Search

Retinoic acid receptor-related orphan receptor γ (RORγ) is a nuclear hormone receptor with multiple biological functions in circadian clock regulation, inflammation, and immunity. Its cyclic temporal role in circadian rhythms, and cell-specific activity in the immune system, make it an intriguing target for spatially and temporally localised pharmacology. To create tools that can study RORγ biology with appropriate spatiotemporal resolution, we designed light-dependent inverse RORγ agonists by building azobenzene photoswitches into ligand consensus structures. Optimizations gave photoswitchable RORγ inhibitors combining a large degree of potency photocontrol, with remarkable on-target potency, and excellent selectivity over related off-target receptors. This still rare combination of performance features distinguishes them as high quality photopharmaceutical probes, which can now serve as high precision tools to study the spatial and dynamic intricacies of RORγ action in signaling and in inflammatory disorders., (© 2024 The Author(s). Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

AMPA receptors (AMPARs) are the main drivers of excitatory glutamatergic transmission in the brain, central to synaptic plasticity, and are key drug targets. However, AMPARs are expressed in virtually every neuron in the central nervous system and are activated with complex temporal dynamics, making it difficult to determine their functional roles with sufficient precision. Here we describe a cell specific, light-controllable competitive antagonist for the AMPA receptor called MP-GluA block that combines the temporal precision of a photo-switchable ligand with the spatial and cellular specificity of a genetically-encoded membrane-anchor protein. This tool could pave the way for controlling endogenous AMPARs in neural circuits with cellular, spatial, and temporal specificity., (© 2024 The Author(s). Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Control of biological activity with light is a fascinating idea. "Caged" compounds, molecules modified with photolabile protecting group, are one of the instruments for this purpose. Adrenergic receptors are essential regulators of neuronal, endocrine, cardiovascular, vegetative, and metabolic functions. These receptors are largely used as pharmacologic targets. Photolabile "caged" analogs of adrenergic receptor agonists has been reported more than 30 years ago. We report that the photolysis of epinephrine analogs, apart from liberation of the epinephrine, is accompanied by a formation of significant amount of adrenochrome, a compound with neuro- and cardiotoxic effect., Competing Interests: Declarations. Conflict of interest: The authors declare that they have no conflict of interest., (© 2024. The Author(s), under exclusive licence to the European Photochemistry Association, European Society for Photobiology.)

Photopharmacology, an emerging interdisciplinary field, enables precise modulation of target biological activity by employing light-controllable photoswitchable molecules. The research on nicotinamidase (Naam), a recently identified target molecule for pesticides, remains relatively underexplored. Therefore, based on the structure of its inhibitor Flonicamid, we designed and synthesized a series of photopharmacological ligands (FAB S ) by incorporating a light-controllable azobenzene switch, further investigating their effects on insect behavior and activity. Compound FAB03 exhibited an LC 50 value of 32.8 μM before light exposure and 154 μM after light exposure against Aedes albopictus , indicating a 4.7-fold difference in activity. Compound FAB05 exhibited an LC 50 value of 0.691 μM before light exposure and 2.85 μM after light exposure against nymphs (1 day old) of Aphis craccivora , indicating a 4.1-fold difference in activity. Additionally, this series of photopharmacological ligands has successfully achieved light-mediated modulation of locomotor behavior in both Drosophila melanogaster and A. albopictus larvae. Fluorescence staining experiments with A. albopictus larvae were conducted to explore potential modes of action of the drug. Calcium content analysis showed that Flonicamid at 20 μg/mL significantly caused an inward Ca 2+ flux in the ovarian of Spodoptera frugiperda (SF9), and the trans-configurations of the photopharmacological ligands FAB03 and FAB05 also led to a certain increase in the concentration of intracellular Ca 2+ . AlphaFold2 was employed to predict the three-dimensional structure of Naam, and potential binding modes were elucidated through molecular docking studies.

Introduction: Intensive and detailed investigations of the molecular function and cellular role of mammalian transient receptor potential canonical (TRPC) channels started back in the early 90 th of the past century. Initial TRPC research was fueled by high hopes to resolve fundamental questions of cellular Ca 2+ signaling. To date, we have learned important lessons in TRPC channel biology and biophysics, while little progress has been made in terms of therapeutic concepts., Areas Covered: This is a critical account of recent progress in building a solid mechanistic basis for therapeutic interventions utilizing TRPC3 as a target. I focus on hurdles and key issues to be resolved, thereby drafting a list of essential scientific 'to-dos' on the way toward drugging of TRPC3., Expert Opinion: Recent advances in the molecular physiology of TRPC3 include unveiling its lipid sensing machinery and the channels´ ability to serve spatiotemporally diverse Ca 2+ signaling in a cell type- and context-dependent manner. The ongoing development of technology for high-precision manipulation of the channel opens up a view on novel therapeutic strategies. It is now to unravel the complex role of TRPC3 in human physiopathology, to pinpoint the channels´ therapeutic relevance, and to further refine technologies for targeted interventions.

