Biophysical characterization and identification of bioactive compounds against the pharmacological target mecp2 and evaluation of drug nanocarriers

• Autores: Rafael Claveria Gimeno
• Directores de la Tesis: Olga Abian Franco (dir. tes.), Adrian Velazquez Campoy (dir. tes.)
• Lectura: En la Universidad de Zaragoza ( España ) en 2017
• Idioma: español
• Tribunal Calificador de la Tesis: Juan Iovanna (presid.), Teresa Sierra Travieso (secret.), Manel Esteller Badosa (voc.)
• Programa de doctorado: Programa de Doctorado en Bioquímica y Biología Molecular por la Universidad de Zaragoza
• Materias:
  • Química
    • Bioquímica
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• Resumen

  • Rett syndrome, a rare neurodevelopmental disease causing mental, motor and growth impairment, the leading cause of intellectual retardation in girls, is linked directly to mutations in the methyl CpG binding protein 2 (MeCP2). MeCP2, an intrinsically disordered protein, is a DNA binding protein involved in epigenetic regulation and chromatin remodeling that preferentially interacts with methylated DNA regions, though it also interacts with multiple protein partners. Unstructured regions are important because they provide the required structural plasticity for establishing multiple interactions with different binding partners through processes where partial folding and binding are coupled, resulting in allosteric regulation.

    This thesis includes a biophysical characterization of the structural stability and the interaction between MeCP2 and the DNA employing spectroscopic and calorimetric techniques, contributing to increase the knowledge about MeCP2 intracellular functions, and providing the tools to assess the deleterious effects caused by Rett Syndrome mutations and allowing us identifying small chemical compounds able to stabilize the interaction and, potentially, to reverse these damaging effects.

    In vitro primary screening for identifying bioactive compounds (activators or pharmacological chaperones) against MeCP2-dsDNA interaction led to the discovery of 14 hit compounds that must be tested in cell-based efficacy secondary screenings. Frequently hit compounds do not succeed due to an apparent low potency in cell assays, despite an excellent performance in primary screening. Cellular internalization and intracellular metabolism are some of the difficulties the compounds must confront in cell/organism assays, and different strategies can be envisaged for minimizing that problem.

    Some of these strategies have been used to improve the cellular internalization of compounds identified against a previously well characterized protein target in our research group, the hepatitis C virus NS3 protease. Some biopolymers and cyclodextrins have been employed as molecular nanocarriers to overcome the mentioned obstacles and enhance the therapeutic effect of the compounds.