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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‐GluAblock 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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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 M1 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 M1 receptor signaling and a proof of concept for photoswitchable allosteric modulators at Class A receptors. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Isotonitazene belongs to a potent class of μ‐opioid receptor (μOR) ligands, known as nitazenes. The lack of knowledge surrounding this agonist and others in its class has sparked thorough re‐investigations. To aid in these investigations, the purportedly covalent yet underexplored nitazene BIT was biochemically re‐evaluated in this work, along with a newly synthesized analogue, Iso‐BIT. Moreover, in the pursuit of understanding the mechanism, function and interactions of the μOR, this study involved developing photoswitchable nitazene derivatives as potential probe molecules. Converting known ligands into azo‐containing photoswitchable derivatives offers the opportunity to modulate ligand structure with light, allowing for photocontrol of compound activity. While photocontrol of μOR activity could not be entirely achieved, photophysical evaluation of these 2‐benzimidazole azo‐arenes revealed a novel photoswitch scaffold that responds to visible light. Furthermore, azo‐containing 2 e and 3 e emerged as promising nitazene derivatives that were able to form an exceptionally high fraction of covalent‐ligand receptor complexes with wild‐type μOR at physiological pH. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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+. [ABSTRACT FROM AUTHOR]

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+. [ABSTRACT FROM AUTHOR]

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+. [ABSTRACT FROM AUTHOR]

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+. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Azobenzenes are a series of compounds that can be isomerized upon irradiation with light. These molecules can modify the physical, chemical, and biological properties of a diverse range of materials. They can control protein structure and function with temporal and spatial precision. In this work, we investigated the possible interaction between azobenzene and aromatic amino acids. We hypothesized that aromatic amino acids, such as tryptophan, would show altered photochemical properties when conjugated with azobenzene. When irradiated at either 365 nm or 465 nm, the molecule now lacks the usually characteristic photoswitch capabilities and is visibly fluorescent at 365 nm. To our knowledge, this is the first evidence to suggest that primary protein structure could affect photoswitch activity. The knowledge gained from this research will help to further the understanding of azobenzenes as they are used in biomolecules. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

β2‐adrenergic receptor (β2‐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 β2‐AR remain vastly elusive, we developed tools for this investigation with high temporal and spatial resolution. Several photoswitchable β2‐AR agonists with nanomolar activity were synthesized. The most potent agonist for β2‐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. [ABSTRACT FROM AUTHOR]

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 % O2) and hypoxic (1 % O2) 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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The cannabinoid 2 receptor (CB2R) 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 CB2R "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 CB2R. βΑrr2 bias was observed in CB2R internalization and βarr2 recruitment, while no activation occurred when looking at Gα16 or mini‐Gαi. Overall, compound 10 d is the first light‐dependent functionally selective agonist to investigate the complex mechanisms of CB2R‐βarr2 dependent endocytosis. [ABSTRACT FROM AUTHOR]

We previously developed, synthesized and tested light‐activated sulfonylureas for optical control of KATP 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 KATP 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 Ca2+‐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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Photopharmacology is an attractive approach for achieving targeted drug action with the use of light. In photopharmacology, molecular photoswitches are introduced into the structure of biologically active small molecules to allow for the optical control of their potency. Going beyond trial and error, photopharmacology has progressively applied rational drug design methodologies to devise light‐controlled bioactive ligands. In this review, we categorize photopharmacological efforts from the standpoint of medicinal chemistry strategies, focusing on diffusible photochromic ligands modified with photoswitches that operate through E‐Z bond isomerization. In the vast majority of cases, photoswitchable ligands are designed as analogs of existing compounds, through a variety of approaches. By analyzing in detail a comprehensive list of instructive examples, we describe the state of the art and discuss future opportunities for rational design in photopharmacology. [ABSTRACT FROM AUTHOR]

