ES2510396T3 - Direccionamiento a células de cáncer usando nanopartículas - Google Patents
Direccionamiento a células de cáncer usando nanopartículas Download PDFInfo
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- ES2510396T3 ES2510396T3 ES11186037.5T ES11186037T ES2510396T3 ES 2510396 T3 ES2510396 T3 ES 2510396T3 ES 11186037 T ES11186037 T ES 11186037T ES 2510396 T3 ES2510396 T3 ES 2510396T3
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Classifications
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Abstract
Una nanopartícula furtiva específica de diana que comprende: una matriz polimérica y un agente terapéutico; donde la matriz polimérica comprende un polímero no funcionalizado que comprende un copolímero de ácido poliláctico y polietilenglicol o poli(ácido láctico-co-ácido glicólico) y polietilenglicol; y un polímero conjugado con un ligando unido covalentemente a un ligando PSMA de bajo peso molecular; donde el ligando PSMA de bajo peso molecular es de Fórmulas I, II, III o IV:^**Fórmula** y sus enantiómeros, estereoisómeros, rotámeros, tautómeros, diastereómeros o racematos; donde m y n son cada uno, independientemente, 0, 1, 2 o 3; p es 0 o 1; R1, R2, R4 y R5 se seleccionan cada uno, independientemente, del grupo que consiste en alquilo sustituido o no sustituido, arilo sustituido o no sustituido y cualquier combinación de estos; y R3 es H o alquilo C1-6; y donde R1, R2, R4 y R5 comprenden un punto de unión a la nanopartícula.
Description
E11186037
29-09-2014
- Agente terapéutico
- Polímero biocompatible Polímero furtivo Resto de dirección
- mitoxantrona
- PLA PEG GL1
- mitoxantrona
- PGA PEG GL1
- mitoxantrona
- PLGA PEG GL2
- mitoxantrona
- PLA PEG GL2
- mitoxantrona
- PGA PEG GL2
- mitoxantrona
- PLGA PEG-DSPE GL1
- mitoxantrona
- PLA PEG-DSPE GL1
- mitoxantrona
- PGA PEG-DSPE GL1
- mitoxantrona
- PLGA PEG-DSPE GL2
- mitoxantrona
- PLA PEG-DSPE GL2
- mitoxantrona
- PGA PEG-DSPE GL2
- docetaxel
- PLGA PEG GL1
- docetaxel
- PLA PEG GL1
- docetaxel
- PGA PEG GL1
- docetaxel
- PLGA PEG GL2
- docetaxel
- PLA PEG GL2
- docetaxel
- PGA PEG GL2
- docetaxel
- PLGA PEG-DSPE GL1
- docetaxel
- PLA PEG-DSPE GL1
- docetaxel
- PGA PEG-DSPE GL1
- docetaxel
- PLGA PEG-DSPE GL2
- docetaxel
- PLA PEG-DSPE GL2
- docetaxel
- PGA PEG-DSPE GL2
- doxorubicina
- PLGA PEG GL1
- doxorubicina
- PLA PEG GL1
- doxorubicina
- PGA PEG GL1
- doxorubicina
- PLGA PEG GL2
- doxorubicina
- PLA PEG GL2
- doxorubicina
- PGA PEG GL2
- doxorubicina
- PLGA PEG-DSPE GL1
- doxorubicina
- PLA PEG-DSPE GL1
- doxorubicina
- PGA PEG-DSPE GL1
- doxorubicina
- PLGA PEG-DSPE GL2
- doxorubicina
- PLA PEG-DSPE GL2
36
E11186037
29-09-2014
- Agente terapéutico
- Polímero biocompatible Polímero furtivo Resto de dirección
- doxorubicina
- PGA PEG-DSPE GL2
- gemcitabina
- PLGA PEG GL1
- gemcitabina
- PLA PEG GL1
- gemcitabina
- PGA PEG GL1
- gemcitabina
- PLGA PEG GL2
- gemcitabina
- PLA PEG GL2
- gemcitabina
- PGA PEG GL2
- gemcitabina
- PLGA PEG-DSPE GL1
- gemcitabina
- PLA PEG-DSPE GL1
- gemcitabina
- PGA PEG-DSPE GL1
- gemcitabina
- PLGA PEG-DSPE GL2
- gemcitabina
- PLA PEG-DSPE GL2
- gemcitabina
- PGA PEG-DSPE GL2
- 5-fluorouracilo
- PLGA PEG GL1
- 5-fluorouracilo
- PLA PEG GL1
- 5-fluorouracilo
- PGA PEG GL1
- 5-fluorouracilo
- PLGA PEG GL2
- 5-fluorouracilo
- PLA PEG GL2
- 5-fluorouracilo
- PGA PEG GL2
- 5-fluorouracilo
- PLGA PEG-DSPE GL1
- 5-fluorouracilo
- PLA PEG-DSPE GL1
- 5-fluorouracilo
- PGA PEG-DSPE GL1
- 5-fluorouracilo
- PLGA PEG-DSPE GL2
- 5-fluorouracilo
- PLA PEG-DSPE GL2
- 5-fluorouracilo
- PGA PEG-DSPE GL2
- paclitaxel
- PLGA PEG GL1
- paclitaxel
- PLA PEG GL1
- paclitaxel
- PGA PEG GL1
- paclitaxel
