Generation of HER-2/neu-specific cytotoxic neutrophils in vivo: Efficient arming of neutrophils by combined administration of granulocyte colony-stimulating factor and Fc gamma receptor I bispecific antibodies

Abs are able to induce inflammatory antitumor responses by recruiting IgG Fc receptor (Fc gamma R)-bearing cytotoxic effector cells. We recently described the capacity of the high affinity Fc gamma RI (CD64) to trigger cytotoxic activity of neutrophils (PMN) during granulocyte CSF (G-CSF) treatment. To take advantage of Fc gamma RI as a cytotoxic trigger molecule on PMN, two Ab constructs were prepared. We show that a chimeric human IgG1 Ab (Ch520C9) and an anti-Fc gamma RI bispecific Ab (BsAb; 22x520C9), both directed to the proto-oncogene product HER-2/neu, interact with Fc gamma RI. In addition, both Ab constructs mediate enhanced lysis of HER-2/neu-expressing tumor cells by G-CSF-primed PMN. However, engagement of Fc gamma RI by Ch520C9 was inhibited by human serum IgG, thereby abrogating the enhanced Ch520C9-mediated cytotoxicity. BsAb 22x520C9, which binds Fc gamma RI outside the ligand binding domain, effectively recruits the cytotoxic potential of Fc gamma RI on G-CSF-primed PMN regardless of the presence of human serum. These results indicate that under physiologic conditions, serum IgG impairs activation of Fc gamma RI-mediated cytotoxicity by conventional antitumor Abs. The IgG blockade can be circumvented with anti-Fc gamma RI BsAbs. Using human Fc gamma RI transgenic mice we demonstrate that BsAb 22x520C9 is able to engage Fc gamma RI in vivo. BsAb 22x520C9 injected i.v. was readily detected on circulating PMN of G-CSF-treated transgenic animals. In addition, we showed that PMN remain "armed" with BsAb 22x520C9 during migration to inflammatory sites, and that after isolation such PMN specifically lyse HER-2/neu-expressing tumor cells. These results point to the possibility of targeting anti-Fc gamma RI BsAbs to G-CSF-primed PMN in vivo, endowing them with specific anti-tumor activity.