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Nature Oct 2023Type A γ-aminobutyric acid receptors (GABARs) are the principal inhibitory receptors in the brain and the target of a wide range of clinical agents, including...
Type A γ-aminobutyric acid receptors (GABARs) are the principal inhibitory receptors in the brain and the target of a wide range of clinical agents, including anaesthetics, sedatives, hypnotics and antidepressants. However, our understanding of GABAR pharmacology has been hindered by the vast number of pentameric assemblies that can be derived from 19 different subunits and the lack of structural knowledge of clinically relevant receptors. Here, we isolate native murine GABAR assemblies containing the widely expressed α1 subunit and elucidate their structures in complex with drugs used to treat insomnia (zolpidem (ZOL) and flurazepam) and postpartum depression (the neurosteroid allopregnanolone (APG)). Using cryo-electron microscopy (cryo-EM) analysis and single-molecule photobleaching experiments, we uncover three major structural populations in the brain: the canonical α1β2γ2 receptor containing two α1 subunits, and two assemblies containing one α1 and either an α2 or α3 subunit, in which the single α1-containing receptors feature a more compact arrangement between the transmembrane and extracellular domains. Interestingly, APG is bound at the transmembrane α/β subunit interface, even when not added to the sample, revealing an important role for endogenous neurosteroids in modulating native GABARs. Together with structurally engaged lipids, neurosteroids produce global conformational changes throughout the receptor that modify the ion channel pore and the binding sites for GABA and insomnia medications. Our data reveal the major α1-containing GABAR assemblies, bound with endogenous neurosteroid, thus defining a structural landscape from which subtype-specific drugs can be developed.
Topics: Animals; Mice; Binding Sites; Cryoelectron Microscopy; Depression, Postpartum; Flurazepam; gamma-Aminobutyric Acid; Hypnotics and Sedatives; Ion Channel Gating; Neurosteroids; Photobleaching; Pregnanolone; Protein Conformation; Protein Subunits; Receptors, GABA-A; Sleep Initiation and Maintenance Disorders; Zolpidem
PubMed: 37730991
DOI: 10.1038/s41586-023-06556-w -
British Journal of Pharmacology Oct 2021Neurosteroids influence neuronal function and have multiple promising clinical applications. Direct modulation of postsynaptic neurotransmitter receptors by...
BACKGROUND AND PURPOSE
Neurosteroids influence neuronal function and have multiple promising clinical applications. Direct modulation of postsynaptic neurotransmitter receptors by neurosteroids is well characterized, but presynaptic effects remain poorly understood. Here, we report presynaptic glutamate release potentiation by neurosteroids pregnanolone and pregnanolone sulfate and compare their mechanisms of action to phorbol 12,13-dibutyrate (PDBu), a mimic of the second messenger DAG.
EXPERIMENTAL APPROACH
We use whole-cell patch-clamp electrophysiology and pharmacology in rat hippocampal microisland cultures and total internal reflection fluorescence (TIRF) microscopy in HEK293 cells expressing GFP-tagged vesicle priming protein Munc13-1, to explore the mechanisms of neurosteroid presynaptic modulation.
KEY RESULTS
Pregnanolone sulfate and pregnanolone potentiate glutamate release downstream of presynaptic Ca influx, resembling the action of a phorbol ester PDBu. PDBu partially occludes the effect of pregnanolone, but not of pregnanolone sulfate. Calphostin C, an inhibitor that disrupts DAG binding to its targets, reduces the effect PDBu and pregnanolone, but not of pregnanolone sulfate, suggesting that pregnanolone might interact with a well-known DAG/phorbol ester target Munc13-1. However, TIRF microscopy experiments found no evidence of pregnanolone-induced membrane translocation of GFP-tagged Munc13-1, suggesting that pregnanolone may regulate Munc13-1 indirectly or interact with other DAG targets.
CONCLUSION AND IMPLICATIONS
We describe a novel presynaptic effect of neurosteroids pregnanolone and pregnanolone sulfate to potentiate glutamate release downstream of presynaptic Ca influx. The mechanism of action of pregnanolone, but not of pregnanolone sulfate, partly overlaps with that of PDBu. Presynaptic effects of neurosteroids may contribute to their therapeutic potential in the treatment of disorders of the glutamate system.
Topics: Animals; Glutamic Acid; HEK293 Cells; Humans; Neurosteroids; Pregnanolone; Rats; Sulfates
PubMed: 33988248
DOI: 10.1111/bph.15529 -
The Lancet. Neurology May 2022CDKL5 deficiency disorder (CDD) is a rare, X-linked, developmental and epileptic encephalopathy characterised by severe global developmental impairment and seizures that... (Randomized Controlled Trial)
Randomized Controlled Trial
BACKGROUND
CDKL5 deficiency disorder (CDD) is a rare, X-linked, developmental and epileptic encephalopathy characterised by severe global developmental impairment and seizures that can begin in the first few months after birth and are often treatment refractory. Ganaxolone, an investigational neuroactive steroid, reduced seizure frequency in an open-label, phase 2 trial that included patients with CDD. We aimed to further assess the efficacy and safety of ganaxolone in patients with CDD-associated refractory epilepsy.
