Peptides: from synthesis to biomedical application in two types of cancer
[eng] The aim of this work was to synthesize peptides that would perform two functions in the potential treatment of two types of cancer. The first group of peptides was used as the antigenic component of a nanovaccine formulation that represents an immunotherapeutic approach to treating pancreatic...
| Autor: | |
|---|---|
| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2024 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:diposit.ub.edu:2445/216676 |
| Acceso en línea: | https://hdl.handle.net/2445/216676 http://hdl.handle.net/10803/692591 |
| Access Level: | acceso abierto |
| Palabra clave: | Pèptids Síntesi de pèptids Vacunes Antígens Peptides Peptide synthesis Vaccines Antigens |
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España |
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Peptides: from synthesis to biomedical application in two types of cancer |
| title |
Peptides: from synthesis to biomedical application in two types of cancer |
| spellingShingle |
Peptides: from synthesis to biomedical application in two types of cancer Luna González, Omar Fernando Pèptids Síntesi de pèptids Vacunes Antígens Peptides Peptide synthesis Vaccines Antigens |
| title_short |
Peptides: from synthesis to biomedical application in two types of cancer |
| title_full |
Peptides: from synthesis to biomedical application in two types of cancer |
| title_fullStr |
Peptides: from synthesis to biomedical application in two types of cancer |
| title_full_unstemmed |
Peptides: from synthesis to biomedical application in two types of cancer |
| title_sort |
Peptides: from synthesis to biomedical application in two types of cancer |
| dc.creator.none.fl_str_mv |
Luna González, Omar Fernando |
| author |
Luna González, Omar Fernando |
| author_facet |
Luna González, Omar Fernando |
| author_role |
author |
| dc.contributor.none.fl_str_mv |
Royo Expósito, Miriam Albericio Palomera, Fernando Universitat de Barcelona. Departament de Química Orgànica |
| dc.subject.none.fl_str_mv |
Pèptids Síntesi de pèptids Vacunes Antígens Peptides Peptide synthesis Vaccines Antigens |
| topic |
Pèptids Síntesi de pèptids Vacunes Antígens Peptides Peptide synthesis Vaccines Antigens |
| description |
[eng] The aim of this work was to synthesize peptides that would perform two functions in the potential treatment of two types of cancer. The first group of peptides was used as the antigenic component of a nanovaccine formulation that represents an immunotherapeutic approach to treating pancreatic ductal adenocarcinoma. In some cases, the peptides were modified at the N-terminus through palmitoylation and PEGylation, with the objective of enhancing their immunogenic potential. Additionally, they were synthesized as single epitopes or as multi-epitope constructs derived from tumor-associated antigen proteins. The peptides were formulated in poly(lactic-co-glycolic) acid-based nanoparticles, and the resulting nanoformulation was tested in a mouse model to assess its immunogenic activity. Two multiepitope peptides demonstrated a markedly positive response in vitro. These were the Palmitoyl-PLTVAEVQKLLGPHVKKALPLDLLLFLKKSLLFLLFSL-NH2 peptide and the H-KVYLRVRPLLKKSYGVLLWEIKKRFVPDGNRI-NH2 peptide. These findings suggest that long multi-epitope constructs are the most effective alternative for use as nanovaccine components, as single epitope peptides were demonstrated to lack immunogenicity. However, preliminary in vivo assays of the two multi-epitope peptides exhibited minimal activity against the tumor in a mouse model. Additional validation is necessary through the repetition of these assays. The second group of peptides served as targeting units in a quatsome nanovesicle delivery system that is designed to carry a therapeutic nucleic acid for the treatment of neuroblastoma. The peptides were initially synthesized with fluorescein as a probe. In parallel, a small molecule ligand, a thiolated p-aminobenzylguanidine derivative, was also synthesized, labeled and evaluated in conjunction with the targeting peptide moieties to determine their internalization capability in a neuroblastoma cell line. A sequence targeting the GD2 receptor in neuroblastoma cells (H-WHWRLPSGGGC-NH2) and the thiolated p-aminobenzylguanidine derivative, demonstrated the greatest capacity to internalize into these cells and were therefore selected for the development of a conjugation methodology in quatsome nanovesicles using a thiol-maleimide click reaction. The methodology was successfully developed, and the optimal conditions were identified as a pH of 7.5, a reaction time of two hours, the presence of a reducing agent and a clean-up methodology of size exclusion chromatography in Sephadex G50 and aqueous elution followed by mild acidic elution. This allowed for the separation of nanovesicles from unreacted ligands and the indirect estimation of the conjugated targeting moiety in the