The field of G protein-coupled receptor (GPCR) research has greatly benefited from the spatiotemporal resolution provided by light controllable, i.e., photoswitchable ligands. Most of the developed tools have targeted the Rhodopsin-like family (Class A), the largest family of GPCRs. However, to date, all such Class A photoswitchable ligands were designed to act at the orthosteric binding site of these receptors. Herein, we report the development of the first photoswitchable allosteric modulators of Class A GPCRs, designed to target the M 1 muscarinic acetylcholine receptor. The presented benzyl quinolone carboxylic acid (BQCA) derivatives, Photo-BQCisA and Photo-BQCtrAns, exhibit complementary photopharmacological behavior and allow reversible control of the receptor using light as an external stimulus. This makes them valuable tools to further investigate M 1 receptor signaling and a proof of concept for photoswitchable allosteric modulators at Class A receptors., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Background and Purpose: Patients suffering from ischaemic heart disease and heart failure are at high risk of recurrent ventricular arrhythmias (VAs), eventually leading to sudden cardiac death. While high-voltage shocks delivered by an implantable defibrillator may prevent sudden cardiac death, these interventions themselves impair quality of life and raise both morbidity and mortality, which accentuates the need for developing novel defibrillation techniques., Experimental Approach: Photopharmacology allows for reversible control of biological processes by light. When relying on synthetic and externally applied chromophores, it renders genetic modification of target cells dispensable and may hence be advantageous over optogenetic approaches. Here, the photochromic ligand azobupivacaine 2 (AB2) was probed as a modulator of cardiac electrophysiology in an ex vivo intact mouse heart model., Key Results: By reversibly blocking voltage-gated Na + and K + channels, photoswitching of AB2 modulated both the ventricular effective refractory period and the conduction velocity in native heart tissue. Moreover, photoswitching of AB2 was able to convert VA into sinus rhythm., Conclusion and Implications: The present study provides the first proof of concept that AB2 enables gradual control of cardiac electrophysiology by light. AB2 may hence open the door to the development of an optical defibrillator based on photopharmacology., (© 2024 The Author(s). British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Small molecule modulators are powerful tools for selectively probing and manipulating proteins in native biological systems. However, the development of versatile modulators that exhibit desired properties is hindered by the lack of a rapid and robust synthetic strategy. Here, we develop a facile and reliable one-step methodology for the generation of multifunctional toolboxes encompassing a wide variety of chemical modulators with different desired features. These modulators bind irreversibly to the protein target via a selective warhead. Key elements are introduced onto the warhead in a single step using multi-component reactions. To illustrate the power of this new technology, we synthesized a library of diverse modulators designed to explore a highly challenging and poorly understood protein, human 15-lipoxygenase-1. Modulators made include; activity-based/photoaffinity probes, chemosensors, photocrosslinkers, as well as light-controlled and high-affinity inhibitors. The efficacy of our compounds was successfully established through the provision of on demand inhibition and labeling of our target protein in vitro, in cellulo and in vivo; thus, proving that this technology has promising potential for applications in many complex biological systems., (© 2024 The Author(s). Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

The cytoskeleton is essential for spatial and temporal organisation of a wide range of cellular and tissue-level processes, such as proliferation, signalling, cargo transport, migration, morphogenesis, and neuronal development. Cytoskeleton research aims to study these processes by imaging, or by locally manipulating, the dynamics and organisation of cytoskeletal proteins with high spatiotemporal resolution: which matches the capabilities of optical methods. To date, no photoresponsive microtubule-stabilising tool has united all the features needed for a practical high-precision reagent: a low potency and biochemically stable non-illuminated state; then an efficient, rapid, and clean photoresponse that generates a high potency illuminated state; plus good solubility at suitable working concentrations; and efficient synthetic access. We now present CouEpo, a photocaged epothilone microtubule-stabilising reagent that combines these needs. Its potency increases approximately 100-fold upon irradiation by violet/blue light to reach low-nanomolar values, allowing efficient photocontrol of microtubule dynamics in live cells, and even the generation of cellular asymmetries in microtubule architecture and cell dynamics. CouEpo is thus a high-performance tool compound that can support high-precision research into many microtubule-associated processes, from biophysics to transport, cell motility, and neuronal physiology., (© 2024 The Author(s). Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Elucidating the mechanisms by which protein synthesis contributes to complex biological processes has remained a challenging endeavor. This is particularly true in the field of neuroscience, where multiple, tightly regulated periods of new protein synthesis in different cell-types are thought to facilitate intricate neurological functions, such as memory formation. Current methods for labeling the de novo proteome have lacked the spatial and temporal resolution to accurately discriminate these overlapping and often competing windows of mRNA translation. To address this technological limitation, here we describe a novel, light-inducible specific method for labeling newly synthesized proteins within a targeted cell-type.By developing Opto-ANL, a photocaged version of the nonendogenous amino acid azidonorleucine (ANL), we can selectively label newly synthesized proteins in specific cell-types through the targeted expression of a mutant methionyl-tRNA synthetase (L274G-MetRS). We demonstrate that Opto-ANL can be rapidly uncaged by UV light treatment in both cell culture and mouse brain slices, with Opto-ANL labeled proteins being able to be visualized via fluorescent noncanonical amino acid tagging. We also reveal that pretreatment with Opto-ANL not only allows for the period of de novo proteomic labeling to be tightly controlled, but also improves labeling efficiency compared to regular ANL. To demonstrate the potential applications of this novel technique, we use Opto-ANL to detect insulin-induced increases in protein synthesis and to label the excitatory neuronal de novo proteome in mouse brain slices. We believe that this application of photopharmacology will allow researchers to generate novel insights into how the translational landscape is altered across cell-types during complex neurological phenomena such as memory formation.

Photo-switchable nuclear receptor modulators ("photohormones") enable spatial and temporal control over transcription factor activity and are valuable precision tools for biological studies. We have developed a new photohormone chemotype by incorporating a light-switchable motif in the scaffold of a cinalukast-derived PPARα ligand and tuned light-controlled activity by systematic structural variation. An optimized photohormone exhibited PPARα agonism in its light-induced (Z)-configuration and strong selectivity over related lipid-activated transcription factors representing a valuable addition to the collection of light-controlled tools to study nuclear receptor activity., (© 2024 The Authors. ChemMedChem published by Wiley-VCH GmbH.)

A set of arylazopyrazole-based inhibitors targeting the mitotic motor protein CENP-E was discovered through the chemical platform using the quantitative cyclization of 1,3-diketone intermediate with various hydrazines under mild conditions. Through this efficient platform, the structure-activity relationship pertaining to the pyrazole photoswitch in photoswitchable CENP-E inhibitors not only in vitro but also in cells was successfully clarified., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The development of light-responsive molecular tools enables spatiotemporal control of biochemical processes with superior precision. Amongst these molecular tools, photolabile caging groups are employed to prevent critical binding interactions between a bioactive molecule and its corresponding target. Only upon irradiation with light, the bioactive is released in its 'active' form and is now readily available to bind to its target. Coumarin-derived caging groups constitute one of the most popular classes of photolabile protecting groups, due to their facile synthetic accessibility, ease of tuning photophysical properties via structural modification and rapid photolysis reactions. Herein, we highlight the recent progress made on the development of coumarin-derived caging groups, in which the red-shifting of absorption spectra, improving aqueous solubility and tailoring sub-cellular localisation has been of particular interest., (© 2024 The Authors. ChemPlusChem published by Wiley-VCH GmbH.)

