ACS Chemical Neuroscience最新文献

Alzheimer's disease (AD) remains an incurable neurodegenerative disorder, requiring novel therapeutic strategies. We developed multitarget-directed ligands designed to inhibit human butyrylcholinesterase (hBChE) and activate the sigma-1 receptor (S1R), addressing both cholinergic dysfunction and neuroinflammation, the latter being reduced through action on both targets. The (pseudo-)irreversible carbamate inhibitor 18c emerged as the most promising compound, exhibiting potent and selective hBChE inhibition (IC₅₀ = 3.3 nM, 45-fold selectivity over human acetylcholinesterase) and strong S1R agonistic activity (IC₅₀ = 25 nM, EC₅₀ = 99 nM) determined in a radioligand binding assay and by S1R-BiP dissociation, respectively. Its cleavage product 14c (after carbamate hydrolysis by hBChE) retained dual activity (IC₅₀(hBChE) = 269 nM, IC₅₀(S1R) = 20 nM, and EC₅₀(S1R) = 279 nM). Both compounds reduced the lipopolysaccharide-induced pro-inflammatory activation profile in microglial N9 cells while preserving anti-inflammatory marker expression, thereby indicating an overall immunomodulatory effect. In vivo, inhibitor 18c improved cognitive deficits in a mouse model with Aβ_25-35-induced neurotoxicity, enhancing short- and long-term memory in Y-maze and passive avoidance tests at dosages as low as 0.1-1 mg/kg. These findings highlight the potential of dual-targeting hBChE/S1R strategies for AD therapy.

NLRP3 inflammasome activation has been implicated in the pathogenesis of human disorders, including a number of neurodegenerative diseases of the central nervous system. Pharmacological inhibition of NLRP3-mediated neuroinflammation via a brain-penetrant therapeutic agent, therefore, is an attractive target for the treatment of neurodegenerative disorders. The NLRP3 inflammasome inhibitor NT-0796 is an ester-containing prodrug that is metabolized intracellularly into its active metabolite NDT-19795 by carboxylesterase-1 (CES1). NT-0796 shows high in vitro permeability in an induced pluripotent stem cells (iPSC)-derived human brain endothelial blood–brain barrier (BBB) model, while nonhuman primate pharmacokinetics studies demonstrate the ability of NT-0796 to cross the BBB, which is then metabolized locally to give rise to its active metabolite, NDT-19795 in vivo. While human and nonhuman primate cerebrospinal fluid are incapable of NT-0796 metabolism, brain parenchymal tissue homogenates obtained from either species metabolized NT-0796 to NDT-19795. Data presented in this manuscript also offer a mechanistic explanation for the central exposure levels of NDT-19795 achieved following NT-0796 administration in our Phase 1b open-label study conducted in elderly healthy volunteers and patients with Parkinson’s disease, recently published in Movement Disorders. Immunolocalization of nonhuman primate frontal tissues indicates that CES1 is expressed by a subpopulation of cells, many of which were also ionized calcium-binding adapter molecule 1 (IBA1) positive. Collectively, these data show that the ester-containing prodrug, NT-0796, is highly brain penetrant and, after crossing the BBB, NT-0796 is metabolized predominantly by CES1 in brain parenchyma. Immunolocalization studies also suggest that the subset of cells in the brain expressing CES1 is microglial in origin.

This work describes progress toward an M₄ PAM preclinical candidate. The SAR to address potency, clearance, subtype selectivity, CNS exposure, and P-gp efflux are detailed within. A novel 1-(7,8-dimethyl-[1,2,4]triazolo[4,3-b]pyridazin-6-yl)piperidin-4-ol scaffold was identified, and optimization provided a highly potent analog VU6025733 (hM₄ EC₅₀ = 23 nM; rM₄ EC₅₀ = 55 nM). Further characterization revealed a highly selective compound across muscarinic acetylcholine receptor subtypes with exceptional DMPK properties (in vivo rat CL_p = 5.9 mL/min/kg; t_1/2 = 4.8 h; CYP1A2 & CYP2C9 IC₅₀s > 30 μM, CYP2D6 IC₅₀ > 9 μM; CYP3A4 IC₅₀ > 25 μM). Moreover, VU6025733 demonstrated robust in vivo efficacy in a rat amphetamine-induced hyperlocomotion model in a dose-dependent manner. However, hepatotoxicity risk precluded further development.

Herein, we report the structure–activity relationship (SAR) to develop novel mGlu₅ negative allosteric modulator (NAM) scaffolds devoid of the aryl/heterobiaryl acetylene moiety found in many historic mGlu₅ NAMs, which has been linked to metabolic liabilities and hepatotoxicity. This endeavor utilized a scaffold-hopping strategy from the predecessor compound VU6031545, in which we replace an ether-linked tetrahydrofuran with various carbon-linked heteroaryl motifs to generate highly potent and selective mGlu₅ NAMs. One such compound, VU6035386, displayed low nanomolar potency against human mGlu₅ and was highly brain penetrant. Moreover, VU6035386 showed a vast improvement in predicted human hepatic clearance versus predecessor compound VU6031545.