nanovesicle. The methodology yielded conjugation estimates of 50% to 65% for both the GD2-binding peptide and the thiolated p-aminobenzylguanidine derivative. Furthermore, this formulation was demonstrated to have the capacity to deliver a nucleic acid to a neuroblastoma cell line. However, a switch of the PEGyl moiety carrying the maleimide function from PEG2000 to PEG1000 is required to achieve quantitative internalization. Furthermore, a study was conducted to evaluate the suitability of five different carbodiimides for use in solid-phase peptide synthesis, the methodology employed for the production of all peptide compounds in this research. The objective of this comparative study was to identify an optimal alternative to N,N'-diisopropylcarbodiimide (DIC) that can prevent the formation of the toxic compound hydrogen cyanide, which can occur when the reaction is conducted in the presence of oxyma. The study demonstrated that 1-tert-butyl-3-ethylcarbodiimide is an effective alternative to DIC. It exhibited comparable synthetic performance in the production of two peptide models and an antigenic peptide, while reducing the occurrence of hydrogen cyanide by threefold compared to DIC. |
| publishDate |
2024 |
| dc.date.none.fl_str_mv |
2024 |
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info:eu-repo/semantics/doctoralThesis info:eu-repo/semantics/publishedVersion |
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doctoralThesis |
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publishedVersion |
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https://hdl.handle.net/2445/216676 http://hdl.handle.net/10803/692591 |
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https://hdl.handle.net/2445/216676 http://hdl.handle.net/10803/692591 |
| dc.language.none.fl_str_mv |
Inglés |
| language_invalid_str_mv |
Inglés |
| dc.rights.none.fl_str_mv |
cc by (c) Luna González, Omar Fernando, 2024 http://creativecommons.org/licenses/by/3.0/es/ info:eu-repo/semantics/openAccess |
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cc by (c) Luna González, Omar Fernando, 2024 http://creativecommons.org/licenses/by/3.0/es/ |
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openAccess |
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application/pdf |
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Universitat de Barcelona |
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Universitat de Barcelona |
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Tesis Doctorals - Departament - Química Orgànica reponame:Dipòsit Digital de la UB instname:Universidad de Barcelona |
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Universidad de Barcelona |
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Dipòsit Digital de la UB |
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Dipòsit Digital de la UB |
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1869421583997599744 |
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Peptides: from synthesis to biomedical application in two types of cancerLuna González, Omar FernandoPèptidsSíntesi de pèptidsVacunesAntígensPeptidesPeptide synthesisVaccinesAntigens[eng] The aim of this work was to synthesize peptides that would perform two functions in the potential treatment of two types of cancer. The first group of peptides was used as the antigenic component of a nanovaccine formulation that represents an immunotherapeutic approach to treating pancreatic ductal adenocarcinoma. In some cases, the peptides were modified at the N-terminus through palmitoylation and PEGylation, with the objective of enhancing their immunogenic potential. Additionally, they were synthesized as single epitopes or as multi-epitope constructs derived from tumor-associated antigen proteins. The peptides were formulated in poly(lactic-co-glycolic) acid-based nanoparticles, and the resulting nanoformulation was tested in a mouse model to assess its immunogenic activity. Two multiepitope peptides demonstrated a markedly positive response in vitro. These were the Palmitoyl-PLTVAEVQKLLGPHVKKALPLDLLLFLKKSLLFLLFSL-NH2 peptide and the H-KVYLRVRPLLKKSYGVLLWEIKKRFVPDGNRI-NH2 peptide. These findings suggest that long multi-epitope constructs are the most effective alternative for use as nanovaccine components, as single epitope peptides were demonstrated to lack immunogenicity. However, preliminary in vivo assays of the two multi-epitope peptides exhibited minimal activity against the tumor in a mouse model. Additional validation is necessary through the repetition of these assays. The second group of peptides served as targeting units in a quatsome nanovesicle delivery system that is designed to carry a therapeutic nucleic acid for the treatment of neuroblastoma. The peptides were initially synthesized with fluorescein as a probe. In parallel, a small molecule ligand, a thiolated p-aminobenzylguanidine derivative, was also synthesized, labeled and evaluated in conjunction with the