Photoregulation of biomolecules has become crucial tools in chemical biology, because light enables access under mild conditions and with delicate spatiotemporal control. The control of enzyme activity in a reversible way is a challenge. To achieve it, a facile approach is to use photoswitchable inhibitors. This review highlights recent progress in photoswitchable inhibitors based on azobenzenes units. The progress suggests that the incorporation of an azobenzene unit to a known inhibitor is an effective method for preparing a photoswitchable inhibitor, and with these photoswitchable inhibitors, the activity of enzymes can be regulated by optical control, which is valuable in both basic science and therapeutic applications.

Chemical photoswitches have become a widely used approach for the remote control of biological functions with spatiotemporal precision. Several molecular scaffolds have been implemented to improve photoswitch characteristics, ranging from the nature of the photoswitch itself (e.g. azobenzenes, dithienylethenes, hemithioindigo) to fine-tuning of aromatic units and substituents. Herein, we present deuterated azobenzene photoswitches as a general means of enhancing the performance of photopharmacological molecules. Deuteration can improve azobenzene performance in terms of light sensitivity (higher molar extinction coefficient), photoswitch efficiency (higher photoisomerization quantum yield), and photoswitch kinetics (faster macroscopic rate of photoisomerization) with minimal alteration to the underlying structure of the photopharmacological ligand. We report synthesized deuterated azobenzene-based ligands for the optimized optical control of ion channel and G protein-coupled receptor (GPCR) function in live cells, setting the stage for the straightforward, widespread adoption of this approach., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

A problem of systemic pharmacotherapy is off-target activity, which causes adverse effects. Outstanding examples include neuroinhibitory medications like antiseizure drugs, which are used against epilepsy and neuropathic pain but cause systemic side effects. There is a need of drugs that inhibit nerve signals locally and on-demand without affecting other regions of the body. Photopharmacology aims to address this problem with light-activated drugs and localized illumination in the target organ. Here, we have developed photoswitchable derivatives of the widely prescribed antiseizure drug carbamazepine. For that purpose, we expanded our method of ortho azologization of tricyclic drugs to meta/para and to N-bridged diazocine. Our results validate the concept of ortho cryptoazologs (uniquely exemplified by Carbazopine-1) and bring to light Carbadiazocine (8), which can be photoswitched between 400-590 nm light (using violet LEDs and halogen lamps) and shows good drug-likeness and predicted safety. Both compounds display photoswitchable activity in vitro and in translucent zebrafish larvae. Carbadiazocine (8) also offers in vivo analgesic efficacy (mechanical and thermal stimuli) in a rat model of neuropathic pain and a simple and compelling treatment demonstration with non-invasive illumination., (© 2024 The Author(s). Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Aim: Photopharmacology is a new technique for modulating biological phenomena through the photoconversion of substances in a specific target region at precise times. Caged compounds are thought to be compatible with photopharmacology as uncaged ligands are released and function in a light irradiation-dependent manner. Here, we investigated whether a microscale light-emitting diode (MicroLED) probe is applicable for the photoconversion of caged-glutamate (caged-Glu) in vivo., Methods: A needle-shaped MicroLED probe was fabricated and inserted into the mouse hippocampal dentate gyrus (DG) with a cannula for drug injection and a recording electrode for measuring the local field potential (LFP). Artificial cerebrospinal fluid (ACSF) or caged-Glu was infused into the DG and illuminated with light from a MicroLED probe., Results: In the caged-Glu-injected DG, the LFP changed in the 10-20 Hz frequency ranges after light illumination, whereas there was no change in the ACSF control condition., Conclusion: The MicroLED probe is applicable for photopharmacological experiments to modulate LFP with caged-Glu in vivo., (© 2024 The Author(s). Neuropsychopharmacology Reports published by John Wiley & Sons Australia, Ltd on behalf of The Japanese Society of Neuropsychopharmacology.)

We used light-sensitive drugs to identify the brain region-specific role of mGlu5 metabotropic glutamate receptors in the control of pain. Optical activation of systemic JF-NP-26, a caged, normally inactive, negative allosteric modulator (NAM) of mGlu5 receptors, in cingulate, prelimbic, and infralimbic cortices and thalamus inhibited neuropathic pain hypersensitivity. Systemic treatment of alloswitch-1, an intrinsically active mGlu5 receptor NAM, caused analgesia, and the effect was reversed by light-induced drug inactivation in the prelimbic and infralimbic cortices, and thalamus. This demonstrates that mGlu5 receptor blockade in the medial prefrontal cortex and thalamus is both sufficient and necessary for the analgesic activity of mGlu5 receptor antagonists. Surprisingly, when the light was delivered in the basolateral amygdala, local activation of systemic JF-NP-26 reduced pain thresholds, whereas inactivation of alloswitch-1 enhanced analgesia. Electrophysiological analysis showed that alloswitch-1 increased excitatory synaptic responses in prelimbic pyramidal neurons evoked by stimulation of presumed BLA input, and decreased BLA-driven feedforward inhibition of amygdala output neurons. Both effects were reversed by optical silencing and reinstated by optical reactivation of alloswitch-1. These findings demonstrate for the first time that the action of mGlu5 receptors in the pain neuraxis is not homogenous, and suggest that blockade of mGlu5 receptors in the BLA may limit the overall analgesic activity of mGlu5 receptor antagonists. This could explain the suboptimal effect of mGlu5 NAMs on pain in human studies and validate photopharmacology as an important tool to determine ideal target sites for systemic drugs., Competing Interests: SN, NA, MM, XR, SB, FR, FL, XG, TI, MC, CZ, LL, SM, CG, GB, AL, FN, VN No competing interests declared, (© 2024, Notartomaso, Antenucci et al.)

Voltage-gated ion channels are transmembrane proteins responsible for the generation and propagation of action potentials in excitable cells. Over the last decade, advancements have enabled the elucidation of crystal structures of ion channels. This progress in structural understanding, particularly in identifying the binding sites of local anesthetics, opens avenues for the design of novel compounds capable of modulating ion conduction. However, many traditional drugs lack selectivity and come with adverse side effects. The emergence of photopharmacology has provided an orthogonal way of controlling the activity of compounds, enabling the regulation of ion conduction with light. In this review, we explore the central pore region of voltage-gated sodium and potassium channels, providing insights from both structural and pharmacological perspectives. We discuss the different binding modes of synthetic compounds that can physically occlude the pore and, therefore, block ion conduction. Moreover, we examine recent advances in the photopharmacology of voltage-gated ion channels, introducing molecular approaches aimed at controlling their activity by using photosensitive drugs., (© 2024 The Authors. ChemPhysChem published by Wiley-VCH GmbH.)

