Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity
NTRODUCTION: Scorpion venom contains peptides with neurotoxic action primarily active on ion channels in the nervous system of insects and mammals. They are also characterized as cytolytic and anticancer, biological characteristics that have not yet been reported for the Tityus macrochirus venom. OB...
- Autores:
-
Rincón-Cortés, Clara Andrea
Reyes-Montaño, Edgar Antonio
Vega-Castro, Nohora Angélica
- Tipo de recurso:
- Article of journal
- Fecha de publicación:
- 2017
- Institución:
- Universidad de Ciencias Aplicadas y Ambientales U.D.C.A
- Repositorio:
- Repositorio Institucional UDCA
- Idioma:
- spa
- OAI Identifier:
- oai:repository.udca.edu.co:11158/2332
- Acceso en línea:
- https://revistabiomedica.org/index.php/biomedica/article/view/3265
- Palabra clave:
- Células Tumorales Cultivadas
Supervivencia celular
Neoplasias
Venenos de Escorpión
Cell Line
TumorCell Survival
Scorpiones
Neoplasms
Peptides
Arachnida
Scorpion Venoms
Scorpiones
Arachnida
Péptidos
- Rights
- openAccess
- License
- Derechos Reservados - Universidad de Ciencias Aplicadas y Ambientales
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Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| dc.title.alternative.spa.fl_str_mv |
Purificación parcial de péptidos presentes en el veneno del escorpión Tityus macrochirus (Buthidae) y evaluación preliminar de su actividad citotóxica |
| title |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| spellingShingle |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity Células Tumorales Cultivadas Supervivencia celular Neoplasias Venenos de Escorpión Cell Line TumorCell Survival Scorpiones Neoplasms Peptides Arachnida Scorpion Venoms Scorpiones Arachnida Péptidos |
| title_short |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| title_full |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| title_fullStr |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| title_full_unstemmed |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| title_sort |
Partial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicity |
| dc.creator.fl_str_mv |
Rincón-Cortés, Clara Andrea Reyes-Montaño, Edgar Antonio Vega-Castro, Nohora Angélica |
| dc.contributor.author.spa.fl_str_mv |
Rincón-Cortés, Clara Andrea Reyes-Montaño, Edgar Antonio Vega-Castro, Nohora Angélica |
| dc.subject.mesh.spa.fl_str_mv |
Células Tumorales Cultivadas Supervivencia celular Neoplasias Venenos de Escorpión |
| topic |
Células Tumorales Cultivadas Supervivencia celular Neoplasias Venenos de Escorpión Cell Line TumorCell Survival Scorpiones Neoplasms Peptides Arachnida Scorpion Venoms Scorpiones Arachnida Péptidos |
| dc.subject.proposal.spa.fl_str_mv |
Cell Line TumorCell Survival Scorpiones Neoplasms Peptides Arachnida Scorpion Venoms |
| dc.subject.agrovoc.spa.fl_str_mv |
Scorpiones Arachnida Péptidos |
| description |
NTRODUCTION: Scorpion venom contains peptides with neurotoxic action primarily active on ion channels in the nervous system of insects and mammals. They are also characterized as cytolytic and anticancer, biological characteristics that have not yet been reported for the Tityus macrochirus venom. OBJECTIVE: To assess if the total T. macrochirus venom and the fraction of partially purified peptides decrease the viability of various tumor-derived cell lines. MATERIALS AND METHODS: The scorpion venom was collected by electrical stimulation and, subsequently, subjected to chromatography, electrophoresis, and ultrafiltration with Amicon Ultra 0.5® membranes for the partial identification and purification of its peptides. The cytotoxic activity of the venom and the peptides fraction trials on tumor-derived cell lines were carried out by the MTT method. RESULTS: The T. macrochirus scorpion venom has peptides with molecular weights ranging between 3 and 10 kDa. They were partially purified using the ultrafiltration technique, and assessed by the RP-HPLC method. Cytotoxicity trials with the whole T. macrochirus venom showed a higher viability decrease on the PC3 cell line compared to the other cell lines assessed, while the partially purified peptides decreased the HeLa cell line viability. |
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2017 |
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2017 |
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2019-10-23T19:44:55Z |
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2019-10-23T19:44:55Z |
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Artículo de revista |
