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Title: Pushing peptides further: Novel methodologies for the synthesis of backbone-modified peptides
Author: Lobo Ruiz, Ariadna
Director/Tutor: Tulla-Puche, Judit
Keywords: Pèptids
Síntesi de pèptids
Peptide synthesis
Issue Date: 10-May-2019
Publisher: Universitat de Barcelona
Abstract: [eng] Peptides and proteins are essential substances for living organisms, as they can be found in every cell and tissue and are involved in many biological and physiological processes. Given their intrinsic properties and their attractive pharmacological profile, they have emerged as potential tools for drug discovery. However, in vivo instability due to protease degradation and poor bioavailability are the main drawbacks that have hampered their exploitation as therapeutic agents. Of special interest are peptide-based molecules displaying peptide backbone modifications, since they often result in improved pharmacological properties, such as greater stability and bioavailability, enhanced cell permeability and lower toxicity. Among the most relevant types of bioactive backbone-modified peptide families, depsipeptides and stapled peptides are included. In this context, the work presented herein was focused on the development of novel methodologies for the synthesis of depsipeptides and stapled peptides. Up to date, the most general and effective strategy for the preparation of complex cyclodepsipeptides combines solid-phase synthesis and solution chemistry approaches, in which segment condensation is used for the assembly of the building blocks containing the depsipeptide moieties. However, this methodology presents some disadvantages. For instance, the synthetic route must be designed and optimised for each particular case, and therefore a versatile general synthetic method cannot be outlined. Thus, a robust full solid-phase methodology would become a valuable chemical tool for both the preparation of naturally-occurring cyclodepsipeptides and the rapid generation of synthetic analogues. With that purpose, a synthetic analogue of naturally-occurring cyclodepsipeptide YM-254890 was used as a model depsipeptide for the development of such methodology, where the drawbacks commonly encountered during solid-phase depsipeptide synthesis, including: DKP formation, formation of undesired α,β-elimination side-products during Fmoc removal and selection of the optimal protecting group scheme, were extensively studied and solved. Additionally, evaluation of the strategy efficiency was carried out by comparison with conventional segment condensation approaches. Remarkably, similar overall yields as the ones obtained for segment condensation approaches were observed. Thus, the newly developed methodology becomes a versatile and convenient tool for the preparation of complex cyclodepsipeptides. On the other hand, great efforts have been put into the generation of novel stapled peptides mimicking α-helices. Although extensive research has been carried out in this field, a single universal stapling technique cannot be established, since selection of the most suitable cross-linking approach highly depends on the nature of the protein-protein interaction to be addressed. Nevertheless, the ability of stapled peptides to cross the cell membrane, increase in vivo stability and exhibit improved biological activity, has gained raising interest over the past years. It is well known, that backbone N-modified peptides exhibit greater lipophilicity, which ultimately results in enhanced cell internalisation. Additionally, N-modified peptides present higher resistance against proteolytic degradation. Considering these benefits, we envisioned that insertion of N-methyl-rich peptide bridges would be a good approach to develop a novel class of stapled peptides with an enhanced pharmacokinetic profile. With that purpose, the second part of this work was addressed at the design and development of a novel class of single and double highly N-methylated stapled peptides, or the so-called HMSP. Due to the importance of the p53 tumour suppressor, which activates cell death in response to various stress conditions, the p53-MDM2 protein- protein interaction was the focus in the development of our synthetic methodology. To linear p53-based peptides presenting a random coil secondary structure, several N- methyl-rich peptide bridges of different nature and length were inserted at different positions. Insertion of these staple entities makes these molecular constructs highly versatile, as the nature, length and flexibility of the staple can be modulated by the number and nature of NMe-amino acids. Circular dichroism experiments confirmed that helicity was induced, and allowed evaluation of the helicity increase for each system.
[spa] Dadas sus propiedades intrínsecas y su atractivo perfil farmacológico, los péptidos y las proteínas, surgen como potenciales agentes terapéuticos para el descubrimiento de nuevos fármacos. No obstante, su explotación en este campo ha sido obstaculizada por su escasa estabilidad y biodisponibilidad. De especial interés, son las modificaciones en el esqueleto peptídico, ya que a menudo dan lugar a propiedades farmacológicas mejoradas, tales como una mayor estabilidad, biodisponibilidad, permeabilidad celular y una menor toxicidad. La presente tesis se centra en los depsipéptidos y péptidos grapa, que se encuentran entre las familias de péptidos más relevantes que presentan modificaciones en el esqueleto peptídico. En el primer proyecto, se desarrolló una metodología robusta en fase sólida, basada en la estrategia Fmoc, para la preparación de depsipéptidos que contienen enlaces éster múltiples y consecutivos. Para ello, un análogo sintético del ciclodepsipéptido natural YM-254890 se designó como modelo para estudiar los inconvenientes comúnmente encontrados en la síntesis de depsipéptidos en fase sólida, que a día de hoy, han obstaculizado la instauración de dicha metodología. Por consiguiente, dificultades como la formación de DKP, la eliminación problemática de Fmoc, la selección del esquema de grupos protectores, entre otras, fueron ampliamente estudiadas y resueltas. En el segundo proyecto, con el fin de inducir la conformación hélice alfa, a la vez que preparar compuestos con un perfil farmacológico mejorado, se abordó el diseño y desarrollo de una nueva clase de péptidos grapa altamente N-metilados, los denominados HMSP. Para ello, entre dos posiciones clave de la secuencia bioactiva del péptido p53, que naturalmente no presenta una estructura secundaria organizada, se introdujeron varios péptidos puente ricos en residuos N-metilo. La estructura hélice alfa fue inducida satisfactoriamente. Por otro lado, la inserción de dichos puentes dio lugar a la preparación de construcciones moleculares altamente versátiles, dado que la naturaleza, la longitud y la flexibilidad del puente pueden ser moduladas por el número y la naturaleza de los aminoácidos N-metilados.
Appears in Collections:Tesis Doctorals - Departament - Química Inorgànica i Orgànica

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