Intrabodies: Development and Application in Functional Genomics and Therapy
The human genome project has led to the identification of a large number of genes and respective proteins, thus providing the pharmaceutical industry with thousands of new potential drug targets, most of which function in intracellular compartments (Lander et al., 2001; Venter et al., 2001). This fact opens new perspectives for therapy of human diseases; however, it also demands reliable approaches for understanding the role and the function of these new genes and proteins (functional genomics) and for identifying those that can be validated as drug targets (target validation). Different experimental tools are currently used for investigating the function of these new intracellular proteins, including their potential role in disease, and for evaluating them as potential drug targets. The classical way to investigate the function of genes, and thereby determine the physiological and pathological relevance of gene products, is to interfere with their expression (Ihle, 2000). Approaches such as gene knockout, antisense oligonucleotide or RNA interference (RNAi) are currently used to study gene and protein function and to validate candidate drug targets by analysing the effects of their deletion. One limitation of all these techniques is that they eliminate all functions of a target gene product at once, thus making it difficult to dissect potentially distinct roles of different domains and to mimic the effects of a small molecule that presumably will act at a specific domain of the protein (Kamb and Caponigro, 2001).
KeywordsMigration Ischemia Disulfide Cyclosporine Cytosol
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- Auf der Maur, A., Escher, D., and Barberis, A. (2001). Antigen-independent selection of stable intracellular single-chain antibodies. FEBS Lett 508, 407–412.Google Scholar
- Auf der Maur, A., Zahnd, C., Fischer, F., Spinelli, S., Honegger, A., Cambillau, C., Escher, D., Pluckthun, A., and Barberis, A. (2002). Direct in vivo screening of intrabody libraries constructed on a highly stable single-chain framework. J Biol Chem 277, 45075–45085.Google Scholar
- Caron de Fromentel, C., Gruel, N., Venot, C., Debussche, L., Conseiller, E., Dureuil, C., Teillaud, J. L., Tocque, B., and Bracco, L. (1999). Restoration of transcriptional activity of p53 mutants in human tumour cells by intracellular expression of anti-p53 single chain Fv fragments. Oncogene 18, 551–557.Google Scholar
- Cattaneo, A., and Biocca, S. (1997). Intracellular antibodies: Development and applications (New York, Springer).Google Scholar
- Lichtlen, P., Auf der Maur, A., and Barberis, A. (2002). Target validation through protein-domain knockout: applications of intracellularly stable single-chain antibodies. TARGETS 1, 37–44.Google Scholar
- Nagahara, H., Vocero-Akbani, A. M., Snyder, E. L., Ho, A., Latham, D. G., Lissy, N. A., Becker-Hapak, M., Ezhevsky, S. A., and Dowdy, S. F. (1998). Transduction of full-length TAT fusion proteins into mammalian cells: TAT-p27Kipl induces cell migration. Nat Med 4, 1449–1452.PubMedCrossRefGoogle Scholar
- Willuda, J., Honegger, A., Waibel, R., Schubiger, P. A., Stahel, R., Zangemeister-Wittke, U., and Plückthun, A. (1999). High thermal stability is essential for tumor targeting of antibody fragments: engineering of a humanized anti-epithelial glycoprotein-2 (epithelial cell adhesion molecule) single-chain Fv fragment. Cancer Res 59, 5758–5767.PubMedGoogle Scholar
- Wörn, A., Auf der Maur, A., Escher, D., Honegger, A., Barberis, A., and Plückthun, A. (2000). Correlation between in vitro stability and in vivo performance of anti-GCN4 intrabodies as cytoplasmic inhibitors. J Biol Chem 275, 2795–2803.Google Scholar
- Zhu, Q., Zeng, C., Huhalov, A., Yao, J., Turi, T. G., Danley, D., Hynes, T., Cong, Y., DiMattia, D., Kennedy, S., et al. (1999). Extended half-life and elevated steady-state level of a single-chain Fv intrabody are critical for specific intracellular retargeting of its antigen, caspase-7. J Immunol Methods 231, 207–222.PubMedCrossRefGoogle Scholar