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Schier R , Marks JD , Wolf EJ , Apell G , Wong C , McCartney JE , Bookman MA , Huston JS , Houston LL , Weiner LM , Adams GP
In-Vitro and in-Vivo Characterization of a Human Anti-C-Erbb-2 Single-Chain Fv Isolated from a Filamentous Phage Antibody Library
Immunotechnology. 1995 May;1(1) :73-81
Abstract`Background: Antibody-based reagents have failed to live up to their anticipated role as highly specific targeting agents for cancer therapy. Targeting with human single-chain Fv (sFv) molecules may overcome some of the limitations of murine IgG, but are difficult to produce with conventional hybridoma technology. Alternatively, phage display of antibody gene repertoires can be used to produce human sFv. Objectives: To isolate and characterize human single chain Fvs which bind to c-erB-2, an oncogene product overexpressed by 30-50% of breast carcinomas and other adenocarcinomas. Study design: A non- immune human single-chain Fv (C6.5) was isolated which binds specifically to c-erB-2. C6.5 is entirely human in sequence, expresses at high level as native protein in E. coli, and is easily purified in high yield in two steps. C6.5 is entirely human in sequence, expresses at high level as native protein in E. coli, and is easily purified in high yield in two steps. 6.5 binds to immobilized c-erB-2 extracellular domain with a Kd of 1.6 x 10(-8) M and to c-erb-2 on SK-OV-3 cells with a K-d of 2.0 x 10(-8) M, an affinity that is similar to sFv produced against the same antigen from hybridomas. Biodistribution studies demonstrate. 1.47% injected dose/g tumor 24 h after injection of I-125-C6.5 into scid mice bearing SK-OV-3 tumors. Tumor:normal organ ratios range from 8.9:1 for kidney to 283:1 for muscle. Conclusion: These results are the first in vivo biodistribution studies using an sFv isolated from a non-immune human repertoire and confirm the specificity of sFv produced in this manner. The use of phage display to produce C6.5 mutants with higher affinity and slower K-off would permit rigorous evaluation of the role of antibody affinity and binding kinetics in tumor targeting and could result in the production of a therapeutically useful targeting protein for radioimmunotherapy and other applications.
NotesTimes Cited: 51 Article TF482 IMMUNOTECHNOLOGY