ES2215494T1 - Moleculas pequeñas de rna que median la interferencia de rna. - Google Patents

Moleculas pequeñas de rna que median la interferencia de rna.

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Publication number
ES2215494T1
ES2215494T1 ES01985833T ES01985833T ES2215494T1 ES 2215494 T1 ES2215494 T1 ES 2215494T1 ES 01985833 T ES01985833 T ES 01985833T ES 01985833 T ES01985833 T ES 01985833T ES 2215494 T1 ES2215494 T1 ES 2215494T1
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target
gene
cell
rna molecule
nucleic acid
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ES2215494T3 (es
ES2215494T5 (es
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Thomas Tuschl
Sayda Elbashir
Winfried Lendeckel
Matthias Wilm
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Europaisches Laboratorium fuer Molekularbiologie EMBL
Max Planck Gesellschaft zur Foerderung der Wissenschaften eV
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Europaisches Laboratorium fuer Molekularbiologie EMBL
Max Planck Gesellschaft zur Foerderung der Wissenschaften eV
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Priority claimed from PCT/US2001/010188 external-priority patent/WO2001075164A2/en
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Abstract

Molécula de RNA bicatenario aislada, en la cual cada cadena de RNA tiene una longitud de 19-25 nucleótidos, en la cual cada molécula de RNA es capaz de modificaciones de ácido nucleico específicas en cuanto a la diana.

Claims (47)

