Los vectores herpesvirales portadores de genes completos aseguran la persistencia de la expresión génica a largo plazo en ensayos de terapia génica en modelos animales.
Algunas enfermedades hereditarias están causadas por mutaciones, denominadas “recesivas”, que causan una menor producción de una proteína o la producción de una proteína defectuosa, incapaz de realizar correctamente su función. La terapia génica pretende curar este tipo de enfermedades mediante la introducción de los genes “sanos” que así pueden sustituir la función de los genes “defectuosos”. Para introducir dichos genes sanos en las células afectadas por la enfermedad se necesitan vehículos adecuados, los denominados “vectores”, que, en la mayoría de los casos son virus desprovistos de sus componentes más patogénicos.
El éxito de este tipo de terapia génica depende de una adecuada distribución del gen sano a un número suficiente de células afectadas así como de asegurar la persistencia de la expresión del gen “sano” en estas células. Esto último ha resultado ser particularmente más difícil de lo que en principio cabía esperar. Así pues, en numerosas ocasiones, el gen “sano” que se había introducido en una célula, al cabo del tiempo, se quedaba “silencioso”, es decir, dejaba de dar las instrucciones para la correcta producción de la proteína, con lo que la enfermedad nuevamente regresaba. Se cree que este “silenciamiento” puede deberse a la utilización de una serie de elementos “artificiales”: en primer lugar, no se utilizaba el gen de verdad sino una versión simplificada del mismo, el llamado cDNA, que contiene la información codificante de la proteína pero carece de muchos elementos requeridos para la regulación de su producción, y, en segundo lugar, se utilizaba un “promotor” de origen viral para asegurar la expresión de dicho gen. Parece ser que en el organismo, al cabo del tiempo, estos elementos son de alguna manera reconocidos como “extraños” y se produce su “silenciamiento”. Como alternativa a la utilización de estos elementos cabe recurrir a la versión natural del gen, en toda su amplitud. Esto es técnicamente más complicado, ya que los genes son normalmente muy grandes, y no caben en la mayoría de los vectores virales que son utilizados normalmente en los ensayos de terapia génica (como adenovirus, retrovirus y lentivirus). Una excepción interesante son los vectores herpesvirales, derivados del virus herpes simplex (HSV-1), capaces de acomodar hasta 150 kb de material genético.
Los grupos de Javier Diaz-Nido y Filip Lim, de la Universidad Autónoma de Madrid, en colaboración con el grupo de Richard Wade-Martins, de la Universidad de Oxford en el Reino Unido, hemos estado explorando la posibilidad de utilizar este tipo de vectores herpesvirales para la terapia génica de la ataxia de Friedreich (una enfermedad causada por la deficiencia de una proteína denominada frataxina, y que se caracteriza por la degeneración de ciertas neuronas en el sistema nervioso central, así como por otras alteraciones en el corazón y en el páncreas).
En un estudio publicado en 2007 demostramos que un vector herpesviral portador del gen completo de la frataxina era capaz de suplir las deficiencias funcionales encontradas en las células de la piel de los pacientes con ataxia de Friedreich (1).
Ahora, en un trabajo que se acaba de publicar “online” en la revista “Gene Therapy” (2) hemos podido comprobar que los vectores portadores del gen completo de la frataxina permiten una expresión persistente “in vivo” después de ser inyectados en el cerebelo de ratones. Estos resultados refuerzan la hipótesis de que los vectores portadores de genes completos no son silenciados y, en consecuencia, pueden ser muy útiles para una terapia génica a largo plazo. Otra ventaja de este tipo de vectores es que persisten como episomas estables en los núcleos de las células, no integrándose en ningún cromosoma, de manera que no causan alteraciones en el genoma de dichas células (a diferencia de lo que sucede con otros vectores). A la vista de estos datos, consideramos que los vectores herpesvirales portadores de genes completos pueden ser una herramienta muy segura y eficaz para la terapia de las enfermedades hereditarias causadas por mutaciones recesivas (como la ataxia de Friedreich).
1.- Gomez-Sebastian S, Gimenez-Cassina A, Diaz-Nido J, Lim F, Wade-Martins R.
Infectious delivery and expression of a 135 kb human FRDA genomic DNA locus complements Friedreich's ataxia deficiency in human cells.
Mol Ther. 2007 Feb;15(2):248-54.
http://www.ncbi.nlm.nih.gov/pubmed/17235301
http://www.nature.com/mt/journal/v15/n2/full/6300021a.html
2.- Gimenez-Cassina A, Wade-Martins R, Gomez-Sebastian S, Corona JC, Lim F, Diaz-Nido J.
Infectious delivery and long-term persistence of transgene expression in the brain by a 135-kb iBAC-FXN genomic DNA expression vector.