Background and Purpose: Opioid‐based drugs are the gold standard medicines for pain relief. However, tolerance and several side effects (i.e. constipation and dependence) may occur upon chronic opioid administration. Photopharmacology is a promising approach to improve the benefit/risk profiles of these drugs. Thus, opioids can be locally activated with high spatiotemporal resolution, potentially minimizing systemic‐mediated adverse effects. Here, we aimed at developing a morphine photo‐derivative (photocaged morphine), which can be activated upon light irradiation both in vitro and in vivo. Experimental Approach Light‐dependent activity of pc‐morphine was assessed in cell‐based assays (intracellular calcium accumulation and electrophysiology) and in mice (formalin animal model of pain). In addition, tolerance, constipation and dependence were investigated in vivo using experimental paradigms. Key results: In mice, pc‐morphine was able to elicit antinociceptive effects, both using external light‐irradiation (hind paw) and spinal cord implanted fibre‐optics. In addition, remote morphine photoactivation was devoid of common systemic opioid‐related undesired effects, namely, constipation, tolerance to the analgesic effects, rewarding effects and naloxone‐induced withdrawal. Conclusion and Implications: Light‐dependent opioid‐based drugs may allow effective analgesia without the occurrence of tolerance or the associated and severe opioid‐related undesired effects. LINKED ARTICLES: This article is part of a themed issue on Advances in Opioid Pharmacology at the Time of the Opioid Epidemic. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v180.7/issuetoc [ABSTRACT FROM AUTHOR]

BACKGROUND: Ionotropic γ‐aminobutyric acid (GABA) receptor (GABAR) in an insect is the major inhibitory receptor and is one of the most important targets for insecticides. Due to the high spatiotemporal resolution of GABAR, the photopharmacological ligands acting on it in vertebrates but not insect have been developed. RESULTS: In this study, two types of photochromic ligands (PCLs) including DTFIPs (DTFIP1 and DTFIP2) and ABFIPs (p‐, m‐, and o‐ABFIP) were synthesized by incorporating photoswitch azobenzene or dithienylethene into fipronil (FIP), which is the antagonist of insect GABAR. Their photomodulation was measured by mosquito larval behavior, and their potential action mechanism was explored by the two‐electrode voltage clamp (TEVC) technique in vitro. DTFIP1 and m‐ABFIP exhibited the most significant difference of insecticidal activity by about 90‐ and 5‐fold to mosquito larvae between non‐irradiated and irradiated formation, respectively, and allowed for optical control of mosquito swimming activity. TEVC assay results indicated that m‐ABFIP and DTFIP1 enable optical control over the homomeric LsRDL‐type GABAR, which is achieved by regulating the chloride channel of resistance to dieldrin (RDL)‐type GABAR by photoisomerization. CONCLUSION: Our results suggested that PCLs synthesized from fipronil provide an alternative and precise tool for studying insect ionotropic GABARs and GABA‐dependent behavior. © 2022 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Light‐responsive molecules have seen a major advance in modulating biological functions in recent years. Especially photoswitches are highly attractive building blocks due to the reversible nature of their light‐mediated reactivity. They are frequently used to affect both the properties of small bioactive compounds and biomacromolecules if incorporated suitably. Despite their success in a plethora of applications, only a limited set of photochromic core structures is routinely employed and a large number of photochromic couples are under‐investigated in biological context. Broadening the toolbox of photoswitches available to modulate biological activity would open new avenues and unlock the full potential of photoswitchable molecules for biological studies. In this work, we explore the photochemical and thermal properties of the dihydroazulene/vinylheptafulvene photochromic couple as peptide conjugates in aqueous environment. [ABSTRACT FROM AUTHOR]

