Construcción de un vector de expresión derivado de virus adenoasociados para corregir in vitro el defecto genético de la enfermedad de Morquio A

Luis Alejandro Barrera, Mónica A. Gutiérrez, Felipe García Vallejo, Shunji Tomatsu, Flavio Cerón, Carlos J. Alméciga Díaz, Martha C. Domínguez, .

Palabras clave: mucopolisacaridosis IV A [genética], dependovirus, terapia de gen, técnicas de transferencia de gen, cultivo de virus, técnicas de cultivo

Resumen

Introducción. La mucopolisacaridosis IV A (Morquio A) es una enfermedad de depósito lisosómico causada por la deficiencia en la actividad de la enzima N-acetil-galactosamina- 6-sulfato-sulfatasa que produce la acumulación intralisosómica de queratán y condroitín-6-sulfato. Hasta el momento, su manejo es paliativo, por lo que las investigaciones se han enfocado en establecer una terapia que pueda aplicarse tempranamente y garantice la expresión estable de la enzima. En este sentido, la terapia génica se presenta como una de las potenciales alternativas terapéuticas para corregir el defecto genético en la mucopolisacaridosis IV A. Objetivo. Construir vectores de expresión derivados de virus adenoasociados para corregir in vitro la deficiencia enzimática en la mucopolisacaridosis IV A. Materiales y métodos. Se produjeron vectores derivados de virus adenoasociados que portaban el gen humano de la enzima N-acetil-galactosamina-6-sulfato-sulfatasa dirigido por el promotor temprano del citomegalovirus humano, empleando un sistema libre de adenovirus. Se transfectaron células HEK293 y fibroblastos humanos Morquio A con los virus recombinantes, y se determinó la actividad enzimática en el lisado celular a las 24 y 48 horas después de la transfección. Resultados. Se obtuvieron virus adenoasociados recombinantes, libres de adenovirus, con títulos hasta de 2,08 x 1010 cápsides/ml. Tanto en células HEK293 como en fibroblastos Morquio A transfectados, se obtuvieron actividades enzimáticas hasta de 3,05 nmoles/mg por hora, 48 horas después de la transfección. Conclusión. Los virus recombinantes producidos expresaron in vitro la enzima GALNS en las células transfectadas. Estos resultados constituyen el paso inicial para el desarrollo de una terapia génica para la enfermedad de Morquio A empleando vectores derivados de virus adenoasociados.

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  • Luis Alejandro Barrera Instituto de Errores Innatos del Metabolismo, Pontificia Universidad Javeriana, Bogotá, D.C., Colombia
  • Mónica A. Gutiérrez Instituto de Errores Innatos del Metabolismo, Pontificia Universidad Javeriana, Bogotá, D.C., Colombia Department of Pediatrics, Saint Louis University, Saint Louis, MO, United States R. I. P.
  • Felipe García Vallejo Laboratorio de Biología Molecular, Facultad de Salud, Universidad del Valle, Cali, Colombia
  • Shunji Tomatsu Department of Pediatrics, Saint Louis University, Saint Louis, MO, United States
  • Flavio Cerón Laboratorio de Biología Molecular, Facultad de Salud, Universidad del Valle, Cali, Colombia
  • Carlos J. Alméciga Díaz Instituto de Errores Innatos del Metabolismo, Pontificia Universidad Javeriana, Bogotá, D.C., Colombia
  • Martha C. Domínguez Laboratorio de Biología Molecular, Facultad de Salud, Universidad del Valle, Cali, Colombia

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Cómo citar
Barrera, L. A., Gutiérrez, M. A., García Vallejo, F., Tomatsu, S., Cerón, F., Alméciga Díaz, C. J., & Domínguez, M. C. (2008). Construcción de un vector de expresión derivado de virus adenoasociados para corregir in vitro el defecto genético de la enfermedad de Morquio A. Biomédica, 28(3), 448-459. https://doi.org/10.7705/biomedica.v28i3.85
Publicado
2008-09-01
Sección
Nota técnica