Caged-carvedilol as a new tool for visible-light photopharmacology of β-adrenoceptors in native tissues

Biochemistry; Cell biology; PharmacologyBioquímica; Biologia cel·lular; FarmacologiaBioquímica; Biología celular; FarmacologíaAdrenoceptors are G protein-coupled receptors involved in a large variety of physiological processes, also under pathological conditions. This is due in large part to their ubiquitous expression in the body exerting numerous essential functions. Therefore, the possibility to control their activity with high spatial and temporal precision would constitute a valuable research tool. In this study, we present a caged version of the approved non-selective β-adrenoceptor antagonist carvedilol, synthesized by alkylation of its secondary amine with a coumarin derivative. Introducing this photo-removable group abolished carvedilol physiological effects in cell cultures, mouse isolated perfused hearts and living zebrafish larvae. Only after visible light application, carvedilol was released and the different physiological systems were pharmacologically modulated in a similar manner as the control drug. This research provides a new photopharmacological tool for a wide range of research applications that may help in the development of future precise therapies.This work was supported by ERDF-FEDER European Fund (projects CTQ2017-89222-R) and by the Catalan government (2017SGR 1604) to AL. Ministerio de Ciencia e Innovación, Agencia Estatal de Investigación (PID2020-120499RB-I00) supported XR and AL. XR research was financed by the Spanish Ministry of Economy, Industry and Competitiveness (SAF2015-74132-JIN). MF was supported by the “Agencia Estatal deInvestigación” from the Spanish Ministry of Science and Innovation and the IDAEA-CSIC, a Centre of Excellence Severo Ochoa (CEX2018-000794-S). ARS has a consolidated Miguel Servet contract and was financed by by the Catalan government (2017-SGR-1807). ADC received the support of a fellowship from “la Caixa” Foundation (ID 100010434) under the fellowship codeLCF/BQ/DE18/11670012

Caged-carvedilol as a new tool for visible-light photopharmacology of β-adrenoceptors in native tissues

Adrenoceptors are G protein-coupled receptors involved in a large variety of physiological processes, also under pathological conditions. This is due in large part to their ubiquitous expression in the body exerting numerous essential functions. Therefore, the possibility to control their activity with high spatial and temporal precision would constitute a valuable research tool. In this study, we present a caged version of the approved non-selective β-adrenoceptor antagonist carvedilol, synthesized by alkylation of its secondary amine with a coumarin derivative. Introducing this photo-removable group abolished carvedilol physiological effects in cell cultures, mouse isolated perfused hearts and living zebrafish larvae. Only after visible light application, carvedilol was released and the different physiological systems were pharmacologically modulated in a similar manner as the control drug. This research provides a new photopharmacological tool for a wide range of research applications that may help in the development of future precise therapies.We thank Maria José Bleda (IQAC-CSIC, Barcelona), Ignacio Pérez (IQAC-CSIC, Barcelona), Yolanda Pérez (IQAC-CSIC, Barcelona) and Carme Serra (SimChemSiMChem, IQAC-CSIC, Barcelona) for technical support. We thank Dr. Kees Jalink (The Netherlands Cancer Institute, Amsterdam, the Netherlands) for providing the plasmids encoding for the Epac-SH188 biosensor. We thank the University of Vic-Central University of Catalonia (UVic-UCC), Dr. Malu Calle and Dr. Marta Otero for the material assignment which helped in some biological assays. This work was supported by ERDF-FEDER European Fund (projects CTQ2017-89222-R) and by the Catalan government (2017SGR 1604) to AL. Ministerio de Ciencia e Innovación, Agencia Estatal de Investigación (PID2020-120499RB-I00) supported XR and AL. XR research was financed by the Spanish Ministry of Economy, Industry and Competitiveness (SAF2015-74132-JIN). MF was supported by the “Agencia Estatal deInvestigación” from the Spanish Ministry of Science and Innovation and the IDAEA-CSIC, a Centre of Excellence Severo Ochoa (CEX2018-000794-S). ARS has a consolidated Miguel Servet contract and was financed by by the Catalan government (2017-SGR-1807). ADC received the support of a fellowship from “la Caixa” Foundation (ID 100010434) under the fellowship codeLCF/BQ/DE18/11670012.Peer reviewe