Photoremovable protecting groups (PPGs) releasable by two‐photon excitation (2PE) provide spatio‐temporal control over the photoactivation of biological effectors to study biological systems. We synthesized four derivatives of the (8‐cyano‐7‐hydroxyquinolin‐2‐yl)methyl (CyHQ) chromophore by functionalization of position C4 to generate four different derivatives (MeO‐CyHQ, Mor‐CyHQ, pTol‐CyHQ, and TMP‐CyHQ) for studying the release of tertiary amines via 2PE. Sulpiride, an anti‐dopaminergic drug, was selected as a model substrate. All probes had excellent properties for use in biological settings, including high quantum yield (Φu), hydrolytic stability, and good aqueous solubility in simulated physiological buffer. The TMP‐CyHQ probe enhanced the two‐photon uncaging action cross‐section (δu) 8‐fold (2.64 GM) compared to the parent CyHQ‐sulpiride. The optimized PPG was used to mediate the photoactivation of various biological effectors containing the tertiary amine functionality (tamoxifen, 4‐hydroxytamoxifen, yohimbine) using 2PE. All constructs showed excellent efficiency with δu ranging from 1.21 to 2.42 GM and moderate to excellent yields of tertiary amines released. [ABSTRACT FROM AUTHOR]

The natural product jasplakinolide is widely used to stabilize F‐actin. Based on extensive structure–activity relationship studies, we have developed a new generation of photoswitchable jasplakinolides that feature rationally designed red‐shifted azobenzene photoswitches. Our lead compound, nOJ, can be activated with longer wavelengths in the visible range (e.g. 440–475 nm) and rapidly returns to its inactive state through thermal relaxation. nOJ enables the reversible control of F‐actin dynamics, as shown through live‐cell imaging, cell migration, and cell proliferation assays. Short, local irradiation with blue light resulted in highly localized and reversible actin aggregation with subcellular precision. Our optical tool can be useful in diverse fields to study actin dynamics with excellent spatiotemporal resolution. [ABSTRACT FROM AUTHOR]

Photoswitchable ligands as biological tools provide an opportunity to explore the kinetics and dynamics of the clinically relevant μ‐opioid receptor. These ligands can potentially activate or deactivate the receptor when desired by using light. Spatial and temporal control of biological activity allows for application in a diverse range of biological investigations. Photoswitchable ligands have been developed in this work, modelled on the known agonist fentanyl, with the aim of expanding the current "toolbox" of fentanyl photoswitchable ligands. In doing so, ligands have been developed that change geometry (isomerize) upon exposure to light, with varying photophysical and biochemical properties. This variation in properties could be valuable in further studying the functional significance of the μ‐opioid receptor. [ABSTRACT FROM AUTHOR]

The rise of antibiotic resistance causes a serious health care problem, and its counterfeit demands novel, innovative concepts. The combination of photopharmacology, enabling a light‐controlled reversible modulation of drug activity, with antibiotic drug design has led to first photoswitchable antibiotic compounds derived from established scaffolds. In this study, we converted cystobactamids, gyrase‐inhibiting natural products with an oligoaryl scaffold and highly potent antibacterial activities, into photoswitchable agents by inserting azobenzene in the N‐terminal part and/or an acylhydrazone moiety near the C‐terminus, yielding twenty analogs that contain mono‐ as well as double‐switches. Antibiotic and gyrase inhibition properties could be modulated 3.4‐fold and 5‐fold by light, respectively. Notably, the sensitivity of photoswitchable cystobactamids towards two known resistance factors, the peptidase AlbD and the scavenger protein AlbA, was light‐dependent. While irradiation of an analog with an N‐terminal azobenzene with 365 nm light led to less degradation by AlbD, the AlbA‐mediated inactivation was induced. This provides a proof‐of‐principle that resistance towards photoswitchable antibiotics can be optically controlled. [ABSTRACT FROM AUTHOR]