Gene Ther. 2011 Apr 14. doi:10.1038/gt.2011.45 [Epub ahead of print]
http://www.ncbi.nlm.nih.gov/pubmed/21490681
http://www.nature.com/gt/journal/vaop/ncurrent/full/gt201145a.html
Wednesday, April 27, 2011
Herpes viral vectors carrying complete genes ensure long-term gene expression persistence in animal models of gene therapy trials.
Press release: Herpes viral vectors carrying complete genes ensure long-term gene expression persistence in animal models of gene therapy trials.
Some hereditary diseases are caused by "recessive" mutations which cause either a decrease in the amount of a protein or the production of a defective protein, which is unable to properly perform its function. Gene therapy aims to cure such diseases through the introduction of "healthy" genes allowing the functional replacement of the "defective" genes. In order to introduce such healthy genes into the cells affected by the disease, it is necessary to use appropriate carriers, known as "vectors", which, in most cases are viruses devoid of their more pathogenic components.
The success of this type of gene therapy depends on both the proper distribution of the healthy gene to a sufficient number of affected cells and the persistence of the expression of the "healthy" gene in these cells. To ensure a long-term expression of the “healthy” gene has been a goal more difficult to attain than was anticipated. Thus, on many occasions, the "healthy" gene that had been introduced into a cell, as time went by, became "silent", i.e., it failed to give the instructions for the proper production of the protein, so the disease returned again. It is believed that this "silencing" may be due to the use of a number of "artificial" elements: first, the complete gene was not really used but a simplified version, the so-called cDNA, which contains the protein-coding information but lacks many elements required for the regulation of its production, and, second, a "promoter" of viral origin was used to ensure the expression of this gene. It seems that in the body, over time, these elements are in some way recognized as "foreign” and its “silencing” is triggered. As an alternative to the use of these elements, one could resort to the natural version of the gene, in all its length. This is technically more complicated because the genes are normally very large and do not fit into most viral vectors which are normally used in gene therapy trials (such as adenoviruses , retroviruses and lentiviruses). An interesting exception are herpes viral vectors, derived from herpes virus simplex (HSV-1), which are capable of accommodating up to 150 kb of genetic material.
Javier Diaz-Nido and Filip Lim Groups , at the Universidad Autónoma de Madrid in Spain, in collaboration with the Group of Richard Wade-Martins, of the University of Oxford in the United Kingdom, have been exploring the possibility of using this type of Herpes viral vectors for Friedreich's ataxia gene therapy (a disease caused by the deficiency of one protein called frataxin, and that is characterized by the degeneration of certain neurons in the central and peripheral nervous systems, as well as other alterations in the heart and the pancreas).
In a paper published in 2007 they showed that a herpes viral vector carrying the complete frataxin gene was able to make up for the functional deficiencies that were found in cultured skin cells of patients with Friedreich's ataxia (1).
Now, in a paper just published "online" in the "Gene Therapy" magazine (2) they have been able to demonstrate that vectors carrying the full frataxin gene allow a persistent expression "in vivo" after being injected into the mouse cerebellum. These results reinforce the view that vectors carrying complete genes are not silenced and may therefore be very useful for long term gene therapy. Another advantage of this type of herpes viral vectors is that they persist as stable “episomes” in the nuclei of cells, not integrated into any chromosome, and so they do not cause any alterations in the genome of these cells (unlike what happens with other vectors). Furthermore, herpes viral vectors are well known for their effective targeting of neuronal cells. In view of these data, it appears that herpes viral vectors carrying complete genes may be a safe and effective tool for the therapy of neurological hereditary diseases which are caused by recessive mutations (such as Friedreich's ataxia).
1.-Gomez-Sebastian S, Gimenez-Cassina A, Diaz-Nido J, Lim F, Wade-Martins R.
Infectious delivery and expression of 135 KB FRDA human genomic DNA locus complements Friedreich's ataxia deficiency in human cells.
MOL Ther. 2007 Feb; 15 (2): 248-54.
http://www.ncbi.NLM.NIH.gov/PubMed/17235301
http://www.nature.com/MT/journal/V15/N2/full/6300021a.html
2.-Gimenez-Cassina A, Wade-Martins R, Gomez-Sebastian S, Corona JC, Lim F, Diaz-Nido J.
Infectious delivery and long-term persistence of transgene expression in the brain by a 135-kb iBAC-FXN genomic DNA expression vector.
Gene Ther. 2011 Apr 14. DOI:10.1038/gt.2011.45 [Epub ahead of print]
http://www.ncbi.NLM.NIH.gov/PubMed/21490681
http://www.nature.com/gt/journal/vaop/ncurrent/full/gt201145a.html
Some hereditary diseases are caused by "recessive" mutations which cause either a decrease in the amount of a protein or the production of a defective protein, which is unable to properly perform its function. Gene therapy aims to cure such diseases through the introduction of "healthy" genes allowing the functional replacement of the "defective" genes. In order to introduce such healthy genes into the cells affected by the disease, it is necessary to use appropriate carriers, known as "vectors", which, in most cases are viruses devoid of their more pathogenic components.
