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NMR-Based Activity Assays for Determining Compound Inhibition, IC50 Values, Artifactual Activity, and Whole-Cell Activity of Nucleoside Ribohydrolases
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Vinculación o flexión: distinción de los agonistas alostéricos de la Abl quinasa de los antagonistas mediante un
Wolfgang Jahnke1, Robert M Grotzfeld, Xavier Pellé
1Novartis Institutes for Biomedical Research, 4002 Basel, Switzerland. wolfgang.jahnke@novartis.com
Journal of the American Chemical Society
|May 11, 2010
Resumen
Se identificaron nuevos inhibidores Bcr-Abl dirigidos a la bolsa de miristato. La actividad funcional depende de inducir un cambio de conformación específico en helix_I, distinguiendo a los antagonistas de los agonistas en el tratamiento de la leucemia mieloide crónica (LMC).
Área de la Ciencia:
- La bioquímica es la bioquímica.
- Biología Molecular Biología Molecular
- Farmacología Farmacología.
Sus antecedentes:
- Los inhibidores alostéricos dirigidos a Bcr-Abl representan una estrategia terapéutica prometedora para la leucemia mieloide crónica (LMC).
- Se empleó el cribado basado en fragmentos para descubrir nuevos inhibidores de Abl que se unen al bolsillo de miristato.
Objetivo del estudio:
- Para investigar la actividad funcional de los ligandos de bolsillo de miristato.
- Para identificar los determinantes estructurales de la eficacia del inhibidor Bcr-Abl.
- Desarrollar un ensayo para el monitoreo de los estados de conformación Abl.
Principales métodos:
- Cribado basado en fragmentos para los inhibidores de Abl.
- Análisis conformacional basado en RMN para monitorear la hélice C-terminal.
- Pruebas bioquímicas para confirmar la activación de c-Abl.
Principales resultados:
- No todos los ligandos de bolsillo de miristato son inhibidores funcionales de Bcr-Abl.
- El estado conformacional de C-terminal helix_I es un determinante clave de la actividad funcional.
- Los ligandos que inducen la flexión helix_I son antagonistas funcionales; los que no inducen este cambio son agonistas de la quinasa.
- La activación del agonista alostérico de c-Abl se confirmó bioquímicamente.
Conclusiones:
- El estado conformacional de helix_I dicta el resultado funcional de la unión del ligando del bolsillo de miristato.
- Este ensayo conformacional proporciona un método para distinguir entre antagonistas y agonistas funcionales.
- Comprender estos mecanismos puede guiar el desarrollo de terapias más efectivas para la LMC.
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