Photoswitchable reagents can be powerful tools for high‐precision biological control. TRPC5 is a Ca2+‐permeable cation channel with distinct tissue‐specific roles, from synaptic function to hormone regulation. Reagents giving spatiotemporally‐resolved control over TRPC5 activity may be key to understanding and harnessing its biology. Here we develop the first photoswitchable TRPC5‐modulator, BTDAzo, to address this goal. BTDAzo can photocontrol TRPC5 currents in cell culture, as well as controlling endogenous TRPC5‐based neuronal Ca2+ responses in mouse brain slices. BTDAzos are also the first reported azo‐benzothiadiazines, an accessible and conveniently derivatised azoheteroarene with strong two‐colour photoswitching. BTDAzo′s ability to control TRPC5 across relevant channel biology settings makes it suitable for a range of dynamically reversible photoswitching studies in TRP channel biology, with the aim to decipher the various biological roles of this centrally important ion channel. [ABSTRACT FROM AUTHOR]

Photocleavable prodrugs enable controllable drug delivery to target sites modulated by light irradiation. However, the in vivo utility is usually hindered by their insolubility and inefficient delivery. In this study, we report a simple strategy of co‐assembling boron‐dipyrromethene‐chlorambucil prodrug and near‐infrared dye IR783 to fabricate photoresponsive nanoassemblies, which achieved both high prodrug loading capacity (~99%) and efficient light‐triggered prodrug activation. The incorporated IR783 dye not only stabilized the nanoparticles and contributed tumor targeting as usual, but also exhibited degradation after light irradiation and in‐situ monitoring of nanoparticle dissociation by fluorescent imaging. Systemic administration of the nanoparticles and localized light irradiation at tumor sites enabled monitorable and efficient drug release in vivo. Our results demonstrate that such prodrug‐dye co‐assembled nanomedicine is a promising formulation for photoresponsive drug delivery, which would advance the translation of photoresponsive nanomedicines. [ABSTRACT FROM AUTHOR]

A Ru(bpy)3Cl2 photocatalyst is applied to the rapid trans to cis isomerization of a range of alkene‐containing pharmacological agents, including combretastatin A‐4 (CA‐4), a clinical candidate in oncology, and resveratrol derivatives, switching their configuration from inactive substances to potent cytotoxic agents. Selective in cellulo activation of the CA‐4 analog Res‐3M is demonstrated, along with its potent cytotoxicity and inhibition of microtubule dynamics. [ABSTRACT FROM AUTHOR]

Photolabile Protecting Groups (PPGs) are molecular tools used, for example, in photopharmacology for the activation of drugs with light, enabling spatiotemporal control over their potency. Yet, red‐shifting of PPG activation wavelengths into the NIR range, which penetrates the deepest in tissue, has often yielded inefficient or insoluble molecules, hindering the use of PPGs in the clinic. To solve this problem, we report herein a novel concept in PPG design, by transforming clinically‐applied NIR‐dyes with suitable molecular orbital configurations into new NIR‐PPGs using computational approaches. Using this method, we demonstrate how Cy7, a class of NIR dyes possessing ideal properties (NIR‐absorption, high molecular absorptivity, excellent aqueous solubility) can be successfully converted into Cy7‐PPG. We report a facile synthesis towards Cy7‐PPG from accessible precursors and confirm its excellent properties as the most redshifted oxygen‐independent NIR‐PPG to date (λmax=746 nm). [ABSTRACT FROM AUTHOR]

Classical psychedelics are a group of hallucinogens which trigger non‐ordinary states of consciousness through activation of the 5‐HT2A receptor (5‐HT2AR) in the brain. However, the exact mechanism of how 5‐HT2AR agonism alters perception remains elusive. When studying receptor signaling, tools which work at the same spatiotemporal resolution as the receptor are exceptionally useful. To create such a tool, we designed a set of photoswitchable ligands based on the classical psychedelic N,N‐dimethyltryptamine (DMT). By incorporation of the DMT‐indole ring into the photoswitchable system, we obtained red‐shifted ligands which can be operated by visible light. Among these azo‐DMTs, compound 2 h ("Photo‐DMT") stands out as its cis isomer exhibits DMT like activity while the trans isomer acts as weak partial agonist. Such a cis‐on "efficacy switch" substantially expands the pharmacological toolbox to investigate the complex mechanisms of 5‐HT2AR signaling. [ABSTRACT FROM AUTHOR]

