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Antibodies pp 157-168 | Cite as

Intrabodies: Development and Application in Functional Genomics and Therapy

  • Alcide Barberis
  • Adrian Auf Der Maur
  • Kathrin Tissot
  • Peter Lichtlen

Abstract

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).

Keywords

Antibody Fragment Potential Drug Target Complementarity Determine Region Transcriptional Activation Domain Antibody Library 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. Aramburu, J., Yaffe, M. B., Lopez-Rodriguez, C., Cantley, L. C., Hogan, P. G., and Rao, A. (1999). Affinity-driven peptide selection of an NFAT inhibitor more selective than cyclosporin A. Science 285, 2129–2133.PubMedCrossRefGoogle Scholar
  2. Asoh, S., Ohsawa, I., Mori, T., Katsura, K., Hiraide, T., Katayama, Y., Kimura, M., Ozaki, D., Yamagata, K., and Ohta, S. (2002). Protection against ischemic brain injury by protein therapeutics. Proc Natl Acad Sci U S A 99, 17107–17112.PubMedCrossRefGoogle Scholar
  3. 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
  4. 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
  5. Bai, J., Sui, J., Zhu, R. Y., Tallarico, A. S., Gennari, F., Zhang, D., and Marasco, W. A. (2003). Inhibition of Tat-mediated transactivation and HIV-1 replication by human anti-hCyclinT1 intrabodies. J Biol Chem 278, 1433–1442.PubMedCrossRefGoogle Scholar
  6. Barberis, A., and Gaudreau, L. (1998). Recruitment of the RNA polymerase II holoenzyme and its implications in gene regulation. Biol Chem 379, 1397–1405.PubMedGoogle Scholar
  7. Barberis, A., Pearlberg, J., Simkovich, N., Farrell, S., Reinagel, P., Bamdad, C., Sigal, G., and Ptashne, M. (1995). Contact with a component of the polymerase II holoenzyme suffices for gene activation. Cell 81, 359–368.PubMedCrossRefGoogle Scholar
  8. Berger, C., Weber-Bornhauser, S., Eggenberger, J., Hanes, J., Pluckthun, A., and Bosshard, H. R. (1999). Antigen recognition by conformational selection. FEBS Lett 450, 149–153.PubMedCrossRefGoogle Scholar
  9. Biocca, S., Ruberti, F., Tafani, M., Pierandrei-Amaldi, P., and Cattaneo, A. (1995). Redox state of single chain Fv fragments targeted to the endoplasmic reticulum, cytosol and mitochondria. Bio/Technology 13, 1110–1115.PubMedCrossRefGoogle Scholar
  10. Carlson, J. R. (1988). A new means of inducibly inactivating a cellular protein. Mol Cell Biol 8, 2638–2646.PubMedGoogle Scholar
  11. 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
  12. Cattaneo, A., and Biocca, S. (1997). Intracellular antibodies: Development and applications (New York, Springer).Google Scholar
  13. Cattaneo, A., and Biocca, S. (1999). The selection of intracellular antibodies. Trends In Biotechnology 17, 115–121.PubMedCrossRefGoogle Scholar
  14. Derossi, D., Calvet, S., Trembleau, A., Brunissen, A., Chassaing, G., and Prochiantz, A. (1996). Cell internalization of the third helix of the Antennapedia homeodomain is receptor-independent. J Biol Chem 271, 18188–18193.PubMedCrossRefGoogle Scholar
  15. Ewert, S., Huber, T., Honegger, A., and Pluckthun, A. (2003). Biophysical properties of human antibody variable domains. J Mol Biol 325, 531–553.PubMedCrossRefGoogle Scholar
  16. Fields, S., and Sternglanz, R. (1994). The two-hybrid system: an assay for protein-protein interactions. Trends in Genetics 10, 286–292.PubMedCrossRefGoogle Scholar
  17. Hanes, J., and Pluckthun, A. (1997). In vitro selection and evolution of functional proteins by using ribosome display. Proc Natl Acad Sci U S A 94, 4937–4942.PubMedCrossRefGoogle Scholar
  18. Harris, B. (1999). Exploiting antibody-based technologies to manage environmental pollution. Trends Biotechnol 17, 290–296.PubMedCrossRefGoogle Scholar
  19. Hassanzadeh, G. G., De Silva, K. S., Dambly-Chaudiere, C., Brys, L., Ghysen, A., Hamers, R., Muyldermans, S., and De Baetselier, P. (1998). Isolation and characterization of single-chain Fv genes encoding antibodies specific for Drosophila Poxn protein. FEBS Lett 437, 75–80.CrossRefGoogle Scholar
  20. Hope, I. A., and Struhl, K. (1987). GCN4, a eukaryotic transcriptional activator protein, binds as a dimer to target DNA. Embo J 6, 2781–2784.PubMedGoogle Scholar
  21. Hudson, P. J., and Souriau, C. (2003). Engineered antibodies. Nat Med 9, 129–134.PubMedCrossRefGoogle Scholar
  22. Ihle, J. N. (2000). The challenges of translating knockout phenotypes into gene function. Cell 102, 131–134.PubMedCrossRefGoogle Scholar
  23. Jean, D., Tellez, C., Huang, S., Davis, D. W., Bruns, C. J., McConkey, D. J., Hinrichs, S. H., and Bar-Eli, M. (2000). Inhibition of tumor growth and metastasis of human melanoma by intracellular anti-ATF-1 single chain Fv fragment. Oncogene 19, 2721–2730.PubMedCrossRefGoogle Scholar
