TherapeuticEfficacyofanFc-EnhancedTCR-likeAntibodyto the Intracellular WT1 Oncoprotein

Purpose: RMFPNAPYL (RMF), a Wilms’ tumor gene 1 (WT1)–derived CD8 T-cell epitope presented by HLA-A � 02:01, is a validated target for T-cell–based immunotherapy. We previously reported ESK1, a high avidity (Kd < 0.2 nmol/L), fully-human monoclonal antibody (mAb) specific for the WT1 RMF peptide/ HLA-A � 02:01 complex, which selectively bound and killed WT1 þ and HLA-A � 02:01 þ leukemia and solid tumor cell lines. Experimental Design: We engineered a second-generation mAb, ESKM, to have enhanced antibodydependent cell-mediated cytotoxicity (ADCC) function due to altered Fc glycosylation. ESKM was compared with native ESK1 in binding assays, in vitro ADCC assays, and mesothelioma and leukemia therapeutic models and pharmacokinetic studies in mice. ESKM toxicity was assessed in HLA-A � 02:01 þ transgenic mice. Results: ESK antibodies mediated ADCC against hematopoietic and solid tumor cells at concentrations below 1 mg/mL, but ESKM was about 5- to 10-fold more potent in vitro against multiple cancer cell lines. ESKM was more potent in vivo against JMN mesothelioma, and effective against SET2 AML and fresh ALL xenografts.ESKMhadashortenedhalf-life(4.9daysvs.6.5days),butanidenticalbiodistributionpatternin C57BL/6Jmice.AttherapeuticdosesofESKM,therewasnodifferenceinhalf-lifeorbiodistribution inHLAA � 02:01 þ transgenicmicecomparedwiththeparentstrain.Importantly,therapeuticdosesofESKMinthese mice caused no depletion of total WBCs or hematopoetic stem cells, or pathologic tissue damage. Conclusions:Thedataprovideproof ofconceptthatanFc-enhancedmAbcanimproveefficacy againsta low-density, tumor-specific, peptide/MHC target, and support further development of this mAb against an important intracellular oncogenic protein. Clin Cancer Res; 20(15); 4036–46. � 2014 AACR.