The therapeutic effects of many pharmacotherapies have been explored, but disadvantages such as low drug specificity, drug resistance and side effects makes their effective delivery to target sites a great challenge. Consequently, a distinctive prodrug-based technology have emerged as an effective treatments because of their distinctive advantages, such as high drug loading capacity, precise targeting, reduced side effects and spatial and temporal controllability. In particular, the use of gamma/X-ray-mediated strategies in radiotherapy is a new strategy that could enable the precise drug release from implanted devices. This review presents readers with the current state of prodrug therapy and reports the design protocols of rational and effective prodrugs for clinical use., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Photopharmacology is a young and rapidly developing field of research that offers significant potential for new insights into targeted therapy. While it primarily focuses on cancer treatment, it also holds promise for other diseases. The key feature of photopharmacological agents is the presence of a photosensitive and biologically active component in the same molecule. In our current study, we synthesized a spiropyran-based meta-stable state photoacid containing a fragment of β-estradiol. This compound exhibits negative photochromism and photocontrolled fluorescence under visible-light irradiation due to the initial stabilization of its self-protonated form in solution. We conducted comprehensive biological studies on the HeLa cells model to assess the short- and long-term cytotoxicity of the photoacid, its metabolic effects, its influence on signaling and epithelial-mesenchymal transition super-system pathways, and the proportion of the population enriched with cancer stem cells. Our findings reveal that this derivative demonstrates low cytotoxicity to HeLa cells, yet it is capable of dramatically reducing malignant cells side population enriched in cancer stem cells. Additionally, appropriate structural modification lead to an increase in some other biological effects compared to β-estradiol. In particular, our substance possesses rare properties of AP-1 suppression and demonstrates some pro-oxidant and metabolic effects, which can be regulated by visible light irradiation. As a result, the new estradiol-based photoacid may be considered a promising multi-acting photopharmacological agent for the next-generation anti-cancer research & development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Photopharmacology, incorporating photoswitches such as azobenezes into drugs, is an emerging therapeutic method to realize spatiotemporal control of pharmacological activity by light. However, most photoswitchable molecules are triggered by UV light with limited tissue penetration, which greatly restricts the in vivo application. Here, this study proves that 131 I can trigger the trans-cis photoisomerization of a reported azobenezen incorporating PROTACs (azoPROTAC). With the presence of 50 µCi mL -1 131 I, the azoPROTAC can effectively down-regulate BRD4 and c-Myc levels in 4T1 cells at a similar level as it does under light irradiation (405 nm, 60 mW cm -2 ). What's more, the degradation of BRD4 can further benefit the 131 I-based radiotherapy. The in vivo experiment proves that intratumoral co-adminstration of 131 I (300 µCi) and azoPROTC (25 mg kg -1 ) via hydrogel not only successfully induce protein degradation in 4T1 tumor bearing-mice but also efficiently inhibit tumor growth with enhanced radiotherapeutic effect and anti-tumor immunological effect. This is the first time that a radioisotope is successfully used as a trigger in photopharmacology in a mouse model. It believes that this study will benefit photopharmacology in deep tissue., (© 2024 Wiley‐VCH GmbH.)

Orexinergic neurons are critically involved in regulating arousal, wakefulness, and appetite. Their dysfunction has been associated with sleeping disorders, and non-peptide drugs are currently being developed to treat insomnia and narcolepsy. Yet, no light-regulated agents are available to reversibly control their activity. To meet this need, a photoswitchable peptide analogue of the endogenous neuroexcitatory peptide orexin-B was designed, synthesized, and tested in vitro and in vivo. This compound - photorexin - is the first photo-reversible ligand reported for orexin receptors. It allows dynamic control of activity in vitro (including almost the same efficacy as orexin-B, high nanomolar potency, and subtype selectivity to human OX 2 receptors) and in vivo in zebrafish larvae by direct application in water. Photorexin induces dose- and light-dependent changes in locomotion and a reduction in the successive induction reflex that is associated with sleep behavior. Molecular dynamics calculations indicate that trans and cis photorexin adopt similar bent conformations and that the only discriminant between their structures and activities is the positioning of the N-terminus. This, in the case of the more active trans isomer, points towards the OX 2 N-terminus and extra-cellular loop 2, a region of the receptor known to be involved in ligand binding and recognition consistent with a "message-address" system. Thus, our approach could be extended to several important families of endogenous peptides, such as endothelins, nociceptin, and dynorphins among others, that bind to their cognate receptors through a similar mechanism: a "message" domain involved in receptor activation and signal transduction, and an "address" sequence for receptor occupation and improved binding affinity., (© 2024. The Author(s).)

The emerging field of photopharmacology is a promising chemobiological methodology for optical control of drug activities that could ultimately solve the off-target toxicity outside the disease location of many drugs for the treatment of a given pathology. The use of photolytic reactions looks very attractive for a light-activated drug release but requires to develop photolytic reactions sensitive to red or near-infrared light excitation for better tissue penetration. This review will present the concepts of triplet-triplet annihilation upconversion-based photolysis and their recent in vivo applications for light-induced drug delivery using photoactivatable nanoparticles., (© 2024 The Authors. Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

Photopharmacology holds a huge untapped potential to locally treat diseases involving photoswitchable drugs via the elimination of drugs' off-target effects. The growth of this field has created a pressing demand to develop such light-active drugs. We explored the potential for creating photoswitchable antibiotic hybrids by attaching pharmacophores norfloxacin/ciprofloxacin and azoisoxazole (photoswitch). All compounds exhibited a moderate to a high degree of bidirectional photoisomerization, long thermal cis half-lives, and impressive photoresistance. Gram-negative pathogens were found to be insensitive to these hybrids, while against Gram-positive pathogens, all hybrids in their trans states exhibited antibacterial activity that is comparable to that of the parent drugs. Notably, the toxicity of the irradiated hybrid 6 was found to be 2-fold lower than the nonirradiated trans isomer, indicating that the pre-inactivated cis-enriched drug can be employed for the site-specific treatment of bacterial infection using light, which could potentially eliminate the unwanted exposure of toxic antibiotics to both beneficial and untargeted harmful microbes in our body. Molecular docking revealed different binding affinity of the cis and trans isomers with the topoisomerase IV enzyme, due to their different shapes., (© 2024 Wiley‐VCH GmbH.)