Optical control of pain in vivo with a photoactive mGlu5 receptor negative allosteric modulator

Light-operated drugs constitute a major target in drug discovery, since they may provide spatiotemporal resolution for the treatment of complex diseases (i.e. chronic pain). JF-NP-26 is an inactive photocaged derivative of the metabotropic glutamate type 5 (mGlu5) receptor negative allosteric modulator raseglurant. Violet light illumination of JF-NP-26 induces a photochemical reaction prompting the active-drug's release, which effectively controls mGlu5 receptor activity both in ectopic expressing systems and in striatal primary neurons. Systemic administration in mice followed by local light-emitting diode (LED)-based illumination, either of the thalamus or the peripheral tissues, induced JF-NP-26-mediated light-dependent analgesia both in neuropathic and in acute/tonic inflammatory pain models. These data offer the first example of optical control of analgesia in vivo using a photocaged mGlu5 receptor negative allosteric modulator. This approach shows potential for precisely targeting, in time and space, endogenous receptors, which may allow a better management of difficult-to-treat disorders

Total synthesis of C<inf>37</inf> alken-2-one temperature geomarkers

The total synthesis of the natural (8E,15E,22E)-heptatriaconta-8,15,22-trien-2-one and (15E,22E)-heptatriaconta-15,22-dien-2-one is reported. Our synthesis involved a modular and convergent approach, based on the copper-catalyzed addition of Grignard reagents to allylic olefins. The obtention of these methyl ketones is an important asset for the use of these compounds as analytical patterns with application in Earth sciences.We gratefully acknowledge the Geochemistry and Pollution Research Group from the Institute of Environmental Assessment and Water Research (IDAEA-CSIC) and CSIC (201980E011) for financial support.Peer reviewe

Caged-carvedilol as a new tool for visible-light photopharmacology of β-adrenoceptors in native tissues

Adrenoceptors are G protein-coupled receptors involved in a large variety of physiological processes, also under pathological conditions. This is due in large part to their ubiquitous expression in the body exerting numerous essential functions. Therefore, the possibility to control their activity with high spatial and temporal precision would constitute a valuable research tool. In this study, we present a caged version of the approved non-selective β-adrenoceptor antagonist carvedilol, synthesized by alkylation of its secondary amine with a coumarin derivative. Introducing this photo-removable group abolished carvedilol physiological effects in cell cultures, mouse isolated perfused hearts and living zebrafish larvae. Only after visible light application, carvedilol was released and the different physiological systems were pharmacologically modulated in a similar manner as the control drug. This research provides a new photopharmacological tool for a wide range of research applications that may help in the development of future precise therapies

Development and validation of a mass spectrometry binding assay for mGlu5 receptor

International audienceMass spectrometry (MS) binding assays are a label-free alternative to radioligand or fluorescence binding assays, so the readout is based on direct mass spectrometric detection of the test ligand. The study presented here describes the development and validation of a highly sensitive, rapid, and robust MS binding assay for the quantification of the binding of the metabotropic glutamate 5 (mGlu5) negative allosteric modulator (NAM), MPEP (2-methyl-6-phenylethynylpyridine) at the mGlu5 allosteric binding site. The LC-ESI-MS/MS (liquid chromatography-electrospray ionization-tandem mass spectrometric) analytical method was established and validated with a deuterated analogue of MPEP as an internal standard. The developed MS binding assay described here allowed for the determination of MS binding affinity estimates that were in agreement with affinity estimates obtained from a tritiated MPEP radioligand saturation binding assay, indicating the suitability of this methodology for determining affinity estimates for compounds that target mGlu5 allosteric binding sites

Mechanistic insights into light-driven allosteric control of GPCR biological activity

