Azoheteroarene and Diazocine Molecular Photoswitches: Self-Assembly,Responsive Materials and Photopharmacology

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.     

Fluorinated Azobenzenes Switchable with Red Light

Molecular photoswitches triggered with red or NIR light are optimal for photomodulation of complex biological systems, including efficient penetration of the human body for therapeutic purposes (“therapeutic window”). Yet, they are rarely reported, and even more rarely functional under aqueous conditions. In this work, fluorinated azobenzenes are shown to exhibit efficient E→Z photoisomerization with red light (PSS_660nm >75 % Z) upon conjugation with unsaturated substituents. Initially demonstrated for aldehyde groups, this effect was also observed in a more complex structure by incorporating the chromophore into a cyclic dipeptide with propensity for self-assembly. Under physiological conditions, the latter molecule formed a supramolecular material that reversibly changed its viscosity upon irradiation with red light. Our observation can lead to design of new photopharmacology agents or phototriggered materials for in vivo use.     

Taking Photochromism beyond Visible: Direct One‐Photon NIR Photoswitches Operating in the Biological Window

The success of photopharmacology is inevitably tied to the availability of photoswitches, which can be operated within the biological window (λ=650–1450 nm) to maximize penetration in tissue. A general design strategy has been devised and a dihydropyrene derivative is described here that displays negative T‐type photochromism, allowing for efficient and nearly quantitative (95 %) switching induced by NIR light λ>800 nm. The thermal half‐life of the decolored ring‐open meta‐cyclophanediene isomer ranges from minutes to hours, depending on the solvent polarity and hence serves as a probe of the local environment. Due to the rather subtle geometrical differences between the two isomers, suitably modified NIR photoswitches are potential candidates for switching when bound in the pocket of the biological target, in principle allowing for reversible light‐induced inhibitor deactivation as an alternative approach to externally regulate biological functions.     

Tuning of Bistability,Thermal Stability of the Metastable States,and Application Prospects in the C3-Symmetric Designs of Multiple Azo(hetero)arenes Systems

Light-responsive molecular systems with multiple photoswitches in C₃-symmetric designs have enormous application potential. The design part of such molecular systems is critical due to its influence in several properties associated with the photoswitches. In order to tune, and in the evaluation of the design–property relationship, we synthesized 18 tripodal systems with variations in the core, linkers, connectivity, and azo(hetero)arene photoswitches. Through extensive spectroscopic and computational studies, we envisaged the factors controlling near-quantitative photoisomerization in both the directions (bistability) and the thermal stability of the metastable states. Furthermore, we also evaluated the impact of designs in obtaining reversible photo-responsive sol-gel phase transitions, solvatochromism, photo- and thermochromism.     

The Quest for Photoswitches Activated by Near‐Infrared Light: A Theoretical Study of the Photochemistry of BF2‐Coordinated Azo Derivatives

Recently synthesized BF₂‐coordinated azo derivatives have been proposed as photoswitches that operate in the optical window (λ=600–1200 nm) for use in bioimaging applications. Herein, we have theoretically analyzed these compounds and modified some substituents to analyze which properties of the molecule govern its photochemistry. Our results compare rather well with the available experimental data, so our methodology, based on density functional theory (DFT) calculations for the ground electronic state and time‐dependent‐DFT for the first excited electronic state, is validated. Through systematic modification of different substituents of the parent system, we designed compounds that are predicted to operate fully within the optical window. We also analyzed several molecules for which the cis isomer is the more stable isomer, a quite unusual result for azobenzene derivatives that is a much coveted property for some applications of these photoactive molecules in pharmacology. Our results also provide insight into other properties relevant for photoswitches, such as the thermal stability of the less stable isomer and the magnitude of the gap between the wavelengths of the radiation that activates each isomerization process, which must be as large as possible to improve the yield of each photoisomerization. From a more general perspective, our results may provide a step towards the rational design of new photoswitches that fulfill a set of desired characteristics.     

