Pharmacokinetics and microdistribution of polyethylene glycol-modified humanized A33 antibody targeting colon cancer xenografts

Therapeutic proteins have been conjugated with polyethylene glycol (PEGylat ion) to reduce immunogenicity and enhance circulating dose. Here we have in vestigated the effect of PEGylation on immunogenicity, pharmacokinetics, an d histologic microdistribution of tumor-targeting antibodies with humanized A33 antibody (huA33) as a model system. Conjugation of huA33 with methoxy- PEG of M-r 5,000 (32%-34% of primary amines modified) or M-r 20,000 (16%-18 % modification) preserved >50% of native huA33 binding to SW1222 colon canc er cells. In mice, both PEGylated forms cleared from serum moderately slowe r than native huA33, After repeated immunization with PEG-huA33, anti-antib ody titers in immunocompetent mice were <5% of those in huA33-treated contr ols. Both PEG-huA33 forms reached approx. 75% of the maximum tumor dose of huA33 in SW1222-xenografted mice, but their tumor:blood ratios were conside rably reduced. To demonstrate immunologic specificity of PEG-huA33 targetin g in SW1222 tumor-bearing mice, antigenic sites were presaturated by inject ing excess native huA33, This reduced subsequent uptake of PEG-huA33 by up to 80%, whereas presaturation with hu3S193 control antibody had no signific ant effect. To assess the microdistribution of antibody uptake in the same xenograft model, tumor tissue resected at different time points after antib ody administration was examined for human IgG by immunohistochemistry. Both PEG preparations achieved the same peak staining intensity and homogeneity as native huA33 with a delay of several hours. Given the measured reductio n in immunoreactivity in vitro, these results demonstrate that the tumor ta rgeting potential of huA33 in vivo is preserved at PEGylation levels suffic ient to suppress immunogenicity, Int. J. Cancer 87:382-390, 2000. (C) 2000 Wiley-Liss, Inc.