The development of new and effective therapeutics is reliant on the ability to study the underlying mechanisms of potential drug targets in live cells and multicellular systems. A persistent challenge in many drug development programmes is poor selectivity, which can obscure the mechanisms involved and lead to poorly understood modes of action. In efforts to improve our understanding of these complex processes, small molecule inhibitors have been developed in which their OFF/ON therapeutic activity can be toggled using light. Photopharmacology is devoted to using light to modulate drugs. Herein, we highlight the recent progress made towards the development of light‐responsive small molecule inhibitors of selected enzymatic targets. Given the size of this field, literature from 2015 onwards has been reviewed. [ABSTRACT FROM AUTHOR]

Serotonin receptors play central roles in neuromodulation and are critical drug targets for psychiatric disorders. Optical control of serotonin receptor subtypes has the potential to greatly enhance our understanding of the spatiotemporal dynamics of receptor function. While other neuromodulatory receptors have been successfully rendered photoswitchable, reversible photocontrol of serotonin receptors has not been achieved, representing a major gap in GPCR photopharmacology. Herein, we develop the first tools that allow for such control. Azo5HT‐2 shows light‐dependent 5‐HT2AR agonism, with greater activity in the cis‐form. Based on docking and test compound analysis, we also develop photoswitchable orthogonal, remotely‐tethered ligands (PORTLs). These BG‐Azo5HTs provide rapid, reversible, and repeatable optical control following conjugation to SNAP‐tagged 5‐HT2AR. Overall, this study provides a foundation for the broad extension of photopharmacology to the serotonin receptor family. [ABSTRACT FROM AUTHOR]

Optical methods to modulate microtubule dynamics show promise for reaching the micron‐ and millisecond‐scale resolution needed to decrypt the roles of the cytoskeleton in biology. However, optical microtubule stabilisers are under‐developed. We introduce "STEpos" as GFP‐orthogonal, light‐responsive epothilone‐based microtubule stabilisers. They use a novel styrylthiazole photoswitch in a design to modulate hydrogen‐bonding and steric effects that control epothilone potency. STEpos photocontrol microtubule dynamics and cell division with micron‐ and second‐scale spatiotemporal precision. They substantially improve potency, solubility, and ease‐of‐use compared to previous optical microtubule stabilisers, and the structure‐photoswitching‐activity relationship insights in this work will guide future optimisations. The STEpo reagents can contribute greatly to high‐precision research in cytoskeleton biophysics, cargo transport, cell motility, cell division, development, and neuroscience. [ABSTRACT FROM AUTHOR]

Eg5 is a kinesin motor protein that is responsible for bipolar spindle formation and plays a crucial role during mitosis. Loss of Eg5 function leads to the formation of monopolar spindles, followed by mitotic arrest, and subsequent cell death. Several cell‐permeable small molecules have been reported to inhibit Eg5 and some have been evaluated as anticancer agents. We now describe the design, synthesis, and biological evaluation of photoswitchable variants with five different pharmacophores. Our lead compound Azo‐EMD is a cell permeable azobenzene that inhibits Eg5 more potently in its light‐induced cis form. This activity decreased the velocity of Eg5 in single‐molecule assays, promoted formation of monopolar spindles, and led to mitotic arrest in a light dependent way. [ABSTRACT FROM AUTHOR]