Stalis: A Computational Method for Template-Based Ab Initio Ligand Design

Proteins interact with small molecules through specific molecular recognition, which is central to essential biological functions in living systems. Therefore, understanding such interactions is crucial for basic sciences and drug discovery. Here, we present S tructure t emplate-based a b initio li gand design s olution (Stalis), a knowledge-based approach that uses structure templates from the Protein Data Bank libraries of whole ligands and their fragments and generates a set of molecules (virtual ligands) whose structures represent the pocket shape and chemical features of a given target binding site. Our benchmark performance evaluation shows that ligand structure-based virtual screening using virtual ligands from Stalis outperforms a receptor structure-based virtual screening using AutoDock Vina, demonstrating reliable overall screening performance applicable to computational high-throughput screening. However, virtual ligands from Stalis are worse in recognizing active compounds at the small fraction of a rank-ordered list of screened library compounds than crystal ligands, due to the low resolution of the virtual ligand structures. In conclusion, Stalis can facilitate drug discovery research by designing virtual ligands that can be used for fast ligand structure-based virtual screening. Moreover, Stalis provides actual three-dimensional ligand structures that likely bind to a target protein, enabling to gain structural insight into potential ligands. Stalis can be an efficient computational platform for high-throughput ligand design for fundamental biological study and drug discovery research at the proteomic level. © 2019 Wiley Periodicals, Inc.     

Optical Modulation of Antibiotic Resistance by Photoswitchable Cystobactamids

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.     

A Photochromic Agonist for μ‐Opioid Receptors

Opioid receptors (ORs) are widely distributed in the brain, the spinal cord, and the digestive tract and play an important role in nociception. All known ORs are G‐protein‐coupled receptors (GPCRs) of family A. Another well‐known member of this family, rhodopsin, is activated by light through the cis/trans isomerization of a covalently bound chromophore, retinal. We now show how an OR can be combined with a synthetic azobenzene photoswitch to gain light sensitivity. Our work extends the reach of photopharmacology and outlines a general strategy for converting Family A GPCRs, which account for the majority of drug targets, into photoreceptors.     

Reversible Handedness Inversion and Circularly Polarized Light Reflection Tuning in Self-Organized Helical Superstructures Using Visible-Light-Driven Macrocyclic Chiral Switches

A series of macrocyclic azobenzene-based chiral photoswitches have been judiciously designed, synthesized, and characterized. In the molecular structures, binaphthyl is covalently linked to ortho-positions of azobenzene, and four different substituents are linked to 6,6′-positions of binaphthyl. The photoswitches show enhanced helical twisting power (HTP) when doping in commercially available achiral liquid crystals to form self-organized helical superstructures, i.e., cholesteric liquid crystals (CLCs). All the photoswitches exhibit reversible photoisomerization driven by visible light of different wavelengths in both organic solvent and liquid crystals. The photoswitches with shorter substituents enable handedness inversion of CLCs upon photoisomerization. These are the first examples of ortho-linked azobenzene-based photoswitches that enable handedness inversion in CLCs. The photoswitches with longer substituents display only HTP values decreasing while maintaining the same handedness.     

Photoresponsive molecular tools for emerging applications of light in medicine

Light-based therapeutic and imaging modalities, which emerge in clinical applications, rely on molecular tools, such as photocleavable protecting groups and photoswitches that respond to photonic stimulus and translate it into a biological effect. However, optimisation of their key parameters (activation wavelength, band separation, fatigue resistance and half-life) is necessary to enable application in the medical field. In this perspective, we describe the applications scenarios that can be envisioned in clinical practice and then we use those scenarios to explain the necessary properties that the photoresponsive tools used to control biological function should possess, highlighted by examples from medical imaging, drug delivery and photopharmacology. We then present how the (photo)chemical parameters are currently being optimized and an outlook is given on pharmacological aspects (toxicity, solubility, and stability) of light-responsive molecules. With these interdisciplinary insights, we aim to inspire the future directions for the development of photocontrolled tools that will empower clinical applications of light.

This perspective article explores the current state of light-controlled molecular tools for medical therapy and imaging and offers an outlook on clinical application scenarios and optimisation strategies.     

Visible-Light Photoswitchable Benzimidazole Azo-Arenes as β-Arrestin2-Biased Selective Cannabinoid 2 Receptor Agonists

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-Gα_i. Overall, compound 10 d is the first light-dependent functionally selective agonist to investigate the complex mechanisms of CB₂R-βarr2 dependent endocytosis.     

