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Title: Development and applications of photoswitchable muscarinic ligands
Author: Riefolo, Fabio
Director/Tutor: Gorostiza Langa, Pablo Ignacio
Keywords: Farmacologia
Receptors colinèrgics
Lligands (Bioquímica)
Trastorns cognitius
Malalties del cor
Acetylcholine receptors
Ligands (Biochemistry)
Cognition disorders
Heart diseases
Issue Date: 29-Jul-2020
Publisher: Universitat de Barcelona
Abstract: [eng] Modern medical treatments are largely based on the ability of pharmacological drugs to interact with a molecular target in the human body (e.g., receptors) in order to evoke a physiological response. Such targets are expressed throughout the human body in both healthy and diseased tissues: this hampers a truly selective interaction between drug and target in a disease state, giving rise to undesired effects and limiting the effective dose at the desired site. Light-regulated drugs are small molecules that can be tested, validated, and approved using standard drug development procedures, and can be applied directly to wildtype organisms, including humans. This medical approach is known as photopharmacology and offers various benefits for new pharmacological treatments. A reversible photoisomerisable drug can be controlled with light over the body, dramatically improving its selectivity for a target expressed in a specific location and reducing the side effects to the minimal expression. The clinical pharmacology of the muscarinic cholinergic system has important limitations. The complex roles of muscarinic receptors (mAChRs) need further studies to be completely understood. They belong to the class A of G-protein coupled receptors (GPCRs) and are pharmacologically classified into five subtypes (M1-M5). Muscarinic subtype-selective bioactive compounds can potentially be very effective for therapies against Alzheimer's and Parkinson's disease, asthma, pain, intestinal motility disorders, heart disease, and urinary function disorders. Despite several decades of efforts on identifying novel muscarinic agonists and antagonists, their full therapeutic potential is not yet in our hands. The cause is mainly the lack of subtype selectivity of these drugs. Alternative approaches for achieving such subtype specificity are necessary. The success of this challenge can provide a revolution of the pharmacology against a large variety of diseases and disorders, as well as advancing the current knowledge of the metabotropic role of the ACh. This Ph.D. project aims to rational design and develop light-regulated ligands that can enable the optical modulation of muscarinic receptors and the physiological processes in which they are involved. Iperoxo is a charming ligand for mAChRs. Despite its atypical structure compared to acetylcholine, this drug is one of the most potent muscarinic agonists. However, its pharmacological profile is lacking subtype selectivity. Here, we designed, synthesized, and characterized the pharmacological profile of monovalent, bivalent, and dualsteric photoswitchable derivatives of Iperoxo. We tested their activity on the mAChRs subtypes M1 and M2. Among the others, the light-regulated dualsteric drug named Phthalimide-Azobenzene-Iperoxo (PAI) turned out to be a potent photoswitchable activator of M2 receptors. This compound demonstrates to effectively modulate the cardiac function with light in vivo in different wild type animal models, standing as a potential alternative for classical antiarrhythmic drugs that cannot be manipulated spatiotemporally. Moreover, PAI administration in the central nervous system (isolated cortical slices and in anesthetized living mice) can manipulate brain state transitions with light, a function that bears high therapeutic interest in several neurological disorders. In order to overcome the scattering and low penetration of violet and visible illumination, we effectively demonstrated that PAI can activate M2 receptors using two-photon excitation with near-infrared light, offering new therapeutic opportunities for this photopharmacological tool. Because of their therapeutic relevance, we also developed photoswitchable M1 antagonists. We used the tricyclic M1 antagonist pirenzepine as model compound. We designed the new strategy of “crypto-azologization”, replacing the three-ringed core of pirenzepine with different azobenzene scaffolds. We validated this novel design by inhibiting M1 receptors only upon illumination in vitro and in cardiac atria ex vivo. This “crypto-azologization” strategy has the potential to be used for controlling the action of many other mainstream tricyclic drugs with light.
[spa] Los receptores muscarínicos (mAChR) son receptores acoplados a proteína G (GPCRs) de clase A y están clasificados en cinco subtipos (M1-M5) que modulan numerosas respuestas fisiológicas centrales y periféricas. Los medicamentos muscarínicos podrían ser muy efectivos en muchas terapias, como para los trastornos cognitivos y las enfermedades cardíacas, pero su baja selectividad de subtipo y sus efectos secundarios obstaculizan el potencial terapéutico de estos fármacos. La fotofarmacología crea medicamentos que se pueden activar con luz de manera reversible, mejorando su selectividad en un sitio específico del cuerpo y ofreciendo la posibilidad de reducir sus efectos secundarios. Con el fin de introducir alternativas para mejorar la especificidad de los fármacos muscarínicos, hemos desarrollados ligandos fotoconmutables para modular con luz los receptores muscarínicos y los procesos fisiológicos controlados por ellos. Hemos diseñado y creado diferentes derivados fotosensibles de Iperoxo, un agonista muscarínico muy potente. Entre todos, el derivado PAI resultó ser un potente activador de los receptores M2 controlado por luz. Con este compuesto hemos demostrado el fotocontrol de la actividad cardíaca in vivo en diferentes modelos animales, creando una potencial alternativa a los fármacos antiarrítmicos clásicos que no se pueden controlar espaciotemporalmente. La administración de PAI en el sistema nervioso central de ratones vivos anestesiados permite fotomanipular las transiciones entre diferentes actividades oscilatorias neuronales. Esta función que tiene un gran interés terapéutico porque ofrece la posibilidad de controlar varios trastornos neurológicos. PAI se puede activar con la luz infrarroja (840 nm) mediante la excitación de dos fotones, y estas longitudes de onda permiten penetrar profundamente en los tejidos biológicos, minimizando los daños y dando lugar a aplicaciones no invasivas in vivo. Debido a su relevancia terapéutica, también hemos desarrollados derivados fotoconmutables de la pirenzepina, un antagonista tricíclico de los receptores M1. Hemos reemplazado el heterotriciclo del compuesto original con diferentes azobencenos, creando una nueva estrategia para diseñar análogos de fármacos fotoconmutables que hemos denominado "cripto-azologización". Hemos validado este nuevo concepto mediante el fotocontrol de la inhibición de los receptores M1 en las aurículas cardiacas ex vivo. Este método ofrece el potencial de fotocontrolar la acción de muchos fármacos tricíclicos convencionales para realizar estudios funcionales de sus receptores in vivo y para reducir los efectos secundarios de las terapias farmacológicas actuales.
Appears in Collections:Tesis Doctorals - Facultat - Química

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