exist in and around a tumor and in normal brain tissue.