METHODS
In the double-blind phase of this randomised, placebo-controlled, phase 3 trial, done at 39 outpatient clinics in eight countries (Australia, France, Israel, Italy, Poland, Russia, the UK, and the USA), patients were eligible if they were aged 2-21 years with a pathogenic or probably pathogenic CDKL5 variant and at least 16 major motor seizures (defined as bilateral tonic, generalised tonic-clonic, bilateral clonic, atonic, or focal to bilateral tonic-clonic) per 28 days in each 4-week period of an 8-week historical period. After a 6-week prospective baseline period, patients were randomly assigned (1:1) via an interactive web response system to receive either enteral adjunctive ganaxolone or matching enteral adjunctive placebo (maximum dose 63 mg/kg per day for patients weighing ≤28 kg or 1800 mg/day for patients weighing >28 kg) for 17 weeks. Patients, caregivers, investigators (including those analysing data), trial staff, and the sponsor (other than the investigational product manager) were masked to treatment allocation. The primary efficacy endpoint was percentage change in median 28-day major motor seizure frequency from the baseline period to the 17-week double-blind phase and was analysed (using a Wilcoxon-rank sum test) in all patients who received at least one dose of trial treatment and for whom baseline data were available. Safety (compared descriptively across groups) was analysed in all patients who received at least one dose of trial treatment. This study is registered with ClinicalTrials.gov, NCT03572933, and the open-label extension phase is ongoing.
FINDINGS
Between June 25, 2018, and July 2, 2020, 114 patients were screened for eligibility, of whom 101 (median age 6 years [IQR 3 to 10]) were randomly assigned to receive either ganaxolone (n=50) or placebo (n=51). All patients received at least one dose of a study drug, but seizure frequency for one patient in the ganaxolone group was not recorded at baseline and so the primary endpoint was analysed in a population of 100 patients. There was a median percentage change in 28-day major motor seizure frequency of -30·7% (IQR -49·5 to -1·9) in the ganaxolone group and of -6·9% (-24·1 to 39·7) in the placebo group (p=0·0036). The Hodges-Lehmann estimate of median difference in responses to ganaxolone versus placebo was -27·1% (95% CI -47·9 to - 9·6). Treatment-emergent adverse events occurred in 43 (86%) of 50 patients in the ganaxolone group and in 45 (88%) of 51 patients in the placebo group. Somnolence, pyrexia, and upper respiratory tract infections occurred in at least 10% of patients in the ganaxolone group and more frequently than in the placebo group. Serious adverse events occurred in six (12%) patients in the ganaxolone group and in five (10%) patients in the placebo group. Two (4%) patients in the ganaxolone group and four (8%) patients in the placebo group discontinued the trial. There were no deaths in the double-blind phase.
INTERPRETATION
Ganaxolone significantly reduced the frequency of CDD-associated seizures compared with placebo and was generally well tolerated. Results from what is, to our knowledge, the first controlled trial in CDD suggest a potential treatment benefit for ganaxolone. Long-term treatment is being assessed in the ongoing open-label extension phase of this trial.
FUNDING
Marinus Pharmaceuticals.
Topics: Child; Child, Preschool; Double-Blind Method; Epileptic Syndromes; Humans; Infant; Pregnanolone; Prospective Studies; Protein Serine-Threonine Kinases; Seizures; Spasms, Infantile; Treatment Outcome
PubMed: 35429480
DOI: 10.1016/S1474-4422(22)00077-1 -
Drugs Jun 2022Ganaxolone (ZTALMY; Marinus Pharmaceuticals) is a synthetic neuroactive steroid that acts as a positive allosteric modulator of the gamma-aminobutyric acid (GABA)... (Review)
Review
Ganaxolone (ZTALMY; Marinus Pharmaceuticals) is a synthetic neuroactive steroid that acts as a positive allosteric modulator of the gamma-aminobutyric acid (GABA) receptor complex. Ganaxolone received its first approval in March 2022 in the USA for the treatment of seizures associated with cyclin-dependent kinase-like 5 (CDKL5) deficiency disorder (CDD) in patients 2 years of age and older. Approval was based on the results of a multinational phase III trial, in which ganaxolone was effective in reducing seizure frequency in children and adolescents with CDD. In the EU, a Marketing Authorization Application has been filed for ganaxolone in the treatment of seizures associated with CDD and an opinion from the Committee for Medicinal Products for Human Use is expected later this year. Oral ganaxolone is also currently undergoing phase III evaluation in the treatment of tuberous sclerosis complex-related epilepsy, while an intravenous formulation of ganaxolone is being evaluated in refractory status epilepticus. This article summarizes the milestones in the development of ganaxolone leading to this first approval for seizures associated with CDD.