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Rincón-Cortés, C. A., Reyes-Montaño, E. A., & Vega-Castro, N. A. (2017). Partial purification of peptides present in the tityus macrochirus (buthidae) scorpion venom and preliminary assessment of their cytotoxicity. Biomedica : Revista Del Instituto Nacional De Salud, 37(2), 238-249. doi:10.7705/biomedica.v37i3.3265 |
| dc.identifier.uri.spa.fl_str_mv |
https://revistabiomedica.org/index.php/biomedica/article/view/3265 |
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10.7705/biomedica.v37i3.3265 |
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Rincón-Cortés, C. A., Reyes-Montaño, E. A., & Vega-Castro, N. A. (2017). Partial purification of peptides present in the tityus macrochirus (buthidae) scorpion venom and preliminary assessment of their cytotoxicity. Biomedica : Revista Del Instituto Nacional De Salud, 37(2), 238-249. doi:10.7705/biomedica.v37i3.3265 10.7705/biomedica.v37i3.3265 |
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https://revistabiomedica.org/index.php/biomedica/article/view/3265 |
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Biomedica : revista del Instituto Nacional de Salud;Vol. 37, No.,2, Jun. 1 2017, páginas 238-249 |
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Derechos Reservados - Universidad de Ciencias Aplicadas y Ambientales |
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Derechos Reservados - Universidad de Ciencias Aplicadas y Ambientales https://creativecommons.org/licenses/by-nc-sa/4.0/ Atribución-NoComercial-CompartirIgual 4.0 Internacional (CC BY-NC-SA 4.0) http://purl.org/coar/access_right/c_abf2 |
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Rincón-Cortés, Clara AndreaReyes-Montaño, Edgar AntonioVega-Castro, Nohora Angélica2019-10-23T19:44:55Z2019-10-23T19:44:55Z2017Rincón-Cortés, C. A., Reyes-Montaño, E. A., & Vega-Castro, N. A. (2017). Partial purification of peptides present in the tityus macrochirus (buthidae) scorpion venom and preliminary assessment of their cytotoxicity. Biomedica : Revista Del Instituto Nacional De Salud, 37(2), 238-249. doi:10.7705/biomedica.v37i3.3265https://revistabiomedica.org/index.php/biomedica/article/view/326510.7705/biomedica.v37i3.3265NTRODUCTION: Scorpion venom contains peptides with neurotoxic action primarily active on ion channels in the nervous system of insects and mammals. They are also characterized as cytolytic and anticancer, biological characteristics that have not yet been reported for the Tityus macrochirus venom. OBJECTIVE: To assess if the total T. macrochirus venom and the fraction of partially purified peptides decrease the viability of various tumor-derived cell lines. MATERIALS AND METHODS: The scorpion venom was collected by electrical stimulation and, subsequently, subjected to chromatography, electrophoresis, and ultrafiltration with Amicon Ultra 0.5® membranes for the partial identification and purification of its peptides. The cytotoxic activity of the venom and the peptides fraction trials on tumor-derived cell lines were carried out by the MTT method. RESULTS: The T. macrochirus scorpion venom has peptides with molecular weights ranging between 3 and 10 kDa. They were partially purified using the ultrafiltration technique, and assessed by the RP-HPLC method. Cytotoxicity trials with the whole T. macrochirus venom showed a higher viability decrease on the PC3 cell line compared to the other cell lines assessed, while the partially purified peptides decreased the HeLa cell line viability.Introducción. El veneno del escorpión posee péptidos con actividad neurotóxica que actúan principalmente en los canales iónicos del sistema nervioso de insectos y mamíferos. También se ha establecido su acción citolítica y anticancerígena, características biológicas que aún no se han explorado en el veneno del escorpión Tityus macrochirus.Objetivo. Evaluar si tanto el veneno total de T. macrochirus como la fracción de péptidos parcialmente purificados disminuyen el porcentaje de viabilidad de diferentes líneas celulares provenientes de tumores.Materiales y métodos. Mediante métodos cromatográficos, electroforéticos y de ultrafiltración con membranas de Amicon Ultra 0.5®, se identificaron y purificaron parcialmente los péptidos del veneno de T. macrochirus obtenido mediante estimulación eléctrica. Los ensayos de actividad citotóxica del veneno y de la fracción de péptidos se hicieron en líneas celulares provenientes de tumores con el método colorimétrico de reducción de la sal de tetrazolio (Mossman’s Tetrazole Test, MTT).Resultados. El veneno de T. macrochirus posee péptidos con pesos moleculares entre 3 y 10 kDa, los cuales se purificaron parcialmente mediante ultrafiltración y se evaluaron mediante cromatografía líquida de alta resolución en fase inversa (Reverse Phase-High Pressure Liquid Chromatography, RP-HPLC). Los