1. Molécula de RNA bicatenario aislada, en la cual cada cadena de RNA tiene una longitud de 19-25 nucleótidos, en la cual cada molécula de RNA es capaz de modificaciones de ácido nucleico específicas en cuanto a la diana.
2. La molécula de RNA de la reivindicación 1, en la cual al menos una cadena tiene un saliente 3' de
1-5 nucleótidos.
3. La molécula de RNA de la reivindicación 1 ó 2, capaz de interferencia de RNA específica en cuanto a la diana y/o metilación del DNA.
4. La molécula de RNA de una cualquiera de las reivindicaciones 1-3, en la cual cada cadena tiene una longitud de 19-23, particularmente de 20-22 nucleótidos.
5. La molécula de RNA de una cualquiera de las reivindicaciones 2-4, en la cual el saliente 3' es de
1-3 nucleótidos.
6. La molécula de RNA de una cualquiera de las reivindicaciones 2-5, en la cual el saliente 3' está estabilizado contra la degradación.
7. La molécula de RNA de una cualquiera de las reivindicaciones 1-6, que contiene al menos un análogo de nucleótido modificado.
8. La molécula de RNA de la reivindicación 7, en la cual el análogo de nucleótido modificado se selecciona de ribonucleótidos modificados en el azúcar o en la cadena principal.
9. La molécula de RNA de acuerdo con la reivindicación 7 ó 8, en la cual el análogo de nucleótido es un ribonucleótido modificado en el azúcar, en el cual el grupo 2'-OH está reemplazado por un grupo seleccionado de H, OR, R, halo, SH, SR^{1}, NH_{2}, NHR, NR^{2} o CN, en el cual R es C_{1}-C_{6} alquilo, alquenilo o alquinilo y halo es F, Cl, Br o I.
10. La molécula de RNA de la reivindicación 7 u 8, en la cual el análogo de nucleótido es un ribonucleótido modificado en la cadena principal que contiene un grupo fosfotioato.
11. La molécula de RNA de una cualquiera de las reivindicaciones 1-10, que tiene una secuencia que tiene una identidad de al menos 50 por ciento con una molécula diana de mRNA predeterminada.
12. La molécula de RNA de la reivindicación 11, en la cual la identidad es al menos 70 por ciento.
13. Un método de preparación de una molécula de RNA bicatenario de una cualquiera de las reivindicaciones 1-12 que comprende los pasos:
(a)
sintetizar dos cadenas de RNA cada una de las cuales tiene una longitud de 19-25 nucleótidos, siendo dichas cadenas de RNA capaces de formar una molécula de RNA bicatenario,
(b)
combinar las cadenas de RNA sintetizadas en condiciones en las cuales se forma una molécula de RNA bicatenario, que es susceptible de modificaciones de ácido nucleico específicas en cuanto a la diana.
14. El método de la reivindicación 13, en el cual las cadenas de RNA se sintetizan químicamente.
15. El método de la reivindicación 13, en el cual las cadenas de RNA se sintetizan enzimáticamente.
16. Un método de mediación de modificaciones de ácido nucleico específicas en cuanto a la diana en una célula o un organismo que comprende los pasos de:
(a)
poner en contacto dicha célula u organismo con la molécula de RNA bicatenario de una cualquiera de las reivindicaciones 1-12 en condiciones en las cuales pueden producirse modificaciones de ácido nucleico específicas en cuanto a la diana, y
(b)
mediar una modificación de ácido nucleico específica en cuanto a la diana efectuada por el RNA bicatenario hacia un ácido nucleico diana que tiene una porción de secuencia que corresponde sustancialmente al RNA bicatenario.
17. El método de la reivindicación 16, en el cual la modificación de ácido nucleico es interferencia de RNA y/o metilación de DNA.
18. El método de la reivindicación 16 y 17 en el cual dicha puesta en contacto comprende introducir dicha molécula de RNA bicatenario en una célula diana en la cual puede producirse la modificación de ácido nucleico específica en cuanto a la diana.
19. El método de la reivindicación 18, en el cual la introducción comprende un suministro o inyección mediado por un vehículo.
20. Uso del método de una cualquiera de las reivindicaciones 16-19 para determinar la función de un gen en una célula o un organismo.
21. Uso del método de una cualquiera de las reivindicaciones 16-19 para modulación de la función de un gen en una célula o un organismo.
22. El uso de la reivindicación 20 ó 21, en el cual el gen está asociado con una condición patológica.
23. El uso de la reivindicación 22, en el cual el gen es un gen asociado con un patógeno.
24. El uso de la reivindicación 23, en el cual el gen es un gen vírico.
25. El uso de la reivindicación 22, en el cual el gen es un gen asociado con un tumor.
26. El uso de la reivindicación 22, en el cual el gen es un gen asociado a una enfermedad autoinmune.
27. Composición farmacéutica que contiene como agente activo al menos una molécula de RNA bicatenario de una cualquiera de las reivindicaciones 1-12 y un vehículo farmacéutico.
28. La composición de la reivindicación 27 para aplicaciones de diagnóstico.
29. La composición de la reivindicación 27 para aplicaciones terapéuticas.
30. Una célula eucariota o un organismo eucariota no humano que exhibe un fenotipo mutado por modificación génica ("knockout") específico en cuanto al gen diana en la cual dicha célula u organismo está transfectada con al menos una molécula de RNA bicatenario capaz de inhibir la expresión de un gen diana endógeno o con un DNA que codifica al menos una molécula de RNA bicatenario capaz de inhibir la expresión de al menos un gen diana endógeno.
31. La célula u organismo de la reivindicación 30, que es una célula de mamífero.
32. La célula u organismo de la reivindicación 31 que es una célula humana.
33. La célula u organismo de una cualquiera de las reivindicaciones 30-32 que está transfectada adicionalmente con al menos un ácido nucleico diana exógeno que codifica la proteína diana o una forma variante o mutada de la proteína diana, en la cual dicho ácido nucleico diana exógeno difiere del gen diana endógeno en el nivel de ácido nucleico de tal modo que la expresión del ácido nucleico diana exógeno se ve sustancialmente menos inhibida por la molécula de RNA bicatenario que la expresión del gen diana endógeno.
34. La célula o el organismo de la reivindicación 33 en la cual el ácido nucleico diana exógeno está fusionado con una secuencia de ácido nucleico adicional que codifica un péptido o polipéptido detectable.
35. Uso de la célula u organismo de una cualquiera de las reivindicaciones 30-34 para procedimientos analíticos.
36. El uso de la reivindicación 35 para el análisis de perfiles de expresión génica.
37. El uso de la reivindicación 35 para un análisis del proteoma.
38. El uso de una cualquiera de las reivindicaciones 35-37 en el cual se efectúa un análisis de una forma variante o mutante de la proteína diana codificada por un ácido nucleico diana exógeno.
39. El uso de la reivindicación 38 para identificar dominios funcionales de la proteína diana.
40. El uso de una cualquiera de las reivindicaciones 35-39 en el cual se efectúa una comparación de al menos dos células u organismos seleccionados de:
(i)
una célula de control u organismo de control sin inhibición del gen diana,
(ii)
una célula u organismo con inhibición del gen diana y
(iii)
una célula u organismo con inhibición del gen diana más complementación del gen diana por un ácido nucleico exógeno.
41. El uso de una cualquiera de las reivindicaciones 35-40 en el cual el análisis comprende un análisis funcional y/o fenotípico.
42. Uso de una célula de una cualquiera de las reivindicaciones 30-34 para procedimientos preparativos.
43. El uso de la reivindicación 41 para el aislamiento de proteínas o complejos proteínicos a partir de células eucariotas.
44. El uso de la reivindicación 43 para el aislamiento de complejos proteínicos de peso molecular alto que pueden contener opcionalmente ácidos nucleicos.
45. El uso de una cualquiera de las reivindicaciones 35-44 en un procedimiento para identificación y/o caracterización de agentes farmacológicos.
46. Un sistema para identificación y/o caracterización de un agente farmacológico que actúa sobre al menos una proteína diana que comprende:
(a)
una célula eucariota o un organismo eucariota no humano capaz de expresar al menos un gen diana que codifica dicha al menos una proteína diana,
(b)
al menos una molécula de RNA bicatenario capaz de inhibir la expresión de dicho al menos un gen diana endógeno, y
(c)
una sustancia de ensayo o un conjunto de sustancias de ensayo en lo(s) cual(es) deben identificarse y/o caracterizarse las propiedades farmacológicas de dicha sustancia de ensayo o dicho conjunto.
47. El sistema de la reivindicación 46 que comprende adicionalmente:
(d)
al menos un ácido nucleico diana exógeno que codifica la proteína diana o una forma variante o mutada de la proteína diana, en el cual dicho ácido nucleico diana exógeno difiere del gen diana endógeno en el nivel de ácido nucleico de tal modo que la expresión del ácido nucleico diana exógeno se ve sustancialmente menos inhibida por la molécula de RNA bicatenario que la expresión del gen diana endógeno.
ES01985833.1T 2000-12-01 2001-11-29 Moléculas de RNA pequeñas que median la interferencia de RNA Expired - Lifetime ES2215494T5 (es)

Applications Claiming Priority (7)

Application Number Priority Date Filing Date Title
EP00126325 2000-12-01
EP00126325 2000-12-01
US27966101P 2001-03-30 2001-03-30
PCT/US2001/010188 WO2001075164A2 (en) 2000-03-30 2001-03-30 Rna sequence-specific mediators of rna interference
US279661P 2001-03-30
WOPCT/US01/10188 2001-03-30
PCT/EP2001/013968 WO2002044321A2 (en) 2000-12-01 2001-11-29 Rna interference mediating small rna molecules

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