Humanization of an Anti-CCR4 Antibody That Kills Cutaneous T-Cell Lymphoma Cells and Abrogates Suppression by T-Regulatory Cells

Cutaneous T-cell lymphoma (CTCL) is a heterogeneous group of neoplastic disorders characterized by clonallyderivedandskin-homingmalignantTcellsthatexpresshighlevelofchemokinereceptorCCR4,which is associated with their skin-homing capacity. CCR4 is also highly expressed on T-regulatory cells (Tregs) that can migrate to several different types of chemotactic ligand CCL17- and CCL22-secreting tumors to facilitate tumor cell evasion from immune surveillance. Thus, its high-level expression on CTCL cells and Tregs makes CCR4 a potential ideal target for antibody-based immunotherapy for CTCL and other types of solid tumors. Here, we conducted humanization and affinity optimization of a murine anti-CCR4 monoclonal antibody (mAb), mAb1567, that recognizes both the N-terminal and extracellular domains of CCR4 with high affinity and inhibits chemotaxis of CCR4 þ CTCL cells. In a mouse CTCL tumor model, mAb1567 exhibited a potent antitumor effect andin vitromechanistic studies showed that both complement-dependent cytotoxicity (CDC) and neutrophil-mediated antibody-dependent cellular cytotoxicity (ADCC) likely mediated this effect. mAb1567 also exerts human NK cell‐mediated ADCC activity in vitro. Moreover, mAb1567 also effectively inhibits chemotaxis of CD4 þ CD25 high Tregs via CCL22 and abrogates Treg suppression activity in vitro .A n affinity-optimized variant of humanized mAb1567, mAb2-3, was selected for further preclinical development basedonitshigherbindingaffinityandmorepotentADCCandCDCactivities.Takentogether,thishigh-affinity humanized mAb2-3 with potent antitumor effect and a broad range of mechanisms of action may provide a novel immunotherapy for CTCL and other solid tumors. Mol Cancer Ther; 11(11); 2451–61. � 2012 AACR.