Photopharmacology is an emerging field that utilizes photo-responsive molecules to enable control over the activity of a drug using light. The aim is to limit the therapeutic action of a drug at the level of diseased tissues and organs. Considering the well-known implications of protein kinases in cancer and the therapeutic issues associated with protein kinase inhibitors, the photopharmacology is seen as an innovative and alternative solution with great potential in oncology. In this context, we developed the first photocaged TAM kinase inhibitors based on UNC2025, a first-in-class small molecule kinase inhibitor. These prodrugs showed good stability in biologically relevant buffer and rapid photorelease of the photoremovable protecting group upon UV-light irradiation (<10 min.). These light-activatable prodrugs led to a 16-fold decrease to a complete loss of kinase inhibition, depending on the protein and the position at which the coumarin-type phototrigger was introduced. The most promising candidate was the N,O-dicaged compound, showing the superiority of having two photolabile protecting groups on UNC2025 for being entirely inactive on TAM kinases. Under UV-light irradiation, the N,O-dicaged compound recovered its inhibitory potency in enzymatic assays and displayed excellent antiproliferative activity in RT112 cell lines., (© 2024 Wiley‐VCH GmbH.)

Photoisomerizable peptides are promising drug candidates in photopharmacology. While azobenzene- and diarylethene-containing photoisomerizable peptides have already demonstrated their potential in this regard, reports on the use of spiropyrans to photoregulate bioactive peptides are still scarce. This work focuses on the design and synthesis of a spiropyran-derived amino acid, (S)-2-amino-3-(6'-methoxy-1',3',3'-trimethylspiro-[2H-1-benzopyran-2,2'-indolin-6-yl])propanoic acid, which is suitable for the preparation of photoisomerizable peptides. The utility of this amino acid is demonstrated by incorporating it into the backbone of BP100, a known membrane-active peptide, and by examining the photoregulation of the membrane perturbation by the spiropyran-containing peptides. The toxicity of the peptides (against the plant cell line BY-2), their bacteriotoxicity (E. coli), and actin-auxin oscillator modulation ability were shown to be significantly dependent on the photoisomeric state of the spiropyran unit., (© 2024 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

Proteases are involved in many essential biological processes. Dysregulation of their activity underlies a wide variety of human diseases. Photopharmacology, as applied on various classes of proteins, has the potential to assist protease research by enabling spatiotemporal control of protease activity. Moreover, it may be used to decrease side-effects of protease-targeting drugs. In this review, we discuss the current status of the chemical design of photoactivatable proteases inhibitors and their biological application. Additionally, we give insight into future possibilities for further development of this field of research., (© 2024 Wiley‐VCH GmbH.)

Nitric oxide (NO) is a unique biochemical mediator involved in the regulation of vital processes. Light-controllable NO releasers show promise in the development of smart therapies. Here, we present a novel biocompatible material based on polydimethylsiloxane (PDMS) doped with BODIPY derivatives containing an N-nitroso moiety that is capable of the photoinduced generation of NO. We study the green-light-induced NO-release properties with the following three methods: electrochemical gas-phase sensor, liquid-phase sensor, and the Griess assay. Prolonged release of NO from the polymer films after short irradiation by narrow-band LED light sources and a laser beam is demonstrated. Importantly, this was accompanied by no or little release of the parent compound (BODIPY-based photodonor). Silicone films with the capability of controllable and clean NO release can potentially be used as a highly portable NO delivery system for different therapeutic applications.

Nicotinamide adenine dinucleotide (NAD + ) is an essential coenzyme with diverse biological functions in DNA synthesis. Nicotinamide phosphoribosyltransferase (NAMPT) is a key rate-limiting enzyme involved in NAD + biosynthesis in mammals. We developed the first chemical tool for optical control of NAMPT and NAD + in biological systems using photoswitchable proteolysis-targeting chimeras (PS-PROTACs). An NAMPT activator and dimethylpyrazolazobenzene photoswitch were used to design highly efficient PS-PROTACs, enabling up- and down-reversible regulation of NAMPT and NAD + in a light-dependent manner and reducing the toxicity associated with inhibitor-based PS-PROTACs. PS-PROTAC was activated under 620 nm irradiation, realizing in vivo optical manipulation of antitumor activity, NAMPT, and NAD + ., (© 2024 Wiley-VCH GmbH.)

In addition to the discovery of new (modified) potent antibiotics to combat antibiotic resistance, there is a critical need to develop novel strategies that would restrict their off-target effects and unnecessary exposure to bacteria in our body and environment. We report a set of new photoswitchable arylazopyrazole-modified norfloxacin antibiotics that present a high degree of bidirectional photoisomerization, impressive fatigue resistance and reasonably high cis half-lives. The irradiated isomers of most compounds were found to exhibit nearly equal or higher antibacterial activity than norfloxacin against Gram-positive bacteria. Notably, against norfloxacin-resistant S. aureus bacteria, the visible-light-responsive p-SMe-substituted derivative showed remarkably high antimicrobial potency (MIC of 0.25 μg/mL) in the irradiated state, while the potency was reduced by 24-fold in case of its non-irradiated state. The activity was estimated to be retained for more than 7 hours. This is the first report to demonstrate direct photochemical control of the growth of antibiotic-resistant bacteria and to show the highest activity difference between irradiated and non-irradiated states of a photoswitchable antibiotic. Additionally, both isomers were found to be non-harmful to human cells. Molecular modellings were performed to identify the underlying reason behind the high-affinity binding of the irradiated isomer to topoisomerase IV enzyme., (© 2024 Wiley-VCH GmbH.)

