The Wilms' tumor oncogene protein (WT1) is an intracellular, oncogenic transcription factor that is over-expressed in leukemias and a wide range of cancers. WT1 may be expressed in leukemia stem cells. RMFPNAPYL, “RMF”, a WT1-derived CD8 T cell epitope presented by HLA-A0201, is a validated target for T cell-based immunotherapy, used in multiple clinical trials of vaccines and cellular therapies. Therefore, we hypothesized that a “TCR-like antibody” specific for the WT1 peptide/HLA-A0201 complex might be an effective therapeutic agent. Using phage display technology, we generated 2 lead, high avidity (Kd < 0.2nM), fully human monoclonal antibodies (mAb) specific for the WT1 RMF peptide/HLA-A0201 complex. One version, ESK1, is a native human IgG1. A second version, ESKM, with enhanced antibody dependent human effector cell cytotoxicity (ADCC) function due to altered Fc glycolsylation was also prepared. ESK mAb bind to leukemia lines and other cancer cell lines, as well as primary leukemia cells that are both WT1+ and HLA-A0201+. In vitro, both ESK mAb mediated ADCC against CML cells, at concentrations below 3 ug/ml, but ESKM was about 8–10 fold more potent and could kill targets with far fewer peptide/MHC complexes expressed on the cell surface. At therapeutic doses of ESKM, there was no difference in biodistribution between the 2 mAb in C57BL6 mice or in mice that were transgenic for HLA-A0201. Low doses of ESK (25–100ug twice) effectively treated an established disseminated, ALL or human bcr/abl + lymphoid leukemias in a NSG mouse model (T, B and NK deficient) and prolonged survival. In mice, ESKM was slightly more effective than ESK1. An F(Ab')2 version of the antibody had no anti-tumor effect, indicating that an Fc-mediated mechanism plays the major role in therapy. When combined with a single infusion of human CD34-, CD3-, human NK and monocyte effectors in the NSG mice, therapeutic effects of the mAb were more pronounced and more durable. There was no therapeutic effect of either mAb on WT1 low/A0201 negative disseminated Daudi ALL in mice. There was no observed toxicity in HLA-A0201 transgenic mice at the therapeutic mAb dose and schedule. ESK mAb are potential therapeutic agents for ALL, CML, other leukemias and cancers over-expressing the WT1 oncoprotein. Its expression in early leukemia cells may allow for elimination of the progenitors. The data also provide proof of concept for developing therapeutic mAb targeting important intracellular oncogenic proteins.
Yan:Eureka: Employment, Equity Ownership. Liu:Eureka: Employment, Equity Ownership.
Asterisk with author names denotes non-ASH members.