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 rece ntly described the capacity of the high affinity Fc gamma RI (CD64) to trigger cytotoxic activity of neutrophils (PMN) during granulocyte CS F (G-CSF) treatment, To take advantage of Fc gamma RI as a cytotoxic t rigger 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 media te 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 hu man serum IgG, thereby abrogating the enhanced Ch520C9-mediated cytoto xicity, BsAb 22x520C9, which binds Fc gamma RI outside the ligand bind ing domain, effectively recruits the cytotoxic potential of Fc gamma R I 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 antit umor Abs, The IgG blockade can be circumvented with anti-Fc gamma RI B sAbs, Using human Fc gamma RI transgenic mice we demonstrate that BsAb 22x520C9 is able to engage Fc gamma RI in vivo. BsAb 22x520C9 injecte d i.v. was readily detected on circulating PMN of G-CSF-treated transg enic animals, In addition, we showed that PMN remain ''armed'' with Bs Ab 22x520C9 during migration to inflammatory sites, and that after iso lation such PMN specifically lyse HER-2/neu-expressing tumor cells, Th ese results point to the possibility of targeting anti-Fc gamma RI BsA bs to G-CSF-primed PMN in vivo, endowing them with specific anti-tumor activity.