The success of this type of gene therapy depends on both the proper distribution of the healthy gene to a sufficient number of affected cells and the persistence of the expression of the "healthy" gene in these cells. To ensure a long-term expression of the “healthy” gene has been a goal more difficult to attain than was anticipated. Thus, on many occasions, the "healthy" gene that had been introduced into a cell, as time went by, became "silent", i.e., it failed to give the instructions for the proper production of the protein, so the disease returned again. It is believed that this "silencing" may be due to the use of a number of "artificial" elements: first, the complete gene was not really used but a simplified version, the so-called cDNA, which contains the protein-coding information but lacks many elements required for the regulation of its production, and, second, a "promoter" of viral origin was used to ensure the expression of this gene. It seems that in the body, over time, these elements are in some way recognized as "foreign” and its “silencing” is triggered. As an alternative to the use of these elements, one could resort to the natural version of the gene, in all its length. This is technically more complicated because the genes are normally very large and do not fit into most viral vectors which are normally used in gene therapy trials (such as adenoviruses , retroviruses and lentiviruses). An interesting exception are herpes viral vectors, derived from herpes virus simplex (HSV-1), which are capable of accommodating up to 150 kb of genetic material.
Javier Diaz-Nido and Filip Lim Groups , at the Universidad Autónoma de Madrid in Spain, in collaboration with the Group of Richard Wade-Martins, of the University of Oxford in the United Kingdom, have been exploring the possibility of using this type of Herpes viral vectors for Friedreich's ataxia gene therapy (a disease caused by the deficiency of one protein called frataxin, and that is characterized by the degeneration of certain neurons in the central and peripheral nervous systems, as well as other alterations in the heart and the pancreas).
In a paper published in 2007 they showed that a herpes viral vector carrying the complete frataxin gene was able to make up for the functional deficiencies that were found in cultured skin cells of patients with Friedreich's ataxia (1).
Now, in a paper just published "online" in the "Gene Therapy" magazine (2) they have been able to demonstrate that vectors carrying the full frataxin gene allow a persistent expression "in vivo" after being injected into the mouse cerebellum. These results reinforce the view that vectors carrying complete genes are not silenced and may therefore be very useful for long term gene therapy. Another advantage of this type of herpes viral vectors is that they persist as stable “episomes” in the nuclei of cells, not integrated into any chromosome, and so they do not cause any alterations in the genome of these cells (unlike what happens with other vectors). Furthermore, herpes viral vectors are well known for their effective targeting of neuronal cells. In view of these data, it appears that herpes viral vectors carrying complete genes may be a safe and effective tool for the therapy of neurological hereditary diseases which are caused by recessive mutations (such as Friedreich's ataxia).
1.-Gomez-Sebastian S, Gimenez-Cassina A, Diaz-Nido J, Lim F, Wade-Martins R.
Infectious delivery and expression of 135 KB FRDA human genomic DNA locus complements Friedreich's ataxia deficiency in human cells.
MOL Ther. 2007 Feb; 15 (2): 248-54.
http://www.ncbi.NLM.NIH.gov/PubMed/17235301
http://www.nature.com/MT/journal/V15/N2/full/6300021a.html
2.-Gimenez-Cassina A, Wade-Martins R, Gomez-Sebastian S, Corona JC, Lim F, Diaz-Nido J.
Infectious delivery and long-term persistence of transgene expression in the brain by a 135-kb iBAC-FXN genomic DNA expression vector.
Gene Ther. 2011 Apr 14. DOI:10.1038/gt.2011.45 [Epub ahead of print]
http://www.ncbi.NLM.NIH.gov/PubMed/21490681
http://www.nature.com/gt/journal/vaop/ncurrent/full/gt201145a.html
Mitochondrial aconitase knockdown attenuates paraquat-induced dopaminergic cell death via decreased cellular metabolism and release of iron and H2O2*
Journal of Neurochemistry, 2011, DOI: 10.1111/j.1471-4159.2011.07290.x Accepted Article (Accepted, unedited articles published online for future issues)
David Cantu 1,3, Ruth E. Fulton 2, Derek A. Drechsel 2, Manisha Patel 1,2
1 Graduate Program in Neuroscience, 2 Department of Pharmaceutical Sciences, University of Colorado, 3 Department of Neuroscience, Tufts University School of Medicine
Keywords: mitochondrial aconitase, oxidative stress, paraquat, neurotoxicity, hydrogen peroxide, iron, cellular metabolism
David Cantu 1,3, Ruth E. Fulton 2, Derek A. Drechsel 2, Manisha Patel 1,2
1 Graduate Program in Neuroscience, 2 Department of Pharmaceutical Sciences, University of Colorado, 3 Department of Neuroscience, Tufts University School of Medicine
Keywords: mitochondrial aconitase, oxidative stress, paraquat, neurotoxicity, hydrogen peroxide, iron, cellular metabolism
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