In patients with prostate carcinoma as well as in some other cancer types, the reduction of testosterone levels is desired because the hormone stimulates cancer cell growth. One molecular target for this goal is the inhibition of 17β-hydroxysteroid dehydrogenase type 3 (17βHSD3), which produces testosterone from its direct precursor androstenedione. Recent research in this field is trying to harness photopharmacological properties of certain compounds so that the inhibitory effect could be turned on and off by irradiation. Seven new light-switchable diazocines were investigated with regard to their inhibition of 17βHSD3. For this purpose, transfected HEK-293 cells and isolated microsomes were treated with the substrate and the potential inhibitors with and without irradiation for an incubation period of 3 or 5 h. The amount of generated testosterone was measured by UHPLC and compared between samples and control as well as between irradiated and non-irradiated samples. There was no significant difference between samples with and without irradiation. However, four of the seven diazocines led to a significantly lower testosterone production both in cell and in microsome assays. In some of the irradiated samples, a partial destruction of the diazocines was observed, indicated by an additional UHPLC peak. However, the influence on the inhibition is negligible, because the majority of the substance remained intact. In conclusion, new inhibitors of 17βHSD3 have been found, but so far without the feature of a light switch, since the configurational alteration of the diazocines by irradiation did not lead to a change in bioactivity. Further modification might help to find a light-switching molecule that inhibits only in one configuration., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

β 2 -adrenergic receptor (β 2 -AR) agonists are used for the treatment of asthma and chronic obstructive pulmonary disease, but also play a role in other complex disorders including cancer, diabetes and heart diseases. As the cellular and molecular mechanisms in various cells and tissues of the β 2 -AR remain vastly elusive, we developed tools for this investigation with high temporal and spatial resolution. Several photoswitchable β 2 -AR agonists with nanomolar activity were synthesized. The most potent agonist for β 2 -AR with reasonable switching is a one-digit nanomolar active, trans-on arylazopyrazole-based adrenaline derivative and comprises valuable photopharmacological properties for further biological studies with high structural accordance to the native ligand adrenaline., (© 2024 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)

Photolipids have emerged as attractive tools for the optical control of lipid functions. They often contain an azobenzene photoswitch that imparts a cis double-bond upon irradiation. Herein, we present the application of photoswitching to a lipidated natural product, the potent proteasome inhibitor cepafungin I. Several azobenzene-containing lipids were attached to the cyclopeptide core, yielding photoswitchable derivatives. Most notably, PhotoCep4 exhibited a 10-fold higher cellular potency in its light-induced cis-form, matching the potency of natural cepafungin I. The length of the photolipid tail and distal positioning of the azobenzene photoswitch with respect to the macrocycle is critical for this activity. In a proteome-wide experiment, light-triggered PhotoCep4 modulation showed high overlap with constitutively active cepafungin I. The mode of action was studied using crystallography and revealed an identical binding of the cyclopeptide in comparison to cepafungin I, suggesting that differences in their cellular activity originate from switching the tail structure. The photopharmacological approach described herein could be applicable to many other natural products as lipid conjugation is common and often necessary for potent activity. Such lipids are often introduced late in synthetic routes, enabling facile chemical modifications., (© 2023 Wiley-VCH GmbH.)

In recent years, there has been growing interest in the potential therapeutic use of inhibitors of adenosine A 2A receptors (A 2A R) for the treatment of neurodegenerative diseases and cancer. Nevertheless, the widespread expression of A 2A R throughout the body emphasizes the importance of temporally and spatially selective ligands. Photopharmacology is an emerging strategy that utilizes photosensitive ligands to attain high spatiotemporal precision and regulate the function of biomolecules using light. In this study, we combined photochemistry and cellular and in vivo photopharmacology to investigate the light sensitivity of the FDA-approved antagonist istradefylline and its potential use as an A 2A R photopharmacological tool. Our findings reveal that istradefylline exhibits rapid trans-to-cis isomerization under near-UV light, and prolonged exposure results in the formation of photocycloaddition products. We demonstrate that exposure to UV light triggers a time-dependent decrease in the antagonistic activity of istradefylline in A 2A R-expressing cells and enables real-time optical control of A 2A R signaling in living cells and zebrafish. Together, these data demonstrate that istradefylline is a photoinactivatable A 2A R antagonist and that this property can be utilized to perform photopharmacological experiments in living cells and animals.

Photoactivated chemotherapy (PACT) is a promising cancer treatment modality that kills cancer cells via photochemical uncaging of a cytotoxic drug. Most ruthenium-based photocages used for PACT are activated with blue or green light, which penetrates sub-optimally into tumor tissues. Here, we report amide functionalization as a tool to fine-tune the toxicity and excited states of a terpyridine-based ruthenium photocage. Due to conjugation of the amide group with the terpyridine π system in the excited state, the absorption of red light (630 nm) increased 8-fold, and the photosubstitution rate rose 5-fold. In vitro, red light activation triggered inhibition of tubulin polymerization, which led to apoptotic cell death both in normoxic (21 % O 2 ) and hypoxic (1 % O 2 ) cancer cells. In vivo, red light irradiation of tumor-bearing mice demonstrated significant tumor volume reduction (45 %) with improved biosafety, thereby demonstrating the clinical potential of this compound., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Traditional methods for site-specific drug delivery in the brain are slow, invasive, and difficult to interface with recordings of neural activity. Here, we demonstrate the feasibility and experimental advantages of in vivo photopharmacology using "caged" opioid drugs that are activated in the brain with light after systemic administration in an inactive form. To enable bidirectional manipulations of endogenous opioid receptors in vivo, we developed photoactivatable oxymorphone (PhOX) and photoactivatable naloxone (PhNX), photoactivatable variants of the mu opioid receptor agonist oxymorphone and the antagonist naloxone. Photoactivation of PhOX in multiple brain areas produced local changes in receptor occupancy, brain metabolic activity, neuronal calcium activity, neurochemical signaling, and multiple pain- and reward-related behaviors. Combining PhOX photoactivation with optical recording of extracellular dopamine revealed adaptations in the opioid sensitivity of mesolimbic dopamine circuitry in response to chronic morphine administration. This work establishes a general experimental framework for using in vivo photopharmacology to study the neural basis of drug action., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

