Structure-based identification of a G protein-biased allosteric modulator of cannabinoid receptor CB1.

Cannabis sativa is known for its therapeutic benefit in various diseases including pain relief by targeting cannabinoid receptors. The primary component of cannabis, Δ9-tetrahydrocannabinol (THC), and other agonists engage the orthosteric site of CB1, activating both Gi and ß-arrestin signaling pathways. The activation of diverse pathways could result in on-target side effects and cannabis addiction, which may hinder therapeutic potential. A significant challenge in pharmacology is the design of a ligand that can modulate specific signaling of CB1. By leveraging insights from the structure-function selectivity relationship (SFSR), we have identified Gi signaling-biased agonist-allosteric modulators (ago-BAMs). Further, two cryoelectron microscopy (cryo-EM) structures reveal the binding mode of ago-BAM at the extrahelical allosteric site of CB1. Combining mutagenesis and pharmacological studies, we elucidated the detailed mechanism of ago-BAM-mediated biased signaling. Notably, ago-BAM CB-05 demonstrated analgesic efficacy with fewer side effects, minimal drug toxicity and no cannabis addiction in mouse pain models. In summary, our finding not only suggests that ago-BAMs of CB1 provide a potential nonopioid strategy for pain management but also sheds light on BAM identification for GPCRs.

Astrocytes Modulate a Specific Paraventricular Thalamus→Prefrontal Cortex Projection to Enhance Consciousness Recovery from Anesthesia.

Current anesthetic theory is mostly based on neurons and/or neuronal circuits. A role for astrocytes also has been shown in promoting recovery from volatile anesthesia, while the exact modulatory mechanism and/or the molecular target in astrocytes is still unknown. In this study by animal models in male mice and electrophysiological recordings in vivo and in vitro, we found that activating astrocytes of the paraventricular thalamus (PVT) and/or knocking down PVT astrocytic Kir4.1 promoted the consciousness recovery from sevoflurane anesthesia. Single-cell RNA sequencing of the PVT reveals two distinct cellular subtypes of glutamatergic neurons: PVT GRM and PVT ChAT neurons. Patch-clamp recording results proved astrocytic Kir4.1-mediated modulation of sevoflurane on the PVT mainly worked on PVT ChAT neurons, which projected mainly to the mPFC. In summary, our findings support the novel conception that there is a specific PVT→prefrontal cortex projection involved in consciousness recovery from sevoflurane anesthesia, which is mediated by the inhibition of sevoflurane on PVT astrocytic Kir4.1 conductance.

Identification of (4-chlorophenyl)(5-hydroxynaphtho[1,2-b]furan-3-yl)methanone as novel COX-2 inhibitor with analgesic profile.

Chronic pain is a serious problem that affects billions of people worldwide, but current analgesic drugs limit their use in chronic pain management due to their respective side effects. As a first-line clinical drug for chronic pain, COX-2 selective inhibitors can relieve mild to moderate pain, but they also have some problems. The most prominent one is that their analgesic intensity is not enough, and they cannot well meet the treatment needs of chronic pain. Therefore, there is an urgent need to develop COX-2 inhibitors with stronger analgesic intensity. In this article, we used virtual screening method to screen out the structurally novel COX-2 inhibitor for chronic pain management, and conducted a preliminary study on its mechanism of action using molecular dynamics simulation.

Design, synthesis, and biological evaluation of new bis-benzylisoquinoline-based analogues as potential neuromuscular blocking agents.

Neuromuscular blocking agents (NMBAs) are widely used in anesthesia for intubation and surgical muscle relaxation. Novel atracurium and mivacurium derivatives were developed, with compounds 18c, 18d, and 29a showing mivacurium-like relaxation at 27.27 nmol/kg, and 15b, 15c, 15e, and 15h having a shorter duration at 272.7 nmol/kg. The structure-activity and configuration-activity relationships of these derivatives and 29a's binding to nicotinic acetylcholine receptors were analyzed through molecular docking. Rabbit trials showed 29a has a shorter duration compared to mivacurium. This suggests that linker properties, ammonium group substituents, and configuration are crucial for NMBA activity and duration, with compound 29a emerging as a potential ultra-short-acting NMBA.

Discovery of a novel series of pyridone amides as NaV1.8 inhibitors.

The NaV1.8 channel, mainly found in the peripheral nervous system, is recognized as one of the key factors in chronic pain. The molecule VX-150 was initially promising in targeting this channel, but the phase II trials of VX-150 did not show expected pain relief results. By analyzing the interaction mode of VX-150 and NaV1.8, we developed two series with a total of 19 molecules and examined their binding affinity to NaV1.8 in vitro and analgesic effect in vivo. One compound, named 2j, stood out with notable activity against the NaV1.8 channel and showed effective pain relief in models of chronic inflammatory pain and neuropathic pain. Our research points to 2j as a strong contender for developing safer pain-relief treatments.

Identification of N-(((1S,3R,5S)-adamantan-1-yl)methyl)-3-((4-chlorophenyl)sulfonyl)benzenesulfonamide as novel Nav1.8 inhibitor with analgesic profile.