targeting peptide moieties to determine their internalization capability in a neuroblastoma cell line. A sequence targeting the GD2 receptor in neuroblastoma cells (H-WHWRLPSGGGC-NH2) and the thiolated p-aminobenzylguanidine derivative, demonstrated the greatest capacity to internalize into these cells and were therefore selected for the development of a conjugation methodology in quatsome nanovesicles using a thiol-maleimide click reaction. The methodology was successfully developed, and the optimal conditions were identified as a pH of 7.5, a reaction time of two hours, the presence of a reducing agent and a clean-up methodology of size exclusion chromatography in Sephadex G50 and aqueous elution followed by mild acidic elution. This allowed for the separation of nanovesicles from unreacted ligands and the indirect estimation of the conjugated targeting moiety in the nanovesicle. The methodology yielded conjugation estimates of 50% to 65% for both the GD2-binding peptide and the thiolated p-aminobenzylguanidine derivative. Furthermore, this formulation was demonstrated to have the capacity to deliver a nucleic acid to a neuroblastoma cell line. However, a switch of the PEGyl moiety carrying the maleimide function from PEG2000 to PEG1000 is required to achieve quantitative internalization. Furthermore, a study was conducted to evaluate the suitability of five different carbodiimides for use in solid-phase peptide synthesis, the methodology employed for the production of all peptide compounds in this research. The objective of this comparative study was to identify an optimal alternative to N,N'-diisopropylcarbodiimide (DIC) that can prevent the formation of the toxic compound hydrogen cyanide, which can occur when the reaction is conducted in the presence of oxyma. The study demonstrated that 1-tert-butyl-3-ethylcarbodiimide is an effective alternative to DIC. It exhibited comparable synthetic performance in the production of two peptide models and an antigenic peptide, while reducing the occurrence of hydrogen cyanide by threefold compared to DIC.[spa] Este trabajo tuvo como objetivo sintetizar péptidos que pudiesen cumplir dos funciones en el tratamiento de dos tipos de cáncer. El primer grupo de péptidos es el componente antigénico en una formulación de nanovacuna terapéutica contra el adenocarcinoma ductal pancreático. Para incrementar su potencial inmunogénico, se incluyeron péptidos modificados en la amina N-terminal y se formularon en nanopartículas de ácido poli(láctico-co-glicólico). Las formulaciones se ensayaron en un modelo de ratón. Dos péptidos multiepitópicos: Palmitoil-PLTVAEVQKLLGPHVKKALPLDLLLFLKKSLLFLLFSL-NH2 y H-KVYLRVRPLLKKSYGVLLWEIKKRFVPDGNRI-NH2 mostraron una actividad inmunogénica óptima in vitro, lo que sugiere que los péptidos largos multiepitópicos son una excelente opción como componentes de vacunas. Ensayos in vivo, sin embargo, demostraron poca actividad antitumoral. El segundo grupo de péptidos son unidades de direccionamiento en un sistema de entrega tipo quatsoma, que porta un ácido nucleico terapéutico para el tratamiento del neuroblastoma. Estos péptidos se sintetizaron como sondas fluorescentes introduciendo fluoresceína. Conjuntamente, se sintetizó una molécula orgánica, un derivado tiolado de p-aminobencilguanidinio. Estos compuestos se ensayaron en su capacidad de acumulación en células de neuroblastoma. El péptido H-WHWRLPSGGGC-NH2 y el derivado tiolado de p-aminobencilguanidinio fueron seleccionados por su excelente actividad y se desarrolló una metodología de conjugación de estas unidades en quatsomas, alcanzando tasas de conjugación de entre el 50 % y el 65 %. Además, se demostró que esta formulación puede entrar en una línea celular de neuroblastoma y entregar un ácido nucleico siempre que la función maleimida se introduzca en un PEG1000. Todos los péptidos se sintetizaron mediante síntesis en fase sólida. Se realizó un estudio sobre la reacción de acoplamiento de esta metodología, en el que se compararon cinco carbodiimidas, con el fin de evaluar la mejor alternativa a la N,N’-diisopropilcarbodiimida (DIC), tanto en cuanto a la calidad de la síntesis como a la minimización de una reacción secundaria de la DIC en presencia de oxyma: la formación de cianuro de hidrógeno. Se demostró que la 1-tert-butil-3-etil-carbodiimida cumple ambos criterios y puede usarse para este fin.Universitat de BarcelonaRoyo Expósito, MiriamAlbericio Palomera, FernandoUniversitat de Barcelona. Departament de Química Orgànica2024info:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/publishedVersionapplication/pdfhttps://hdl.handle.net/2445/216676http://hdl.handle.net/10803/692591Tesis Doctorals - Departament - Química Orgànicareponame:Dipòsit Digital de la UBinstname:Universidad de BarcelonaIngléscc by (c) Luna González, Omar Fernando, 2024http://creativecommons.org/licenses/by/3.0/es/info:eu-repo/semantics/openAccessoai:diposit.ub.edu:2445/2166762026-05-27T06:46:51Z |
| score |
15,812455 |