ensayos de citotoxicidad del veneno total de T. macrochirus evidenciaron una mayor disminución de la viabilidad en la línea celular PC3 que en las demás líneas celulares evaluadas, en tanto que la fracción parcialmente purificada de péptidos logró disminuir la viabilidad de la línea celular HeLa.Conclusión. Los péptidos del veneno de T. macrochirus presentaron actividad citotóxica en algunas de las líneas celulares provenientes de tumores, y se observó algún grado de selectividad frente a ellaapplication/pdfspaBiomedica : revista del Instituto Nacional de Salud;Vol. 37, No.,2, Jun. 1 2017, páginas 238-249Derechos Reservados - Universidad de Ciencias Aplicadas y Ambientaleshttps://creativecommons.org/licenses/by-nc-sa/4.0/info:eu-repo/semantics/openAccessAtribución-NoComercial-CompartirIgual 4.0 Internacional (CC BY-NC-SA 4.0)http://purl.org/coar/access_right/c_abf2https://udca.elogim.com:2092/record/display.uri?eid=2-s2.0-85040188599&origin=resultslist&sort=plf-f&src=s&nlo=1&nlr=20&nls=afprfnm-t&affilName=UDCA&sid=0e299568affc46b586426220856d6e50&sot=afnl&sdt=cl&cluster=scopubyr%2c%222017%22%2ct&sl=67&s=%28AF-ID%28%22Universidad+de+Ciencias+Aplicadas+y+Ambientales%22+60106980%29%29&relpos=27&citeCnt=1&searchTerm=Células Tumorales CultivadasSupervivencia celularNeoplasiasVenenos de EscorpiónCell LineTumorCell SurvivalScorpionesNeoplasmsPeptidesArachnidaScorpion VenomsScorpionesArachnidaPéptidosPartial purification of peptides present in the Tityus macrochirus (Buthidae) scorpion venom and preliminary assessment of their cytotoxicityPurificación parcial de péptidos presentes en el veneno del escorpión Tityus macrochirus (Buthidae) y evaluación preliminar de su actividad citotóxicaArtículo de revistahttp://purl.org/coar/resource_type/c_6501http://purl.org/coar/resource_type/c_2df8fbb1info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionTexthttp://purl.org/redcol/resource_type/ARThttp://purl.org/coar/version/c_970fb48d4fbd8a85PublicationORIGINAL3265-Texto del manuscrito completo (cuadros y figuras insertos)-18186-1-10-20170519.pdf3265-Texto del manuscrito completo (cuadros y figuras 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en las Obras Colectivas;
b.	Distribuir copias o fonogramas de las Obras, exhibirlas públicamente, ejecutarlas públicamente y/o ponerlas a disposición pública, incluyéndolas como incorporadas en Obras Colectivas, según corresponda;
c.	Distribuir copias de las Obras Derivadas que se generen, exhibirlas públicamente, ejecutarlas públicamente y/o ponerlas a disposición pública.
Los derechos mencionados anteriormente pueden ser ejercidos en todos los medios y formatos, actualmente conocidos o que se inventen en el futuro. Los derechos antes mencionados incluyen el derecho a realizar dichas modificaciones en la medida que sean técnicamente necesarias para ejercer los derechos en otro medio o formatos, pero de otra manera usted no está autorizado para realizar obras derivadas.Todos los derechos no otorgados expresamente por el Licenciante quedan por este medio reservados, incluyendo pero sin limitarse a aquellos que se mencionan en las secciones 4(d) y 4(e).
4. Restricciones.
La licencia otorgada en la anterior Sección 3 está expresamente sujeta y limitada por las siguientes restricciones:
a.	Usted puede distribuir, exhibir públicamente, ejecutar públicamente, o poner a disposición pública la Obra sólo bajo las condiciones de esta Licencia, y Usted debe incluir una copia de esta licencia o del Identificador Universal de Recursos de la misma con cada copia de la Obra que distribuya, exhiba públicamente, ejecute públicamente o ponga a disposición pública. No es posible ofrecer o imponer ninguna condición sobre la Obra que altere o limite las condiciones de esta Licencia o el ejercicio de los derechos de los destinatarios otorgados en este documento. No es posible sublicenciar la Obra. Usted debe mantener intactos todos los avisos que hagan referencia a esta Licencia y a la cláusula de limitación de garantías. Usted no puede distribuir, exhibir públicamente, ejecutar públicamente, o poner a disposición pública la Obra con alguna medida tecnológica que controle el acceso o la utilización de ella de una forma que sea inconsistente con las condiciones de esta Licencia. Lo anterior se aplica a la Obra incorporada a una Obra Colectiva, pero esto no exige que la Obra Colectiva aparte de la obra misma quede sujeta a las condiciones de esta Licencia. Si Usted crea una Obra Colectiva, previo aviso de cualquier Licenciante debe, en la medida de lo posible, eliminar de la Obra Colectiva cualquier referencia a dicho Licenciante o al Autor Original, según lo solicitado por el Licenciante y conforme lo exige la cláusula 4(c).