Trigeminal neuralgia is a debilitating neuropathic pain disorder characterized by facial hypersensitivity, yet its underlying molecular mechanisms remain incompletely understood. Using a mouse model of partial infraorbital nerve transection (pT-ION), we investigated transcriptomic alterations in the trigeminal ganglion (TG) to identify molecular contributors to orofacial pain. Microarray analysis identified 200 differentially expressed genes, with functional enrichment highlighting immune-related processes, calcium signaling, and lysosomal pathways. Among these, Calb2 (calretinin) emerged as a hub gene in both coexpression and protein–protein interaction networks. Immunofluorescence analysis revealed prominent colocalization of calretinin with the voltage-gated calcium channel auxiliary subunit α2δ1 (Cavα2δ1) in TG neurons. Functionally, a single perineural injection of calretinin siRNA into the trigeminal nerve significantly alleviated mechanical and cold hypersensitivity in both the maxillary and mandibular facial regions within 48 h. Pharmacological inhibition of Cavα2δ1 with gabapentin similarly attenuated pain behaviors and reduced calretinin expression in the TG. Conversely, targeted overexpression of Cavα2δ1 in naïve mice was sufficient to induce orofacial hypersensitivity and to upregulate calretinin expression in the TG. Together, these findings identify calretinin as a key downstream contributor to Cavα2δ1-associated trigeminal pain signaling and suggest that modulation of the Cavα2δ1–calretinin axis may represent a potential therapeutic strategy for trigeminal neuropathic pain.

Circadian rhythms (CRs) are intrinsic 24 h cycles that regulate critical physiological processes, including sleep-wake behavior, hormonal signaling, and cognition. Disruption of CRs, often caused by chronic aberrant light exposure, has been linked to neurodegenerative diseases such as Alzheimer’s disease (AD), through altered expression of core clock genes and neurotransmitter imbalances. Estrogen is a known neuromodulator that influences both circadian timing and cognitive function, yet the mechanistic interplay between estrogen and circadian dysregulation in neurodegeneration remains underexplored. In this study, we investigated whether estradiol could mitigate neuropathological and circadian disturbances induced by chronic, constant light (LL) exposure in female C57BL/6J mice. Mice were exposed to LL for 6 or 10 weeks (LL₆, LL₁₀) to model progressive CR disruption. LL₁₀ significantly delayed locomotor rhythms (p < 0.0001), elevated hippocampal amyloid-β (Aβ) levels (p = 0.0018), and reduced SCN GABA and glutamate levels (p < 0.01), compared to LL₆ and light-dark (LD) controls. Both LL₆ and LL₁₀ also showed decreased hippocampal nitric oxide and glutathione levels (p < 0.05), indicating oxidative stress. Estradiol treatment (1.5 or 3 μg/kg) restored activity rhythms, reduced Aβ accumulation (p = 0.0019), and normalized SCN neurotransmitter levels (GABA; p = 0.0046; glutamate: p = 0.0003). These effects were abrogated by tamoxifen, suggesting estrogen receptor-mediated signaling. Histological analysis further showed that estradiol attenuated hippocampal inflammation and neuronal damage in LL₁₀-exposed animals. These results demonstrate that estrogen protects against circadian disruption-induced neuropathology and supports its potential as a therapeutic agent in mitigating cognitive decline via ER-dependent pathways.

Intracellular deposits of neurofibrillary tau tangles and extracellular Aβ plaques are closely associated with Alzheimer’s disease. The mapping of thermodynamic parameters, including solubility limits, indicates when a protein forms amyloid fibrils or remains in solution. This reveals the direction of change of the system and may help in understanding drift and steady states in living systems. Here we have developed methodology for tau solubility quantification and determined the solubility of the amyloidogenic core fragment of tau in vitro. We monitored the concentration of free tau304–380C322S fragment at 37 °C in phosphate buffer at pH 8.0 using three separate methods: HPLC-UV, derivatization with ortho-phthalaldehyde and scintillation counting. The measurements were repeated over time until a stable value was reached, implying that an equilibrium with fibrils had been established. The solubility measurements converged on a free monomer concentration of 6.1 ± 3.5 nM, which represents the solubility of the fragment under the current experimental conditions.