  24. Kamb, A., and Caponigro, G. (2001). Peptide inhibitors expressed in vivo. Curr Opin Chem Biol 5, 74–77.PubMedCrossRefGoogle Scholar
  25. Lander, E. S., Linton, L. M., Birren, B., Nusbaum, C., Zody, M. C., Baldwin, J., Devon, K., Dewar, K., Doyle, M., FitzHugh, W., et al. (2001). Initial sequencing and analysis of the human genome. Nature 409, 860–921.PubMedCrossRefGoogle Scholar
  26. Leder, L., Berger, C., Bornhauser, S., Wendt, H., Ackermann, F., Jelesarov, I., and Bosshard, H. R. (1995). Spectroscopic, calorimetric, and kinetic demonstration of conformational adaptation in peptide-antibody recognition. Biochemistry 34, 16509–16518.PubMedCrossRefGoogle Scholar
  27. Lener, M., Horn, I. R., Cardinale, A., Messina, S., Nielsen, U. B., Rybak, S. M., Hoogenboom, H. R., Cattaneo, A., and Biocca, S. (2000). Diverting a protein from its cellular location by intracellular antibodies. The case of p21Ras. Eur J Biochem 267, 1196–1205.PubMedCrossRefGoogle Scholar
  28. 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
  29. Marasco, W. A. (1997). Intrabodies: turning the humoral immune system outside in for intracellular immunization. Gene Ther 4, 11–15.PubMedCrossRefGoogle Scholar
  30. Martineau, P., Jones, P., and Winter, G. (1998). Expression of an antibody fragment at high levels in the bacterial cytoplasm. J Mol Biol 280, 117–127.PubMedCrossRefGoogle Scholar
  31. Mhashilkar, A. M., Bagley, J., Chen, S. Y., Szilvay, A. M., Heiland, D. G., and Marasco, W. A. (1995). Inhibition of HIV-1 Tat-mediated LTR transactivation and HIV-1 infection by anti-Tat single chain intrabodies. EMBO Journal 14, 1542–1551.PubMedGoogle Scholar
  32. Mössner, E., Koch, H., and Plückthun, A. (2001). Fast selection of antibodies without antigen purification: adaptation of the protein fragment complementation assay to select antigen-antibody pairs. J Mol Biol 308, 115–122.PubMedCrossRefGoogle Scholar
  33. 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
  34. Pörtner-Taliana, A., Russell, M., Froning, K. J., Budworth, P. R., Comiskey, J. D., and Hoeffler, J. P. (2000). In vivo selection of single-chain antibodies using a yeast two-hybrid system. J Immunol Methods 238, 161–172.PubMedCrossRefGoogle Scholar
  35. Ruhlmann, A., and Nordheim, A. (1997). Effects of the immunosuppressive drugs CsA and FK506 on intracellular signalling and gene regulation. Immunobiology 198, 192–206.PubMedCrossRefGoogle Scholar
  36. Schwarze, S. R., Ho, A., Vocero-Akbani, A., and Dowdy, S. F. (1999). In vivo protein transduction: delivery of a biologically active protein into the mouse. Science 285, 1569–1572.PubMedCrossRefGoogle Scholar
  37. Tavladoraki, P., Benvenuto, E., Trinca, S., De Martinis, D., Cattaneo, A., and Galeffi, P. (1993). Transgenic plants expressing a functional single-chain Fv antibody are specifically protected from virus attack. Nature 366, 469–472.PubMedCrossRefGoogle Scholar
  38. Venter, J. C., Adams, M. D., Myers, E. W., Li, P. W., Mural, R. J., Sutton, G. G., Smith, H. O., Yandell, M., Evans, C. A., Holt, R. A., et al. (2001). The sequence of the human genome. Science 291, 1304–1351.PubMedCrossRefGoogle Scholar
  39. Visintin, M., Settanni, G., Maritan, A., Graziosi, S., Marks, J. D., and Cattaneo, A. (2002). The Intracellular Antibody Capture Technology (I ACT): Towards a Consensus Sequence for Intracellular Antibodies. J Mol Biol 317, 73–83.PubMedCrossRefGoogle Scholar
  40. Visintin, M., Tse, E., Axelson, H., Rabbitts, T. H., and Cattaneo, A. (1999). Selection of antibodies for intracellular function using a two-hybrid in vivo system. Proc Natl Acad Sci U S A 96, 11723–11728.PubMedCrossRefGoogle Scholar
  41. Wadia, J. S., and Dowdy, S. F. (2002). Protein transduction technology. Curr Opin Biotechnol 13, 52–56.PubMedCrossRefGoogle Scholar
  42. 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
  43. Wilson, D. S., Keefe, A. D., and Szostak, J. W. (2001). The use of mRNA display to select high-affinity protein-binding peptides. Proc Natl Acad Sci U S A 98, 3750–3755.PubMedCrossRefGoogle Scholar
  44. 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
  45. Wörn, A., and Plückthun, A. (2001). Stability engineering of antibody single-chain Fv fragments. J Mol Biol 305, 989–1010.PubMedCrossRefGoogle Scholar
  46. 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

Copyright information

© Springer Science+Business Media New York 2004

Authors and Affiliations

  • Alcide Barberis
    • 1
  • Adrian Auf Der Maur
    • 1
  • Kathrin Tissot
    • 1
  • Peter Lichtlen
    • 1
  1. 1.ESBATech AG8952 Zürich-SchlierenSwitzerland

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