To interrogate neural circuits and crack their codes, in vivo brain activity imaging must be combined with spatiotemporally precise stimulation in three dimensions using genetic or pharmacological specificity. This challenge requires deep penetration and focusing as provided by infrared light and multiphoton excitation, and has promoted two-photon photopharmacology and optogenetics. However, three-photon brain stimulation in vivo remains to be demonstrated. We report the regulation of neuronal activity in zebrafish larvae by three-photon excitation of a photoswitchable muscarinic agonist at 50 pM, a billion-fold lower concentration than used for uncaging, and with mid-infrared light of 1560 nm, the longest reported photoswitch wavelength. Robust, physiologically relevant photoresponses allow modulating brain activity in wild-type animals with spatiotemporal and pharmacological precision. Computational calculations predict that azobenzene-based ligands have high three-photon absorption cross-section and can be used directly with pulsed infrared light. The expansion of three-photon pharmacology will deeply impact basic neurobiology and neuromodulation phototherapies., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Purpose: Severe side effects prevent the utilization of otherwise promising drugs in treatments. These side effects arise when drugs affect untargeted tissues due to poor target specificity. In photopharmacology, light controls the timing and the location of drug delivery, improving treatment specificity and pharmacokinetic control. Photopharmaceuticals have not seen widespread adoption in part because researchers do not always have access to reliable and reproducible light delivery devices at prices which fit within the larger research budget. Method: In this work, we present a customizable photomodulator for use in both wearable and implantable devices. For experimental validation of the photomodulator, we photolyse JF-NP-26 in rats. Results: We successfully drive in vivo photopharmacology with a tethered photomodulator and demonstrate modifications which enable the photomodulator to operate wirelessly. Conclusion : By documenting our photomodulator development, we hope to introduce researchers to a simple solution which significantly lowers the engineering barriers to photopharmacology research., Graphical Abstract: Researchers present a photomodulator, a device designed to facilitate in vivo photopharmacology. They demonstrate the in vivo capabilities of the photomodulator by photoreleasing raseglurant, an mGluR5 inhibitor, to treat pain in an acute rat model and follow this study by showing how to reconfigure the photomodulator to work wirelessly and interface with other biomedical devices., Supplementary Information: The online version contains supplementary material available at 10.1007/s13534-023-00334-3., Competing Interests: Conflict of interestH. Ajieren and P. Irazoqui have received royalties as part of a licensing agreement with Eli Lilly and are inventors on a patent application submitted by Eli Lilly for technology related to this research., (© Korean Society of Medical and Biological Engineering 2023. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

The cannabinoid 2 receptor (CB 2 R) has high therapeutic potential for multiple pathogenic processes, such as neuroinflammation. Pathway-selective ligands are needed to overcome the lack of clinical success and to elucidate correlations between pathways and their respective therapeutic effects. Herein, we report the design and synthesis of a photoswitchable scaffold based on the privileged structure of benzimidazole and its application as a functionally selective CB 2 R "efficacy-switch". Benzimidazole azo-arenes offer huge potential for the broad extension of photopharmacology to a wide range of optically addressable biological targets. We used this scaffold to develop compound 10 d, a "trans-on" agonist, which serves as a molecular probe to study the β-arrestin2 (βarr2) pathway at CB 2 R. βΑrr2 bias was observed in CB 2 R internalization and βarr2 recruitment, while no activation occurred when looking at Gα 16 or mini-Gα i . Overall, compound 10 d is the first light-dependent functionally selective agonist to investigate the complex mechanisms of CB 2 R-βarr2 dependent endocytosis., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

We previously developed, synthesized and tested light-activated sulfonylureas for optical control of K ATP channels and pancreatic beta cell activity in vitro and in vivo. Such technology relies on installation of azobenzene photoswitches onto the sulfonylurea backbone, affording light-dependent isomerization, alteration in ligand affinity for SUR1 and hence K ATP channel conductance. Inspired by molecular dynamics simulations and to further improve photoswitching characteristics, we set out to develop a novel push-pull closed ring azobenzene unit, before installing this on the sulfonylurea glimepiride as a small molecule recipient. Three fine-tuned, light-activated sulfonylureas were synthesized, encompassing azetidine, pyrrolidine and piperidine closed rings. Azetidine-, pyrrolidine- and piperidine-based sulfonylureas all increased beta cell Ca 2+ -spiking activity upon continuous blue light illumination, similarly to first generation JB253. Notably, the pyrrolidine-based sulfonylurea showed superior switch OFF performance to JB253. As such, third generation sulfonylureas afford more precise optical control over primary pancreatic beta cells, and showcase the potential of pyrrolidine-azobenzenes as chemical photoswitches across drug classes., (© 2023 The Authors. Diabetic Medicine published by John Wiley & Sons Ltd on behalf of Diabetes UK.)

Converting known ligands into photoswitchable derivatives offers the opportunity to modulate compound structure with light and hence, biological activity. In doing so, these probes provide unique control when evaluating G-protein-coupled receptor (GPCR) mechanism and function. Further conversion of such compounds into covalent probes, known as photoswitchable tethered ligands (PTLs), offers additional advantages. These include localization of the PTLs to the receptor binding pocket. Covalent localization increases local ligand concentration, improves site selectivity and may improve the biological differences between the respective isomers. This work describes chemical, photophysical and biochemical characterizations of a variety of PTLs designed to target the μ-opioid receptor (μOR). These PTLs were modeled on fentanyl, with the lead disulfide-containing agonist found to covalently interact with a cysteine-enriched mutant of this medically-relevant receptor., (© 2023 The Authors. ChemMedChem published by Wiley-VCH GmbH.)

The cannabinoid receptor 2 (CB 2 R) has high, unexploited therapeutic potential in several central nervous system disorders due to its involvement in neuroinflammatory processes and pathologies like neurodegeneration. Dualsteric/bitopic ligands are currently developed to achieve receptor subtype selectivity and biased signaling. To obtain a molecular tool compound with photoswitchable potential dualsteric properties, we applied two different approaches to link a positive allosteric modulator with an orthosteric agonist via a photochromic unit. We characterized the photophysical properties of all compounds and determined efficacy in internalization, calcium mobilization, and BRET studies. We report the first potentially dualsteric photoswitchable ligand for studying molecular mechanisms of CB 2 R-associated pathologies. Compound 17 - para is a submicromolar " cis -on" agonist with >10-fold higher potency compared to its trans photoisomer and allows high spatiotemporal control of CB 2 R activation.

