Generation of new cytotoxic human ribonuclease variants directed to the nucleus
| Autor: | Marc Ribó, Jessica Castro, Maria Vilanova, Santiago Ruiz-Martínez, Antoni Benito, Anna Vert, Pere Tubert, Diego Escribano |
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| Přispěvatelé: | Ministerio de Ciencia e Innovación (Espanya) |
| Rok vydání: | 2012 |
| Předmět: |
Nuclear Localization Signals
Pharmaceutical Science Antineoplastic Agents Apoptosis Cancer -- Treatment Medicaments antineoplàstics Jurkat Cells Ribonucleases Cell Line Tumor Drug Discovery Antineoplastic agents Humans Cytotoxic T cell Ribonuclease Cytotoxicity RNA Nuclear Cell Nucleus biology Immunogenicity Ribonuclease Pancreatic Protein engineering Molecular biology Recombinant Proteins Cell biology Mutation Cancer cell biology.protein Molecular Medicine Pancreatic ribonuclease Càncer -- Tractament Nuclear localization sequence HeLa Cells |
| Zdroj: | © Molecular Pharmaceutics, 2012, vol. 9, núm. 10, p. 2894-2902 Articles publicats (D-B) DUGiDocs – Universitat de Girona instname |
| Popis: | Ribonucleases are promising agents for use in anticancer therapy. Engineering a nuclear localization signal into the sequence of the human pancreatic ribonuclease has been revealed as a new strategy to endow this enzyme with cytotoxic activity against tumor cells. We previously described a cytotoxic human pancreatic ribonuclease variant, named PE5, which is able to cleave nuclear RNA, inducing the apoptosis of cancer cells and reducing the amount of P-glycoprotein in different multidrug-resistant cell lines. These results open the opportunity to use this ribonuclease in combination with other chemotherapeutics. In this work, we have investigated how to improve the properties of PE5 as an antitumor drug candidate. When attempting to develop a recombinant protein as a drug, two of the main desirable attributes are minimum immunogenicity and maximum potency. The improvements of PE5 have been designed in both senses. First, in order to reduce the potential immunogenicity of the protein, we have studied which residues mutated on PE5 can be reverted to those of the wild-type human pancreatic ribonuclease sequence without affecting its cytotoxicity. Second, we have investigated the effect of introducing an additional nuclear localization signal at different sites of PE5 in an effort to obtain a more cytotoxic enzyme. We show that the nuclear localization signal location is critical for the cytotoxicity. One of these variants, named NLSPE5, presents about a 10-fold increase in cytotoxicity respective to PE5. This variant induces apoptosis and kills the cells using the same mechanism as PE5 This work has been supported by Grant BFU2009-06935 from MICINN (Spain) and by Grant GRCT04 from the University of Girona. We are very grateful to Dr. Milica Pesic and Dr. Sabera Ruzdijićfor providing us with the NCI-H460/R cell line and to Dr. Ramon Colomer for providing us with the NCI/ ADR-RES cell line. A.V., J.C., and P.T. acknowledge their fellowship from MINEDU, Universitat de Girona, and MEC (Spain), respectively |
| Databáze: | OpenAIRE |
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