BACKGROUND: Photopharmacology is a fast‐growing photonics‐based technology, which realizes the high‐resolution regulation of drugs in time and space through light. The purpose of this research was to introduce photochromic groups into the isoxazoline structure to realize the regulation of γ‐Aminobutyric acid receptors (GABARs) targeting insect behavior. RESULTS: Azobenzene‐Fluralaner analogs ABF02, ABF03 and ABF04 have been proven to have larvicidal activity against mosquito larvae. Cis‐ABF03 had excellent larvicidal activity against mosquito larvae with a median lethal concentration (LC50) value of 1.63, which was better than that of trans‐ABF03 (LC50 = 3.90). In particular, ABF03 also showed insecticidal activity against Mythimna separata. Further experiments showed that ABF03 (1 μm) induced depolarization of dorsal unpaired median neurons after ultraviolet light irradiation, enhanced affinity to the receptor, and blocked ligand‐gated chloride channels of GABARs. ABF03 (1 μm) realized the real‐time photoregulation of the behavior of mosquito larvae, which indicated that the synthesized ligand can complete the binding and off‐target action of drugs and targets in vivo under the regulation of light. CONCLUSION: Azobenzene‐Isoxazoline as photopharmacological ligand was synthesized and evaluated for optical control of insect GABARs and behavior for the first time. ABF03 completed the differential regulation of cockroach neurons and the real‐time reversible regulation of insect behavior. The establishment of photochromic ligands provides a new strategy for basic and convenience‐oriented research on GABARs in invertebrates. © 2021 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

The TAM kinase family arises as a promising therapeutic target for cancer therapy, as well as auto‐immune and viral diseases. In this study, we report the first photoactivatable caged inhibitors of Tyro3 and Mer. This strategy enables spatial and temporal control of the biological activity of the inhibitor upon irradiation with UV light. We describe the design, synthesis, photocleavage properties, and inhibitory activities of four Tyro3 and Mer photoactivatable small molecules. The proof of concept on the TAM kinase family was achieved in vitro, since irradiation by UV light restored the full inhibitory activity of two prodrugs. [ABSTRACT FROM AUTHOR]

We developed a method to control gene activation and base editing in mammalian cells and zebrafish embryos via photochemically activated, caged guide RNAs. Four evenly distributed, caged nucleobases are installed in the 5'‐protospacer region of the gRNA to block interaction between the dCas9:gRNA complex and the dsDNA target. Upon light activation, gRNA:dsDNA hybridization is restored and catalytically dead Cas9‐based CRISPR activation or base editing is achieved. Caged gRNAs are customizable in design, offer spatiotemporal control of transcription activation and base editing in both mammalian cells and zebrafish embryos, and preserve the ability to form dCas9:gRNA complexes for enhanced gRNA stability. We hereby offer a versatile tool for conditional control of interactions between dCas9‐based regulators with their programmed genomic loci, thereby facilitating parsing of spatiotemporally complex genetic events, such as transcription regulation and single‐base mutagenesis. [ABSTRACT FROM AUTHOR]

We report the first cellular application of the emerging near‐quantitative photoswitch pyrrole hemithioindigo, by rationally designing photopharmaceutical PHTub inhibitors of the cytoskeletal protein tubulin. PHTubs allow simultaneous visible‐light imaging and photoswitching in live cells, delivering cell‐precise photomodulation of microtubule dynamics, and photocontrol over cell cycle progression and cell death. This is the first acute use of a hemithioindigo photopharmaceutical for high‐spatiotemporal‐resolution biological control in live cells. It additionally demonstrates the utility of near‐quantitative photoswitches, by enabling a dark‐active design to overcome residual background activity during cellular photopatterning. This work opens up new horizons for high‐precision microtubule research using PHTubs and shows the cellular applicability of pyrrole hemithioindigo as a valuable scaffold for photocontrol of a range of other biological targets. [ABSTRACT FROM AUTHOR]

Light activation is an effective way to impart spatiotemporal control over oligonucleotide probes that are widely applied for gene expression regulation and target function investigation. Among the major oligonucleotide caging strategies, cyclization with a photocleavable linker is an elegant design, which affords both atom efficiency and stability in many biological environments. Here, we introduce an improved protocol for circular oligonucleotide synthesis requiring only one round of HPLC purification. With a series of poly‐U oligonucleotide strands of different sizes and backbone modifications, the pre‐photolysis caging stability and post‐photolysis target binding affinity were studied through a denaturing gel assay and melting temperature measurements. A 14 U 2'‐OMe RNA probe was selected, with strong potential application in transcriptome in vivo analysis (TIVA) for mRNA isolation. [ABSTRACT FROM AUTHOR]