Aromatic units tethered with an azo (-N=N-) functionality comprise a unique class of compounds, known as molecular photoswitches, exhibiting a reversible transformation between their E- and Z-isomers in response to photo-irradiation. Photoswitches have been explored extensively in the recent past to prepare dynamic self-assembled materials, optoelectronic devices, responsive biomaterials, and more. Most of such materials involve azobenzenes as the molecular photoswitch and to date, SciFinder lists more than 7000 articles and 1000 patents. Subsequently, a great deal of effort has been invested to improve the photo-isomerization efficiency and related mesoscopic properties of azobenzenes. Recently, azoheteroarenes and cyclic azobenzenes, such as arylazopyrazoles, arylazoisoxazoles, arylazopyridines, and diazocines, have emerged as second generation molecular photoswitches beyond conventional azobenzenes. These photoswitches offer distinct photoswitching behavior and responsive properties which make them highly promising candidates for multifaceted applications ranging from photoresponsive materials to photopharmacophores. In this minireview, we introduce the structural refinement and photoresponsive properties of azoheteroarenes and diazocines and summarize the state-of-the-art on utilizing these photoswitches as responsive building blocks in supramolecular assembly, material science and photopharmacology, highlighting their versatile photochemical behavior, enhanced functionality, and latest applications., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

The toxicity of local anesthetics is a serious problem, given their widespread use. One of the main causes of the side effects of local anesthetics is their non-selectivity of action in the body. A possible way to increase the selectivity of the action of drugs is to use the photopharmacology approach. Previously, we described the light-controlled local anesthetic ethercaine, the biological effect of which can be controlled using light, thereby increasing its selectivity of action. An important limitation of ethercaine was its low solubility in water, limiting the potential of this compound. In this work, we developed a dosage form of ethercaine, which allowed us to increase its solubility from 0.6% to 2% or more. The resulting 1% solution of ethercaine hydrochloride in 4% Kolliphor ELP had high biological activity on the surface anesthesia model, while demonstrating low acute toxicity in mice with intravenous administration (4-5 times less than that of lidocaine).

In drug development, optical triggering of cancer therapy is increasingly used. Herein, we report a novel photosensitive PI3K inhibitor FD2157, which bears a photoprotecting moiety and can be efficiently cleaved with enhanced anticancer activity upon short-term light irradiation. In biological assessment, FD2157 exhibited remarkably enhanced anticancer activity in inhibition of PI3K pathway against melanoma cell lines upon light irradiation (4 min). Hence, this photosensitive PI3K inhibitor FD2157 may represent a valuable tool compound for studying the PI3K pathway and further optimization toward light-triggered cancer treatment., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Glutamate ionotropic receptors mediate fast excitation processes in the central nervous system of vertebrates and play an important role in synaptic plasticity, learning, and memory. Here, we describe the action of two azobenene-containing compounds, AAQ (acrylamide-azobenzene-quaternary ammonium) and QAQ (quaternary ammonium-azobenzene-quaternary ammonium), which produced rapid and fully reversible light-dependent inhibition of glutamate ionotropic receptors. The compounds demonstrated voltage-dependent inhibition with only minor voltage-independent allosteric action. Calcium-impermeable AMPA receptors had weaker sensitivity compared to NMDA and calcium-permeable AMPA receptors. We further revealed that the compounds bound to NMDA and calcium-permeable AMPA receptors in different modes. They were able to enter the wide selectivity filter of AMPA receptors, and strong negative voltages caused permeation into the cytoplasm. The narrow selectivity filter of the NMDA receptors did not allow the molecules to bypass them; therefore, QAQ and AAQ bound to the shallow channel site and prevented channel closure by a foot-in-the-door mechanism. Computer simulations employing available AMPA and NMDA receptor structures readily reproduced the experimental findings, allowing for the structure-based design of more potent and selective drugs in the future. Thus, our work creates a framework for the development of light-sensitive blockers of calcium-permeable AMPA receptors, which are desirable tools for neuroscience.

Photopharmacology is a booming research area requiring a new generation of agents possessing simultaneous functions of photoswitching and pharmacophore. It is important that any practical implementation of photopharmacology ideally requires spatial control of the medicinal treatment zone. Thus, advances in the study of substances meeting all the listed requirements will lead to breakthrough research in the coming years. In this study, CQDs@phosphonate nanohybrids are presented for the first time and combine biocompatible and nontoxic luminescent carbon quantum dots (CQDs) with photoactive phosphonate enabling inhibition of butyrylcholinesterase (BChE), which is a prognostic marker of numerous diseases. The conjunction of these components in hybrids maintains photoswitching and provides enhancement of BChE inhibition. After laser irradiation with a wavelength of 266 nm, CQDs@phosphonate hybrids demonstrate a drastic increase of butyrylcholinesterase inhibition from 38% up to almost 100% and a simultaneous luminescence decrease. All the listed hybrid properties are demonstrated not only for in vitro experiments but also for complex biological samples, i.e., chicken breast. Thus, the most important achievement is the demonstration of hybrids characterized by a remarkable combination of all-in-one properties important for photopharmacology: (i) bioactivity toward butyrylcholinesterase inhibition, (ii) strong change of inhibition degree as a result of laser irradiation, luminescence as an indicator of (iii) bioactivity state, and of (iv) spatial localization on the surface of a sample.

The presence of signaling-competent G protein-coupled receptors in intracellular compartments is increasingly recognized. Recently, the presence of G i/o protein-coupled melatonin MT 1 receptors in mitochondria has been revealed, in addition to the plasma membrane. Melatonin is highly cell permeant, activating plasma membrane and mitochondrial receptors equally. Here, we present MCS-1145, a melatonin derivative bearing a triphenylphosphonium cation for specific mitochondrial targeting and a photocleavable o-nitrobenzyl group releasing melatonin upon illumination. MCS-1145 displayed low affinity for MT 1 and MT 2 but spontaneously accumulated in mitochondria, where it was resistant to washout. Uncaged MCS-1145 and exogenous melatonin recruited β-arrestin 2 to MT 1 in mitochondria and inhibited oxygen consumption in mitochondria isolated from HEK293 cells only when expressing MT 1 and from mouse cerebellum of WT mice but not from MT 1 -knockout mice. Overall, we developed the first mitochondria-targeted photoactivatable melatonin ligand and demonstrate that melatonin inhibits mitochondrial respiration through mitochondrial MT 1 receptors., Competing Interests: Declaration of interests The authors declare no competing interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)