International audienceG protein-coupled receptors (GPCR), including the metabotrobic glutamate 5 receptor (mGlu5), are important therapeutic targets and the development of allosteric ligands for targeting GPCRs has become a desirable approach toward modulating receptor activity. Traditional pharmacological approaches toward modulating GPCR activity are still limited since precise spatiotemporal control of a ligand is lost as soon as it is administered. Photopharmacology proposes the use of photoswitchable ligands to overcome this limitation, since their activity can be reversibly controlled by light with high precision. As this is still a growing field, our understanding of the molecular mechanisms underlying the light-induced changes of different photoswitchable ligand pharmacology is suboptimal. For this reason, we have studied the mechanisms of action of alloswitch-1 and MCS0331; two freely diffusible, mGlu5 phenylazopyridine photoswitchable negative allosteric modulators. We combined photochemical, cell-based, and in vivo photopharmacological approaches to investigate the effects of trans–cis azobenzene photoisomerization on the functional activity and binding ability of these ligands to the mGlu5 allosteric pocket. From these results, we conclude that photoisomerization can take place inside and outside the ligand binding pocket, and this leads to a reversible loss in affinity, in part, due to changes in dissociation rates from the receptor. Ligand activity for both photoswitchable ligands deviates from high-affinity mGlu5 negative allosteric modulation (in the trans configuration) to reduced affinity for the mGlu5 in their cis configuration. Importantly, this mechanism translates to dynamic and reversible control over pain following local injection and illumination of negative allosteric modulators into a brain region implicated in pain control

A Novel Family of Lysosomotropic Tetracyclic Compounds for Treating Leukemia

In spite of the recent expansion of the therapeutic landscape for acute myeloid leukemia (AML), resistance mechanisms and relapsed disease still pose a serious barrier to achieve curation for most patients. Considering the high inter- and intrapatient heterogeneity, disruptive therapeutic approaches are expected to provide a clinical solution for this unmet need. An in silico drug discovery program identified a new family of lysosome- and mitochondria-targeting compounds that specifically eradicate leukemia ex vivo and in vivo in relevant preclinical models by inducing both mitochondrial damage and apoptosis, and the simultaneous lysosomal membrane leakiness. Moreover, the compounds are effective in a wide panel of cancer cell lines, as their mechanism of action targets a common neoplastic feature. These compounds possess adequate pharmacological properties rendering them promising drug candidates for AML and unrelated neoplasias, and support their further clinical development. Acute myeloid leukemia (AML) is a heterogeneous hematological cancer characterized by poor prognosis and frequent relapses. Aside from specific mutation-related changes, in AML, the overall function of lysosomes and mitochondria is drastically altered to fulfill the elevated biomass and bioenergetic demands. On the basis of previous results, in silico drug discovery screening was used to identify a new family of lysosome-/mitochondria-targeting compounds. These novel tetracyclic hits, with a cationic amphiphilic structure, specifically eradicate leukemic cells by inducing both mitochondrial damage and apoptosis, and simultaneous lysosomal membrane leakiness. Lysosomal leakiness does not only elicit canonical lysosome-dependent cell death, but also activates the terminal differentiation of AML cells through the Ca 2+ -TFEB-MYC signaling axis. In addition to being an effective monotherapy, its combination with the chemotherapeutic arsenic trioxide (ATO) used in other types of leukemia is highly synergistic in AML cells, widening the therapeutic window of the treatment. Moreover, the compounds are effective in a wide panel of cancer cell lines and possess adequate pharmacological properties rendering them promising drug candidates for the treatment of AML and other neoplasias

Optical control of pain in vivo with a photoactive mGlu receptor negative allosteric modulator

Altres ajuts: ICREA (ICREA Academia-2010), Fundació la Marató de TV3 (Grant 20152031) and IWT (SBO-140028).Light-operated drugs constitute a major target in drug discovery, since they may provide spatiotemporal resolution for the treatment of complex diseases (i.e. chronic pain). JF-NP-26 is an inactive photocaged derivative of the metabotropic glutamate type 5 (mGlu) receptor negative allosteric modulator raseglurant. Violet light illumination of JF-NP-26 induces a photochemical reaction prompting the active-drug's release, which effectively controls mGlu receptor activity both in ectopic expressing systems and in striatal primary neurons. Systemic administration in mice followed by local light-emitting diode (LED)-based illumination, either of the thalamus or the peripheral tissues, induced JF-NP-26-mediated light-dependent analgesia both in neuropathic and in acute/tonic inflammatory pain models. These data offer the first example of optical control of analgesia in vivo using a photocaged mGlu receptor negative allosteric modulator. This approach shows potential for precisely targeting, in time and space, endogenous receptors, which may allow a better management of difficult-to-treat disorders