Optical Control of GABAA Receptors with a Fulgimide-Based Potentiator

Optogenetic and photopharmacological tools to manipulate neuronal inhibition have limited efficacy and reversibility. We report the design, synthesis, and biological evaluation of Fulgazepam, a fulgimide derivative of benzodiazepine that behaves as a pure potentiator of ionotropic γ-aminobutyric acid receptors (GABA A Rs) and displays full and reversible photoswitching in vitro and in vivo. The compound enables high-resolution studies of GABAergic neurotransmission, and phototherapies based on localized, acute, and reversible neuroinhibition

A Photoswitchable Antimetabolite for Targeted Photoactivated Chemotherapy

The efficacy and tolerability of systemically administered anticancer agents are limited by their off-target effects. Precise spatiotemporal control over their cytotoxic activity would allow improving chemotherapy treatments, and light-regulated drugs are well suited to this purpose. We have developed phototrexate, the first photoswitchable inhibitor of the human dihydrofolate reductase (DHFR), as a photochromic analogue of methotrexate, a widely prescribed chemotherapeutic drug to treat cancer and psoriasis. Quantification of the light-regulated DHFR enzymatic activity, cell proliferation, and in vivo effects in zebrafish show that phototrexate behaves as a potent antifolate in its photoactivated cis configuration and that it is nearly inactive in its dark-relaxed trans form. Thus, phototrexate constitutes a proof-of-concept to design light-regulated cytotoxic small molecules and a step forward to develop targeted anticancer photochemotherapies with localized efficacy and reduced adverse effect

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

Gorostiza, Pa

Subunit-specific photocontrol of glycine receptors by azobenzene-nitrazepam photoswitcher

© 2021 Maleeva et al. Photopharmacology is a unique approach that through a combination of photochemistry methods and advanced life science techniques allows the study and control of specific biological processes, ranging from intracellular pathways to brain circuits. Recently, a first photochromic channel blocker of anion-selective GABAA receptors, the azobenzene-nitrazepam-based photochromic compound (Azo-NZ1), has been described. In the present study, using patch-clamp technique in heterologous system and in mice brain slices, site-directed mutagenesis and molecular modeling we provide evidence of the interaction of Azo-NZ1 with glycine receptors (GlyRs) and determine the molecular basis of this interaction. Glycinergic synaptic neurotransmission determines an important inhibitory drive in the vertebrate nervous system and plays a crucial role in the control of neuronal circuits in the spinal cord and brain stem. GlyRs are involved in locomotion, pain sensation, breathing, and auditory function, as well as in the development of such disorders as hyperekplexia, epilepsy, and autism. Here, we demonstrate that Azo-NZ1 blocks in a UV-dependent manner the activity of a2 GlyRs (GlyR2), while being barely active on a1 GlyRs (GlyR1). The site of Azo-NZ1 action is in the chloride-selective pore of GlyR at the 2’ position of transmembrane helix 2 and amino acids forming this site determine the difference in Azo-NZ1 blocking activity between GlyR2 and GlyR1. This subunit-specific modulation is also shown on motoneurons of brainstem slices from neonatal mice that switch during development from expressing “fetal” GlyR2 to “adult” GlyR1 receptors

Optical Control of Cardiac Function with a Photoswitchable Muscarinic Agonist

Light-triggered reversible modulation of physiological functions offers the promise of enabling on-demand spatiotemporally controlled therapeutic interventions. Optogenetics has been successfully implemented in the heart, but significant barriers to its use in the clinic remain, such as the need for genetic transfection. Herein, we present a method to modulate cardiac function with light through a photoswitchable compound and without genetic manipulation. The molecule, named PAI, was designed by introduction of a photoswitch into the molecular structure of an M2 mAChR agonist. In vitro assays revealed that PAI enables light-dependent activation of M2 mAChRs. To validate the method, we show that PAI photoisomers display different cardiac effects in a mammalian animal model, and demonstrate reversible, real-time photocontrol of cardiac function in translucent wildtype tadpoles. PAI can also effectively activate M2 receptors using two-photon excitation with near-infrared light, which overcomes the scattering and low penetration of short-wave-length illumination, and offers new opportunities for intravital imaging and control of cardiac function

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

Adrenergic Modulation With Photochromic Ligands

© 2020 Wiley-VCH GmbH Adrenoceptors are ubiquitous and mediate important autonomic functions as well as modulating arousal, cognition, and pain on a central level. Understanding these physiological processes and their underlying neural circuits requires manipulating adrenergic neurotransmission with high spatio-temporal precision. Here we present a first generation of photochromic ligands (adrenoswitches) obtained via azologization of a class of cyclic amidines related to the known ligand clonidine. Their pharmacology, photochromism, bioavailability, and lack of toxicity allow for broad biological applications, as demonstrated by controlling locomotion in zebrafish and pupillary responses in mice