Intrinsically disordered proteins (IDPs) like α-synuclein are pivotal in neurodegenerative diseases but present formidable challenges for drug discovery due to their conformational heterogeneity and lack of defined binding pockets. Here, we report the finding that clinically approved tricyclic antidepressants imipramine, amitriptyline, and doxepin bind directly to monomeric α-synuclein and inhibit its fibrillation, a key process in Parkinson’s pathology. Using a combination of NMR spectroscopy (WaterLOGSY and chemical shift perturbation, CSP) and molecular dynamics simulation methods, we elucidate a nonspecific, multiligand binding mechanism of tricyclic antidepressants predominantly targeting the C-terminal domain of the protein. The experimental CSP magnitudes correlate with the inhibitory potency of ligands, with imipramine emerging as the most potent inhibitor, and agree with the contact probabilities with different residues from simulations. The protein–ligand binding is driven by a dynamic combination of electrostatic attraction of positively charged ligands to anionic side chains, π-stacking with aromatic residues, and hydrophobic contacts. Moreover, simulations show that a single ligand molecule frequently engages in simultaneous salt-bridge and π-stacking interactions, unlike some previously studied α-synuclein binders. Our findings position tricyclic antidepressants as promising scaffolds for targeting α-synuclein and demonstrate the efficiency of molecular dynamics approaches for the description of interactions between small drug molecules and IDPs.

Hypomyelinating diseases are a heterogeneous group of neurodevelopmental disorders caused by genetic anomalies that impair myelin formation or maintenance. Here, we investigate a novel homozygous missense variant in FRMD3 (NM_174938.6:c.898T > C; p.C300R) in a 2-year-old male presenting with global developmental delay, hypotonia, mild ataxia, and MRI features consistent with hypomyelination. The variant affects a highly conserved cysteine within the FERM domain of FRMD3, and predicted to be deleterious by multiple in silico tools. Molecular dynamics simulations and biophysical analyses revealed that p.C300R introduces steric clashes with neighboring residues, destabilizes the FERM domain, increases structural disorder, and exposes hydrophobic aggregation-prone regions. In proband fibroblasts and nonproband neuronal cells, mutant FRMD3 mislocalized from the plasma membrane to the cytosol, forming large aggregates. Thioflavin T assays confirmed elevated aggregation propensity of the mutant. In oligodendrocytes, FRMD3-p.C300R expression markedly impaired neurite formation and failed to restore proteolipid protein 1 (PLP1) and myelin basic protein (MBP) expression following FRMD3 knockdown, in contrast to wild-type rescue. Interactome and single-cell expression analyses place FRMD3 at membrane-trafficking and lipid-handling hubs in oligodendrocytes and white-matter regions, and loss of these interactions through p.C300R-driven destabilization and aggregation likely underlies the regional hypomyelination observed in the proband. Our findings establish FRMD3 as a novel candidate gene for hypomyelinating disease and reveal that structural destabilization and aggregation of FERM-domain peripheral membrane protein can disrupt oligodendrocyte function and myelin protein expression, leading to neurodevelopmental pathology.

Traumatic brain injury (TBI) represents a major global public health challenge. It is propelled by a cascade of secondary injuries. These injuries include inflammation, endothelial dysfunction, hypoxia, cerebral edema, and disruption of epigenetic homeostasis. These processes can precipitate necrosis and apoptosis. They also significantly heighten the risk of long-term cognitive deficits, dementia, and other neurodegenerative disorders. TBI progression is typically segmented into acute and chronic phases. Each phase is characterized by distinct pathological mechanisms and epigenetic alterations. The acute phase is dominated by direct tissue damage and robust inflammatory responses. In contrast, chronic TBI often evolves into long-term neurodegenerative conditions like chronic traumatic encephalopathy (CTE). CTE is marked by persistent neuroinflammation and cognitive decline. A critical gap exists in prior research. It lies in the frequent failure to disentangle the unique epigenetic reprogramming specific to each phase. This failure hinders the development of precisely timed interventions. This review systematically delineates the spatiotemporal dynamics of epigenetic regulation following TBI. It aims to construct a phase-specific framework for precision intervention. Acute-phase hallmarks involve DNA methylation. An example is DNMT3A-mediated silencing of homeostatic genes. They also include histone acetylation and m6A RNA methylation. The WTAP/YTHDF1-Lcn2 axis exemplifies this m6A regulation. Conversely, the chronic phase is defined by sustained neuroinflammation, tau hyperphosphorylation, and ferroptosis. These processes are modulated by noncoding RNAs. Examples include miR-29b and lncRNA 4933431K23Rik. Epigenetic drift also plays a regulatory role. Mitochondrial and endoplasmic reticulum stress further interact with these pathways. They amplify secondary damage. We underscore the clinical promise of time-stratified, personalized epigenetic interventions. These interventions aim to improve long-term outcomes. They forge a critical link between fundamental epigenetic discovery and precision management of neurotrauma. This work deepens the understanding of TBI pathophysiology. It also lays a conceptual and target-oriented groundwork. This groundwork advances neurotrauma care into an era of temporally tailored, individualized precision therapy.