b.	Usted no puede ejercer ninguno de los derechos que le han sido otorgados en la Sección 3 precedente de modo que estén principalmente destinados o directamente dirigidos a conseguir un provecho comercial o una compensación monetaria privada. El intercambio de la Obra por otras obras protegidas por derechos de autor, ya sea a través de un sistema para compartir archivos digitales (digital file-sharing) o de cualquier otra manera no será considerado como estar destinado principalmente o dirigido directamente a conseguir un provecho comercial o una compensación monetaria privada, siempre que no se realice un pago mediante una compensación monetaria en relación con el intercambio de obras protegidas por el derecho de autor.
c.	Si usted distribuye, exhibe públicamente, ejecuta públicamente o ejecuta públicamente en forma digital la Obra o cualquier Obra Derivada u Obra Colectiva, Usted debe mantener intacta toda la información de derecho de autor de la Obra y proporcionar, de forma razonable según el medio o manera que Usted esté utilizando: (i) el nombre del Autor Original si está provisto (o seudónimo, si fuere aplicable), y/o (ii) el nombre de la parte o las partes que el Autor Original y/o el Licenciante hubieren designado para la atribución (v.g., un instituto patrocinador, editorial, publicación) en la información de los derechos de autor del Licenciante, términos de servicios o de otras formas razonables; el título de la Obra si está provisto; en la medida de lo razonablemente factible y, si está provisto, el Identificador Uniforme de Recursos (Uniform Resource Identifier) que el Licenciante especifica para ser asociado con la Obra, salvo que tal URI no se refiera a la nota sobre los derechos de autor o a la información sobre el licenciamiento de la Obra; y en el caso de una Obra Derivada, atribuir el crédito identificando el uso de la Obra en la Obra Derivada (v.g., "Traducción Francesa de la Obra del Autor Original," o "Guión Cinematográfico basado en la Obra original del Autor Original"). Tal crédito puede ser implementado de cualquier forma razonable; en el caso, sin embargo, de Obras Derivadas u Obras Colectivas, tal crédito aparecerá, como mínimo, donde aparece el crédito de cualquier otro autor comparable y de una manera, al menos, tan destacada como el crédito de otro autor comparable.
d.	Para evitar toda confusión, el Licenciante aclara que, cuando la obra es una composición musical:
i.	Regalías por interpretación y ejecución bajo licencias generales. El Licenciante se reserva el derecho exclusivo de autorizar la ejecución pública o la ejecución pública digital de la obra y de recolectar, sea individualmente o a través de una sociedad de gestión colectiva de derechos de autor y derechos conexos (por ejemplo, SAYCO), las regalías por la ejecución pública o por la ejecución pública digital de la obra (por ejemplo Webcast) licenciada bajo licencias generales, si la interpretación o ejecución de la obra está primordialmente orientada por o dirigida a la obtención de una ventaja comercial o una compensación monetaria privada.
ii.	Regalías por Fonogramas. El Licenciante se reserva el derecho exclusivo de recolectar, individualmente o a través de una sociedad de gestión colectiva de derechos de autor y derechos conexos (por ejemplo, SAYCO), una agencia de derechos musicales o algún agente designado, las regalías por cualquier fonograma que Usted cree a partir de la obra (“versión cover”) y distribuya, en los términos del régimen de derechos de autor, si la creación o distribución de esa versión cover está primordialmente destinada o dirigida a obtener una ventaja comercial o una compensación monetaria privada.
e.	Gestión de Derechos de Autor sobre Interpretaciones y Ejecuciones Digitales (WebCasting). Para evitar toda confusión, el Licenciante aclara que, cuando la obra sea un fonograma, el Licenciante se reserva el derecho exclusivo de autorizar la ejecución pública digital de la obra (por ejemplo, webcast) y de recolectar, individualmente o a través de una sociedad de gestión colectiva de derechos de autor y derechos conexos (por ejemplo, Acinpro), las regalías por la ejecución pública digital de la obra (por ejemplo, webcast), sujeta a las disposiciones aplicables del régimen de Derecho de Autor, si esta ejecución pública digital está primordialmente dirigida a obtener una ventaja comercial o una compensación monetaria privada.
