The chimeric
monoclonal antibody ch806 specifically targets the
tumor-associated mutant
epidermal growth factor receptor (de 2-7EGFR or
EGFRVIII) and is currently under investigation for its potential use in
cancer therapy. The humanised
monoclonal antibody hu3S193 specifically targets the Lewis Y epithelial
antigen and is currently in Phase I clinical trials in patients with advanced breast, colon, and ovarian
carcinomas. To assist the clinical evaluation of ch806 and hu3S193, laboratory assays are required to monitor their serum pharmacokinetics and quantitate any immune responses to the
antibodies. Mice immunized with ch806 or hu3S193 were used to generate hybridomas producing
antibodies with specific binding to ch806 or hu3S193 and competitive for
antigen binding. These
anti-idiotype antibodies (designated Ludwig Melbourne Hybridomas, LMH) were investigated as
reagents suitable for use as positive controls for HAHA or HACA analyses and for measuring hu3S193 or ch806 in human serum. Anti-idiotypes with the ability to concurrently bind two target antibody molecules were identified, which enabled the development of highly reproducible, sensitive, specific ELISA assays for determining serum concentrations of hu3S193 and ch806 with a 3 ng/mL limit of quantitation using LMH-3 and LMH-12, respectively. BIAcore analyses determined high apparent binding affinity for both idiotypes: LMH-3 binding immobilized hu3S193, Ka = 4.76 x 10(8) M(-1); LMH-12 binding immobilised ch806, Ka = 1.74 x 10(9) M(-1). Establishment of HAHA or HACA analysis of sera samples using BIAcore was possible using LMH-3 and LMH-12 as positive controls for quantitation of immune responses to hu3S193 or ch806 in patient sera. These anti-idiotypes could also be used to study the penetrance and binding of ch806 or hu3S193 to
tumor cells through immunohistochemical analysis of
tumor biopsies. The generation of
anti-idiotype antibodies capable of concurrently binding a target antibody on each variable domain provides
reagents with high sensitivity for the assessment of safety and pharmacokinetic profiles of target
antibodies administered clinically.