- PLGA PEG GL2
- paclitaxel
- PLA PEG GL2
- paclitaxel
- PGA PEG GL2
- paclitaxel
- PLGA PEG-DSPE GL1
- paclitaxel
- PLA PEG-DSPE GL1
- paclitaxel
- PGA PEG-DSPE GL1
37
E11186037
29-09-2014
- Agente terapéutico
- Polímero biocompatible Polímero furtivo Resto de dirección
- paclitaxel
- PLGA PEG-DSPE GL2
- paclitaxel
- PLA PEG-DSPE GL2
- paclitaxel
- PGA PEG-DSPE GL2
- daunorrubicina
- PLGA PEG GL1
- daunorrubicina
- PLA PEG GL1
- daunorrubicina
- PGA PEG GL1
- daunorrubicina
- PLGA PEG GL2
- daunorrubicina
- PLA PEG GL2
- daunorrubicina
- PGA PEG GL2
- daunorrubicina
- PLGA PEG-DSPE GL1
- daunorrubicina
- PLA PEG-DSPE GL1
- daunorrubicina
- PGA PEG-DSPE GL1
- daunorrubicina
- PLGA PEG-DSPE GL2
- daunorrubicina
- PLA PEG-DSPE GL2
- daunorrubicina
- PGA PEG-DSPE GL2
Ejemplo 4: Unión/captación de nanopartículas mediada por resto de dirección molecular pequeño en células LNcap
La unión y captación de nanopartículas (NP-GL1, NP-GL2) con ligandos unidos a superficie GL1 (basado en ácido glutámico/4-amino-fenilalanina) y GL2 (basado en ácido glutámico/lisina) por células LNCap que expresan altos niveles de PSMA se ensayó por comparación con nanopartículas (NP) de PLGA-PEG desnudas como control
5 negativo y NP que llevaban aptámero (Apt) de antígeno de membrana específico de próstata (PSMA) A10 terminado en amina (NP-Apt) como control positivo. La captación de NP-GL1, NP-GL2, NP y NP-Apt por células LNCap y células PC3 que expresan bajos niveles de PSMA se comparó con la unión/captación mediada por PSMA específica de NP-GL1, NP-GL2.
10 Copolímero dibloque PLGA0,67-PEG5000-CO2H (solución madre 50 mg/ml en ACN); Aptámero (1 mg/ml); ligando basado en ácido glutámico/fenilalanina (GL11); ligando basado en ácido glutámico/lisina (GL2); EDC. HCl (Pierce Biotech); SulfoNHS (Pierce Biotech), solución salina tamponada con fosfato, PBS (Sigma); tampón de fijación: formaldehído al 4% recién preparado en PBS; solución de bloqueo: BSA al 1% recién preparado en PBS; solución de bloqueo y permeabilización: Tritón X100 0,1 recién preparado en solución de bloqueo. Alexa-568 faloidina (5
15 U/ml), NBD Colesterol (Invitrogen); DAPI (Sigma): 0,1 mg/ml; Vectashield (Vector Labs); laca de uñas.
Preparación de nanopartículas:
Se prepararon nanopartículas basadas en copolímeros dibloque de PLGA-PEG-CO2H mediante el procedimiento de nanoprecipitación. Se unió covalentemente GL1, GL2 y Apt a los extremos terminales ácido carboxílico de la corona de PEG de nanopartículas en suspensión acuosa de PBS. La conjugación covalente de GL1, GL2 y Apt con NP
20 estaba basada en la activación por EDC/NHS del extremo terminal de PEG ácido carboxílico y la posterior reacción de los grupos terminales de éster de succinimida activos con la funcionalidad amina en GL1, GL2 y Apt usando el procedimiento siguiente:
Se diluyó solución madre de PLGA-PEG-CO2H (1,2 ml, solución 50 mg/ml en acetonitrilo) con acetonitrilo para dar 6 ml de solución dibloque de 10 mg/ml. Se añadió NBD Colesterol (600 l, solución 1 mg/ml en DMF) a la solución 25 dibloque anterior y la mezcla se añadió gota a gota a 12 ml de agua desionizada agitada (18 M). La suspensión de NP resultante se dejó agitar (400 rpm) abierta en una campana extractora durante 2 h, y posteriormente se purificó por ultrafiltración usando filtros Amicon basados en celulosa regenerada (MWCO 5000 Da) para eliminar el acetonitrilo, DMF y NBD no encapsulado residual de la forma siguiente. Se transfirió la suspensión de NP (16 ml) en cuatro porciones iguales a cuatro tubos de filtración centrífuga Amicon de 15 ml y se concentraron hasta 250-400 l
38
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