5. Representaciones, Garantías y Limitaciones de Responsabilidad.
A MENOS QUE LAS PARTES LO ACORDARAN DE OTRA FORMA POR ESCRITO, EL LICENCIANTE OFRECE LA OBRA (EN EL ESTADO EN EL QUE SE ENCUENTRA) “TAL CUAL”, SIN BRINDAR GARANTÍAS DE CLASE ALGUNA RESPECTO DE LA OBRA, YA SEA EXPRESA, IMPLÍCITA, LEGAL O CUALQUIERA OTRA, INCLUYENDO, SIN LIMITARSE A ELLAS, GARANTÍAS DE TITULARIDAD, COMERCIABILIDAD, ADAPTABILIDAD O ADECUACIÓN A PROPÓSITO DETERMINADO, AUSENCIA DE INFRACCIÓN, DE AUSENCIA DE DEFECTOS LATENTES O DE OTRO TIPO, O LA PRESENCIA O AUSENCIA DE ERRORES, SEAN O NO DESCUBRIBLES (PUEDAN O NO SER ESTOS DESCUBIERTOS). ALGUNAS JURISDICCIONES NO PERMITEN LA EXCLUSIÓN DE GARANTÍAS IMPLÍCITAS, EN CUYO CASO ESTA EXCLUSIÓN PUEDE NO APLICARSE A USTED.
6. Limitación de responsabilidad.
A MENOS QUE LO EXIJA EXPRESAMENTE LA LEY APLICABLE, EL LICENCIANTE NO SERÁ RESPONSABLE ANTE USTED POR DAÑO ALGUNO, SEA POR RESPONSABILIDAD EXTRACONTRACTUAL, PRECONTRACTUAL O CONTRACTUAL, OBJETIVA O SUBJETIVA, SE TRATE DE DAÑOS MORALES O PATRIMONIALES, DIRECTOS O INDIRECTOS, PREVISTOS O IMPREVISTOS PRODUCIDOS POR EL USO DE ESTA LICENCIA O DE LA OBRA, AUN CUANDO EL LICENCIANTE HAYA SIDO ADVERTIDO DE LA POSIBILIDAD DE DICHOS DAÑOS. ALGUNAS LEYES NO PERMITEN LA EXCLUSIÓN DE CIERTA RESPONSABILIDAD, EN CUYO CASO ESTA EXCLUSIÓN PUEDE NO APLICARSE A USTED.
7. Término.
a.	Esta Licencia y los derechos otorgados en virtud de ella terminarán automáticamente si Usted infringe alguna condición establecida en ella. Sin embargo, los individuos o entidades que han recibido Obras Derivadas o Colectivas de Usted de conformidad con esta Licencia, no verán terminadas sus licencias, siempre que estos individuos o entidades sigan cumpliendo íntegramente las condiciones de estas licencias. Las Secciones 1, 2, 5, 6, 7, y 8 subsistirán a cualquier terminación de esta Licencia.
b.	Sujeta a las condiciones y términos anteriores, la licencia otorgada aquí es perpetua (durante el período de vigencia de los derechos de autor de la obra). No obstante lo anterior, el Licenciante se reserva el derecho a publicar y/o estrenar la Obra bajo condiciones de licencia diferentes o a dejar de distribuirla en los términos de esta Licencia en cualquier momento; en el entendido, sin embargo, que esa elección no servirá para revocar esta licencia o que deba ser otorgada , bajo los términos de esta licencia), y esta licencia continuará en pleno vigor y efecto a menos que sea terminada como se expresa atrás. La Licencia revocada continuará siendo plenamente vigente y efectiva si no se le da término en las condiciones indicadas anteriormente.
8. Varios.
a.	Cada vez que Usted distribuya o ponga a disposición pública la Obra o una Obra Colectiva, el Licenciante ofrecerá al destinatario una licencia en los mismos términos y condiciones que la licencia otorgada a Usted bajo esta Licencia.
b.	Si alguna disposición de esta Licencia resulta invalidada o no exigible, según la legislación vigente, esto no afectará ni la validez ni la aplicabilidad del resto de condiciones de esta Licencia y, sin acción adicional por parte de los sujetos de este acuerdo, aquélla se entenderá reformada lo mínimo necesario para hacer que dicha disposición sea válida y exigible.
c.	Ningún término o disposición de esta Licencia se estimará renunciada y ninguna violación de ella será consentida a menos que esa renuncia o consentimiento sea otorgado por escrito y firmado por la parte que renuncie o consienta.
d.	Esta Licencia refleja el acuerdo pleno entre las partes respecto a la Obra aquí licenciada. No hay arreglos, acuerdos o declaraciones respecto a la Obra que no estén especificados en este documento. El Licenciante no se verá limitado por ninguna disposición adicional que pueda surgir en alguna comunicación emanada de Usted. Esta Licencia no puede ser modificada sin el consentimiento mutuo por escrito del Licenciante y Usted.

 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