Steered molecular dynamics simulations of protein-ligand interactions

Molecular recognition and specific protein-ligandinteractions are central to many biochemical processes,such as enzyme catalysis, assembly of organelles, en-ergy transduction, signaling, diverse control functions,and replication, expression and storage of the geneticmaterial[1]. Moreover, protein-ligand interactions pro-vide the mechanism of many drug therapies and un-derstanding of such interactions is thus significant forrational drug design[1,2]. For the experimental studiesof protein-ligan…     

In silico study directed towards identification of the key structural features of GyrB inhibitors targeting MTB DNA gyrase: HQSAR,CoMSIA and molecular dynamics simulations

ABSTRACT

Mycobacterium tuberculosis DNA gyrase subunit B (GyrB) has been identified as a promising target for rational drug design against fluoroquinolone drug-resistant tuberculosis. In this study, we attempted to identify the key structural feature for highly potent GyrB inhibitors through 2D-QSAR using HQSAR, 3D-QSAR using CoMSIA and molecular dynamics (MD) simulations approaches on a series of thiazole urea core derivatives. The best HQSAR and CoMSIA models based on IC₅₀ and MIC displayed the structural basis required for good activity against both GyrB enzyme and mycobacterial cell. MD simulations and binding free energy analysis using MM-GBSA and waterswap calculations revealed that the urea core of inhibitors has the strongest interaction with Asp79 via hydrogen bond interactions. In addition, cation-pi interaction and hydrophobic interactions of the R₂ substituent with Arg82 and Arg141 help to enhance the binding affinity in the GyrB ATPase binding site. Thus, the present study provides crucial structural features and a structural concept for rational design of novel DNA gyrase inhibitors with improved biological activities against both enzyme and mycobacterial cell, and with good pharmacokinetic properties and drug safety profiles.     

Allosteric Optical Control of a Class B G‐Protein‐Coupled Receptor

Allosteric regulation promises to open up new therapeutic avenues by increasing drug specificity at G‐protein‐coupled receptors (GPCRs). However, drug discovery efforts are at present hampered by an inability to precisely control the allosteric site. Herein, we describe the design, synthesis, and testing of PhotoETP, a light‐activated positive allosteric modulator of the glucagon‐like peptide‐1 receptor (GLP‐1R), a class B GPCR involved in the maintenance of glucose homeostasis in humans. PhotoETP potentiates Ca²⁺, cAMP, and insulin responses to glucagon‐like peptide‐1 and its metabolites following illumination of cells with blue light. PhotoETP thus provides a blueprint for the production of small‐molecule class B GPCR allosteric photoswitches, and may represent a useful tool for understanding positive cooperativity at the GLP‐1R.     

Classification of water molecules in protein binding sites

Water molecules play a crucial role in mediating the interaction between a ligand and a macromolecular receptor. An understanding of the nature and role of each water molecule in the active site of a protein could greatly increase the efficiency of rational drug design approaches: if the propensity of a water molecule for displacement can be determined, then synthetic effort may be most profitably applied to the design of specific ligands with the displacement of this water molecule in mind. In this paper, a thermodynamic analysis of water molecules in the binding sites of six proteins, each complexed with a number of inhibitors, is presented. Two classes of water molecules were identified: those conserved and not displaced by any of the ligands, and those that are displaced by some ligands. The absolute binding free energies of 54 water molecules were calculated using the double decoupling method, with replica exchange thermodynamic integration in Monte Carlo simulations. It was found that conserved water molecules are on average more tightly bound than displaced water molecules. In addition, Bayesian statistics is used to calculate the probability that a particular water molecule may be displaced by an appropriately designed ligand, given the calculated binding free energy of the water molecule. This approach therefore allows the numerical assessment of whether or not a given water molecule should be targeted for displacement as part of a rational drug design strategy.     

Inside Cover: Ligand Assisted Thermal Atomization of Palladium Clusters: An Inspiring Approach for the Rational Design of Atomically Dispersed Metal Catalysts (Angew. Chem. Int. Ed. 16/2023)

The transformation from metal nanocluster catalysts to metal single-atom catalysts is an important procedure in the rational design of atomically dispersed metal catalysts (ADCs). However, the conversion methods often involve high annealing temperature as well as reducing atmosphere. Herein, we reported a continuous and convenient approach to transfer Pd nanocluster into Pd single-atom in a ligand assisted annealing procedure, by which means we reduced its activating temperature low to 400 °C. Using ex-situ microscopy and spectroscopy, we comprehensively monitored the structural evolution of Pd species though the whole atomization process. Theoretical calculation revealed that the structural instability caused by remaining Cl ligands was the main reason for this low-temperature transformation. The present atomization strategy and mechanistic knowledge for the conversion from cluster to atomic dispersion provides guidelines for the rational design of ADCs.     