Covalent kinase inhibitors account for some of the most successful drugs that have recently entered the clinic and many others are in preclinical development. A common strategy is to target cysteines in the vicinity of the ATP binding site using an acrylamide electrophile. To increase the tissue selectivity of kinase inhibitors, it could be advantageous to control the reactivity of these electrophiles with light. Here, we introduce covalent inhibitors of the kinase JNK3 that function as photoswitchable affinity labels (PALs). Our lead compounds contain a diazocine photoswitch, are poor non‐covalent inhibitors in the dark, and become effective covalent inhibitors after irradiation with visible light. Our proposed mode of action is supported by X‐ray structures that explain why these compounds are unreactive in the dark and undergo proximity‐based covalent attachment following exposure to light. [ABSTRACT FROM AUTHOR]

The ability to control neural activity is essential for research not only in basic neuroscience, as spatiotemporal control of activity is a fundamental experimental tool, but also in clinical neurology for therapeutic brain interventions. Transcranial‐magnetic, ultrasound, and alternating/direct current (AC/DC) stimulation are some available means of spatiotemporal controlled neuromodulation. There is also light‐mediated control, such as optogenetics, which has revolutionized neuroscience research, yet its clinical translation is hampered by the need for gene manipulation. As a drug‐based light‐mediated control, the effect of a photoswitchable muscarinic agonist (Phthalimide‐Azo‐Iper (PAI)) on a brain network is evaluated in this study. First, the conditions to manipulate M2 muscarinic receptors with light in the experimental setup are determined. Next, physiological synchronous emergent cortical activity consisting of slow oscillations—as in slow wave sleep—is transformed into a higher frequency pattern in the cerebral cortex, both in vitro and in vivo, as a consequence of PAI activation with light. These results open the way to study cholinergic neuromodulation and to control spatiotemporal patterns of activity in different brain states, their transitions, and their links to cognition and behavior. The approach can be applied to different organisms and does not require genetic manipulation, which would make it translational to humans. [ABSTRACT FROM AUTHOR]

Many of the major challenges associated with drug delivery can potentially be addressed by linking drug action to light irradiation. These challenges include the spacing, timing and amount of a drug's activity. Once a drug's activity is linked to light, this activity can be more easily manipulated, because light itself is easy to manipulate. One of the main issues that light control can address is off‐target toxicity. This has the potential to be limited if drugs are activated only in target tissues using light. For drugs that are needed at varying concentrations through the day, varying light has the potential to temporally modulate drug release. Because of these and other advantages, a range of mechanisms for using light to manipulate drug activity has been developed, including photocleavage control, photoconformational control, photothermal control and photodegradation control. These major themes of light control will be described in this minireview, and illustrated with examples. In addition, the issue of tissue light permittivity, arguably the major challenge for the discipline, will be described and analyzed. [ABSTRACT FROM AUTHOR]

Ifenprodil is an important negative allosteric modulator of the N‐methyl‐D‐aspartate (NMDA) receptor. We have synthesized caged and photoswitchable derivatives of this small molecule drug. Caged ifenprodil was biologically inert before photolysis, and UV irradiation efficiently released the drug allowing selective inhibition of GluN2B‐containing NMDA receptors. Azobenzene‐modified ifenprodil, on the other hand, is inert in both its trans and cis configurations, although in silico modeling predicted the trans form to be able to bind to the receptor. The disparity in effectiveness between the two compounds reflects, in part, the inherent ability of each method in manipulating the binding properties of drugs. With appropriate structure‐activity relationship uncaging enables binary control of effector binding, whereas photoswitching using freely diffusible chromophores shifts the dose‐response curve of drug‐receptor interaction. Our data suggest that the efficacy of pharmacophores having a confined binding site such as ifenprodil can be controlled more easily by uncaging in comparison to photoswitching. [ABSTRACT FROM AUTHOR]