Photopharmacological modulation of hippocampal local field potential by caged-glutamate with MicroLED probe.

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.

Photo-BQCA: Positive Allosteric Modulators Enabling Optical Control of the M1 Receptor.

The field of G protein-coupled receptor (GPCR) research has greatly benefited from the spatiotemporal resolution provided by light controllable, photoswitchable agents. 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 over 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.

Advancements of prodrug technologies for enhanced drug selectivity in pharmacotherapies.

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.

In vivo photocontrol of orexin receptors with a nanomolar light-regulated analogue of orexin-B.

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 OX2 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 OX2 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.

A chemical platform for the efficient screening of arylazopyrazole-based photoswitchable CENP-E inhibitors using mild cyclization reactions.

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.

Coumarin-Derived Caging Groups in the Spotlight: Tailoring Physiochemical and Photophysical Properties.

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.

A Photocaged Microtubule-Stabilising Epothilone Allows Spatiotemporal Control of Cytoskeletal Dynamics.

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 violet/blue irradiation 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.

Deuteration as a General Strategy to Enhance Azobenzene-Based Photopharmacology.

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.

Meta-stable state photoacid containing ß-estradiol fragment with photomodulated biological activity and anti-cancer stem cells properties.

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 ß-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 ß-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.

Azologs of the Fatty Acid Mimetic Drug Cinalukast Enable Light-Induced PPARα Activation.

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.

Photoswitchable Carbamazepine Analogs for Non-Invasive Neuroinhibition In Vivo.

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.

Voltage-Gated Ion Channels: Structure, Pharmacology and Photopharmacology.

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.

Triplet-Triplet Annihilation Upconversion-Based Photolysis: Applications in Photopharmacology.

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.

Recent clinical trials and optical control as a potential strategy to develop microtubule-targeting drugs in colorectal cancer management.

Colorectal cancer (CRC) has remained the second and the third leading cause of cancer-related death worldwide and in the United States, respectively. Although significant improvement in overall survival has been achieved, death in adult populations under the age of 55 appears to have increased in the past decades. Although new classes of therapeutic strategies such as immunotherapy have emerged, their application is very limited in CRC so far. Microtubule (MT) inhibitors such as taxanes, are not generally successful in CRC. There may be some way to make MT inhibitors work effectively in CRC. One potential advantage that we can take to treat CRC may be the combination of optical techniques coupled to an endoscope or other fiber optics-based devices. A combination of optical devices and photo-activatable drugs may allow us to locally target advanced CRC cells with highly potent MT-targeting drugs. In this Editorial review, we would like to discuss the potential of optogenetic approaches in CRC management.

Nitric Oxide Photorelease from Silicone Films Doped with N-Nitroso BODIPY.

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.

131I Induced In Vivo Proteolysis by Photoswitchable azoPROTAC Reinforces Internal Radiotherapy.

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 131I can trigger the trans-cis photoisomerization of a reported azobenezen incorporating PROTACs (azoPROTAC). With the presence of 50 µCi mL-1 131I, 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 131I-based radiotherapy. The in vivo experiment proves that intratumoral co-adminstration of 131I (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.

Design and validation of a low-cost photomodulator for in vivo photoactivation of a mGluR5 inhibitor.

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.

Light-Activatable Photocaged UNC2025 for Triggering TAM Kinase Inhibition in Bladder Cancer.

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.

Spiropyran-Based Photoisomerizable α-Amino Acid for Membrane-Active Peptide Modification.

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.

Photoisomerization and Light-Controlled Antibacterial Activity of Fluoroquinolone-Azoisoxazole Hybrids.

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.