Structural and Photophysical Properties of Various Polypyridyl Ligands: A Combined Experimental and Computational Study

Covalent triazine frameworks (CTFs) with polypyridyl ligands are very promising supports to anchor photocatalytic complexes. Herein, we investigate the photophysical properties of a series of ligands which vary by the extent of the aromatic system, the nitrogen content and their topologies to aid in selecting interesting building blocks for CTFs. Interestingly, some linkers have a rotational degree of freedom, allowing both a trans and cis structure, where only the latter allows anchoring. Therefore, the influence of the dihedral angle on the UV-Vis spectrum is studied. The photophysical properties are investigated by a combined computational and experimental study. Theoretically, both static and molecular dynamics simulations are performed to deduce ground- and excited state properties based on density functional theory (DFT) and time-dependent DFT. The position of the main absorption peak shifts towards higher wavelengths for an increased size of the π-system and a higher π-electron deficiency. We found that the position of the main absorption peak among the different ligands studied in this work can amount to 271 nm; which has a significant impact on the photophysical properties of the ligands. This broad range of shifts allows modulation of the electronic structure by varying the ligands and may help in a rational design of efficient photocatalysts.     

Phenothiazine-based chalcones as potential dual-target inhibitors toward cholinesterases (AChE,BuChE) and monoamine oxidases (MAO-A,MAO-B)

Chalcones targeting neurodegenerative diseases have been known as attractive structures in drug design and discovery. In this study, phenothiazine-based chalcones as ChEs and MAOs inhibitors were designed and synthesized via base-catalyzed Claisen-Schmidt condensation, and chemical structures of the compounds were elucidated by NMRs and HRMS. Compounds 3 and 9 showed promising inhibition potency against AChE enzyme with IC₅₀ values of 0.221 μM and 0.053 μM while compound 9 displayed remarkable inhibition potency toward MAO-B enzyme with IC₅₀ value of 0.048 μM. Compound 9 , as a dual-target inhibitor, selectively inhibited AChE and MAO-B enzymes. This promising behavior is an advantage for the compound since MAO-B and AChE inhibition have a role in Alzheimer's disease. Fused tricyclic ring systems such as phenothiazine incorporated with chalcone moiety being multitargeting ligands may help scientists for the rational design of novel lead compounds targeting neurodegenerative illnesses.     

New insights from molecular dynamic simulation studies of the multiple binding modes of a ligand with G-quadruplex DNA

G-quadruplexes are higher-order DNA and RNA structures formed from guanine-rich sequences. These structures have recently emerged as a new class of potential molecular targets for anticancer drugs. An understanding of the three-dimensional interactions between small molecular ligands and their G-quadruplex targets in solution is crucial for rational drug design and the effective optimization of G-quadruplex ligands. Thus far, rational ligand design has been focused mainly on the G-quartet platform. It should be noted that small molecules can also bind to loop nucleotides, as observed in crystallography studies. Hence, it would be interesting to elucidate the mechanism underlying how ligands in distinct binding modes influence the flexibility of G-quadruplex. In the present study, based on a crystal structure analysis, the models of a tetra-substituted naphthalene diimide ligand bound to a telomeric G-quadruplex with different modes were built and simulated with a molecular dynamics simulation method. Based on a series of computational analyses, the structures, dynamics, and interactions of ligand-quadruplex complexes were studied. Our results suggest that the binding of the ligand to the loop is viable in aqueous solutions but dependent on the particular arrangement of the loop. The binding of the ligand to the loop enhances the flexibility of the G-quadruplex, while the binding of the ligand simultaneously to both the quartet and the loop diminishes its flexibility. These results add to our understanding of the effect of a ligand with different binding modes on G-quadruplex flexibility. Such an understanding will aid in the rational design of more selective and effective G-quadruplex binding ligands.     

A Genetically Encoded,Phage‐Displayed Cyclic‐Peptide Library

Superior to linear peptides in biological activities, cyclic peptides are considered to have great potential as therapeutic agents. To identify cyclic‐peptide ligands for therapeutic targets, phage‐displayed peptide libraries in which cyclization is achieved by the covalent conjugation of cysteines have been widely used. To resolve drawbacks related to cysteine conjugation, we have invented a phage‐display technique in which its displayed peptides are cyclized through a proximity‐driven Michael addition reaction between a cysteine and an amber‐codon‐encoded N^?‐acryloyl‐lysine (AcrK). Using a randomized 6‐mer library in which peptides were cyclized at two ends through a cysteine–AcrK linker, we demonstrated the successful selection of potent ligands for TEV protease and HDAC8. All selected cyclic peptide ligands showed 4‐ to 6‐fold stronger affinity to their protein targets than their linear counterparts. We believe this approach will find broad applications in drug discovery.