Development and characterization of in vivo models for Photopharmacology

[eng] The main aim of this thesis is setting up and using in vivo screening techniques to identify and characterize new photopharmacological compounds targeting endogenous proteins. In order to achieve that aim, the thesis has been structured in three specific objectives, which are developed in the corresponding parts and chapters indicated below: Objective I. Photomanipulation of cardiac activity and organism development (part I). - Establish a procedure to identify photoswitchable compounds affecting zebrafish larvae development based on phenotypic outcomes during embryogenesis and hatching (Chapter 2). - Identification of relevant morphological, anatomical, and organotypic traits that are affected by the departure compound of photoswitchable drugs (Chapter 2). - Study of phenotypic markers as a function of photoswitchable drug isoforms (Chapter 2). - Developing a strategy for real-time recording of early stage tadpoles’ cardiac function under controlled illumination (Chapter 3). - Developing a computational approach for video extracting readouts and analysis of cardiac parameters (Chapter 3). - Identification of photoswitchable compounds with reversible effects on tadpoles’ cardiac function (Chapter 3). Objective II. Photomanipulation of locomotion (part II). - Establish an experimental and analytical procedure to categorise differences in locomotion outcomes under two wavelengths illumination (Chapter 4). - Identify swimming profiles related to inhibitory neurotransmission pathways of zebrafish larvae in darkness and illumination (Chapter 4). - Devise a strategy to screen photoswitchable compounds using zebrafish larvae activity profiles and their alteration under varying light conditions (Chapter 4). - Identification and characterization of photoswitchable GABAAR drugs using zebrafish locomotion assays (Chapter 5). - Identify swimming profiles related to inhibitory neural transmission pathways present during locomotion in Xenopus tropicalis tadpoles under long relaxed dark periods and light stimuli (Chapter 6). - Identification and characterization of a photoswitchable potentiator of glycinergic transmission using tapole swimming assays (Chapter 6). - Identification and characterization of a photoswitchable ligands of adrenergic receptors using zebrafish swimming assays (Chapter 7). - Identification and characterization of a photoswitchable ligands of dopaminergic receptors using zebrafish swimming assays (Chapter 8). Objective III. Photomanipulation of visual function (part III). - Develop a model of acute blinding in zebrafish larvae by photoinduced retinal degeneration in order to test vision restoration drugs (Chapter 9). - Identification and characterization of locomotive responses in blinded larvae (Chapter 9). - Development and validation of an optokinetic visual-motor reflex assay to test vision restoration drugs (Chapter 9)

A Photoswitchable Antimetabolite for Targeted Photoactivated Chemotherapy

The efficacy and tolerability of systemically administered anticancer agents are limited by their off-target effects. Precise spatiotemporal control over their cytotoxic activity would allow improving chemotherapy treatments, and light-regulated drugs are well suited to this purpose. We have developed phototrexate, the first photoswitchable inhibitor of the human dihydrofolate reductase (DHFR), as a photochromic analogue of methotrexate, a widely prescribed chemotherapeutic drug to treat cancer and psoriasis. Quantification of the light-regulated DHFR enzymatic activity, cell proliferation, and in vivo effects in zebrafish show that phototrexate behaves as a potent antifolate in its photoactivated cis configuration and that it is nearly inactive in its dark-relaxed trans form. Thus, phototrexate constitutes a proof-of-concept to design light-regulated cytotoxic small molecules and a step forward to develop targeted anticancer photochemotherapies with localized efficacy and reduced adverse effect

A Photoswitchable Antimetabolite for Targeted Photoactivated Chemotherapy

The efficacy and tolerability of systemically administered anticancer agents are limited by their off-target effects. Precise spatiotemporal control over their cytotoxic activity would allow improving chemotherapy treatments, and light-regulated drugs are well suited to this purpose. We have developed phototrexate, the first photoswitchable inhibitor of the human dihydrofolate reductase (DHFR), as a photochromic analogue of methotrexate, a widely prescribed chemotherapeutic drug to treat cancer and psoriasis. Quantification of the light-regulated DHFR enzymatic activity, cell proliferation, and in vivo effects in zebrafish show that phototrexate behaves as a potent antifolate in its photoactivated cis configuration and that it is nearly inactive in its dark-relaxed trans form. Thus, phototrexate constitutes a proof-of-concept to design light-regulated cytotoxic small molecules and a step forward to develop targeted anticancer photochemotherapies with localized efficacy and reduced adverse effect