Poly(ethylene glycol)-conjugated anti-EGF receptor antibody C225 with radiometal chelator attached to the termini of polymer chains

Several biological barriers, including significant liver uptake, limit the clinical application of radiolabeled antibodies in radioimmunoscintigraphy. Here, a general approach is described for radiolabeling of monoclonal anti bodies conjugated with poly(ethylene glycol) (PEG). This strategy is demons trated with C225, a monoclonal antibody directed against epidermal growth f actor (EGF) receptor. We synthesized a heterofunctional PEG with one end at tached to a radiometal chelator, diethylenetriaminepentaacetic acid (DTPA), and the other end to a protected thiol group, S-acetylthioacetate. After a deprotection step, the resulting DTPA-PEG-SH was conjugated to maleimide-a ctivated C225 to yield DTPA-PEG-C225 conjugate. Characterization of DTPA-PE G-C225 with immunoprecipitation and Western blot analysis revealed that the conjugate was biologically active in binding to the EGF receptor in A431 c ells. Competitive EGF receptor binding assay in MDA-MB-468 cells showed tha t DTPA-PEG-C225, with up to 60% of the amino groups in C225 substituted, re tained 66% of C225's binding affinity. Moreover, DTPA-PEG-C225 with increas ing degrees of NH2 substitution from 20% to 70% retained the activity of C2 25 to induce apoptosis in DiFi cells. More importantly, DTPA-PEG-C225 demon strated less nonspecific interaction than DTPA-C225. Pharmacokinetic analys is using In-111-labeled compounds revealed narrower steady-state distributi on of In-111-DTPA-PEG-C225 than In-111-DTPA-C225, probably due to reduced n onspecific binding of PEG-modified antibody to tissues. The terminal half-l ife (t(1/2,gamma)) of In-111-DTPA-PEG-C225, 21.1 h, was shorter than that o f In-111-DTPA-C225, 52.9 h. These data suggest that In-111-DTPA-PEG-C225 ma y provide better imaging characteristics than In-111-DTPA-C225, and that us ing PEG as a linker between the monoclonal antibody and DTPA may be a promi sing strategy in optimizing the imaging characteristics of immunoscintigrap hic agents.