Chronic pain is a common and challenging clinical problem that significantly impacts patients' quality of life. The sodium channel Nav1.8 plays a crucial role in the occurrence and development of chronic pain, making it one of the key targets for treating chronic pain. In this article, we combined virtual screening with cell membrane chromatography techniques to establish a novel method for rapid high-throughput screening of selective Nav1.8 inhibitors. Using this approach, we identified a small molecule compound 6, which not only demonstrated high affinity and inhibitory activity against Nav1.8 but also exhibited significant inhibitory effects on CFA-induced chronic inflammatory pain. Compared to the positive drug VX-150, compound 6 showed a more prolonged analgesic effect, making it a promising candidate as a Nav1.8 inhibitor with potential clinical applications. This discovery provides a new therapeutic option for the treatment of chronic pain.

Direct and Continuous Monitoring of Multicomponent Antibiotic Gentamicin in Blood at Single-Molecule Resolution.

Point-of-care monitoring of small molecules in biofluids is crucial for clinical diagnosis and treatment. However, the inherent low degree of recognition of small molecules and the complex composition of biofluids present significant obstacles for current detection technologies. Although nanopore sensing excels in the analysis of small molecules, the direct detection of small molecules in complex biofluids remains a challenge. In this study, we present a method for sensing the small molecule drug gentamicin in whole blood based on the mechanosensitive channel of small conductance in Pseudomonas aeruginosa (PaMscS) nanopore. PaMscS can directly detect gentamicin and distinguish its main components with only a monomethyl difference. The 'molecular sieve' structure of PaMscS enables the direct measurement of gentamicin in human whole blood within 10 min. Furthermore, a continuous monitoring device constructed based on PaMscS achieved continuous monitoring of gentamicin in live rats for approximately 2.5 h without blood consumption, while the drug components can be analyzed in situ. This approach enables rapid and convenient drug monitoring with single-molecule level resolution, which can significantly lower the threshold for drug concentration monitoring and promote more efficient drug use. Moreover, this work also lays the foundation for the future development of continuous monitoring technology with single-molecule level resolution in the living body.

Spatial transcriptomics and single-nucleus RNA sequencing reveal a transcriptomic atlas of adult human spinal cord.

Despite the recognized importance of the spinal cord in sensory processing, motor behaviors, and neural diseases, the underlying organization of neuronal clusters and their spatial location remain elusive. Recently, several studies have attempted to define the neuronal types and functional heterogeneity in the spinal cord using single-cell or single-nucleus RNA sequencing in animal models or developing humans. However, molecular evidence of cellular heterogeneity in the adult human spinal cord is limited. Here, we classified spinal cord neurons into 21 subclusters and determined their distribution from nine human donors using single-nucleus RNA sequencing and spatial transcriptomics. Moreover, we compared the human findings with previously published single-nucleus data of the adult mouse spinal cord, which revealed an overall similarity in the neuronal composition of the spinal cord between the two species while simultaneously highlighting some degree of heterogeneity. Additionally, we examined the sex differences in the spinal neuronal subclusters. Several genes, such as SCN10A and HCN1, showed sex differences in motor neurons. Finally, we classified human dorsal root ganglia (DRG) neurons using spatial transcriptomics and explored the putative interactions between DRG and spinal cord neuronal subclusters. In summary, these results illustrate the complexity and diversity of spinal neurons in humans and provide an important resource for future research to explore the molecular mechanisms underlying spinal cord physiology and diseases.

Molecular Transformers: Adaptive Multitarget Ligands for Esterase-Induced Transition from Analgesics to Anesthetics.

Multitarget strategies are essential in addressing complex diseases, yet developing multitarget-directed ligands (MTDLs) is particularly challenging when aiming to engage multiple therapeutic targets across different tissues. Here, we present a molecular transformer strategy, enhancing traditional MTDLs. By utilizing esterase-driven hydrolysis, this approach mimics the adaptive nature of transformers for enabling molecules to modify their pharmacological effects in response to the biological milieu. By virtual screening and biological evaluation, we identified KGP-25, a novel compound initially targeting the voltage-gated sodium channel 1.8 (Nav1.8) in the peripheral nervous system (PNS) for analgesia, and later the γ-aminobutyric acid subtype A receptor (GABAA) in the central nervous system (CNS) for general anesthesia. Our findings confirm KGP-25's dual efficacy in cellular and animal models, effectively reducing opioid-related side effects. This study validates the molecular transformer approach in drug design and highlights its potential to overcome the limitations of conventional MTDLs, paving new avenues in innovative therapeutic strategies.

Deep simulated annealing for the discovery of novel dental anesthetics with local anesthesia and anti-inflammatory properties.

Multifunctional therapeutics have emerged as a solution to the constraints imposed by drugs with singular or insufficient therapeutic effects. The primary challenge is to integrate diverse pharmacophores within a single-molecule framework. To address this, we introduced DeepSA, a novel edit-based generative framework that utilizes deep simulated annealing for the modification of articaine, a well-known local anesthetic. DeepSA integrates deep neural networks into metaheuristics, effectively constraining molecular space during compound generation. This framework employs a sophisticated objective function that accounts for scaffold preservation, anti-inflammatory properties, and covalent constraints. Through a sequence of local editing to navigate the molecular space, DeepSA successfully identified AT-17, a derivative exhibiting potent analgesic properties and significant anti-inflammatory activity in various animal models. Mechanistic insights into AT-17 revealed its dual mode of action: selective inhibition of NaV1.7 and 1.8 channels, contributing to its prolonged local anesthetic effects, and suppression of inflammatory mediators via modulation of the NLRP3 inflammasome pathway. These findings not only highlight the efficacy of AT-17 as a multifunctional drug candidate but also highlight the potential of DeepSA in facilitating AI-enhanced drug discovery, particularly within stringent chemical constraints.