Topics: Adolescent; Anticonvulsants; Child; Clinical Trials, Phase III as Topic; Drug Approval; Humans; Multicenter Studies as Topic; Pregnanolone; Seizures
PubMed: 35596878
DOI: 10.1007/s40265-022-01724-0 -
Best Practice & Research. Clinical... Nov 2020The role of progesterone goes beyond the maintenance of pregnancy. The hormone, indeed, protects the developing fetal brain and influences its maturation. Metabolomes... (Review)
Review
The role of progesterone goes beyond the maintenance of pregnancy. The hormone, indeed, protects the developing fetal brain and influences its maturation. Metabolomes analyzed by mass spectrometric methods have revealed the great diversity of steroids in maternal plasma and fetal fluids, but their developmental significance remains to be investigated. Progesterone and its metabolites reach highest levels during the third trimester, when the brain growth spurt occurs: its volume triples, synaptogenesis is particularly active, and axons start to be myelinated. This developmental stage coincides with a period of great vulnerability. Studies in sheep have shown that progesterone and its metabolite allopregnanolone protect the vulnerable fetal brain. Work in rats and mice have demonstrated that progesterone plays an important role in myelin formation. These experimental studies are discussed in relation to preterm birth. Influences of progesterone on very early stages of neural development at the beginning of pregnancy are yet to be explored.
Topics: Animals; Female; Fetus; Humans; Infant, Newborn; Mice; Neuroprotection; Pregnancy; Pregnanolone; Premature Birth; Progesterone; Rats; Sheep
PubMed: 33039311
DOI: 10.1016/j.bpobgyn.2020.09.001 -
Dialogues in Clinical Neuroscience Dec 2023Postpartum depression has deleterious effects on childbearing persons globally. Existing treatments have been largely extrapolated from those for other forms of... (Review)
Review
Postpartum depression has deleterious effects on childbearing persons globally. Existing treatments have been largely extrapolated from those for other forms of depression and have included pharmacotherapy, psychotherapy, and neuromodulation. Hormonal treatments with oestrogen and progestogens, thought to be a rational approach to treatment in response to an emerging literature on the pathophysiology of postpartum depression, have only limited evidence for efficacy to date. Novel antidepressant development with allopregnanolone analogues, in contrast, has proven a promising avenue for the development of rationally designed and efficacious treatments. This state-of-the-art review presents the evidence for the current standard-of-care pharmacotherapy, hormonal treatment, and emerging allopregnanolone analogues for the treatment of postpartum depression along with a discussion of the current understanding of its neuroactive steroid-driven pathophysiology.
Topics: Female; Humans; Depression, Postpartum; Pregnanolone; Antidepressive Agents; Psychotherapy
PubMed: 37796239
DOI: 10.1080/19585969.2023.2262464 -
American Journal of Health-system... Feb 2020Postpartum depression (PPD) is defined as a major depressive episode occurring during pregnancy or within 4 weeks of delivery that may have significant consequences for... (Review)
Review
PURPOSE
Postpartum depression (PPD) is defined as a major depressive episode occurring during pregnancy or within 4 weeks of delivery that may have significant consequences for mother and infant. Antidepressants are used to treat PPD, but their effectiveness may be limited by a slow time to peak effect. Brexanolone is Food and Drug Administration-approved for the management of PPD; its use requires patient participation in a risk evaluation and mitigation strategies (REMS) program. This review evaluates the efficacy and safety of brexanolone in PPD.
SUMMARY
Four completed studies, 1 quasi-experimental study and 3 randomized controlled trials (RCTs), were reviewed. Females who had moderate or severe PPD during the third trimester or within 4 weeks of delivery and were less than 6 months postpartum at initiation of therapy were included. Improvement in Hamilton Rating Scale for Depression (HAM-D) scores was assessed in addition to safety outcomes and scores on other depression rating scales. All studies demonstrated statistical improvement in HAM-D scores from baseline with brexanolone vs placebo use at the end of infusions (ie, hour 60). Results with regard to sustained HAM-D score improvements were mixed in the RCTs at 30-day follow-up. The most frequent adverse events in brexanolone-treated patients were sedation, dizziness, somnolence, and headache. The severe or serious adverse effect of presyncope, syncope, or loss of consciousness was reported by 4% of participants.
CONCLUSION
With a rapid onset of action, brexanolone could be considered advantageous over traditional therapies for PPD in patients for whom a rapid response is required due to severity of disease. Significant concerns remain regarding sustained effect and use in patients outside of the clinical trial setting.
Topics: Antidepressive Agents; Depression, Postpartum; Drug Combinations; Female; Humans; Pregnancy; Pregnanolone; Psychiatric Status Rating Scales; Randomized Controlled Trials as Topic; Risk Evaluation and Mitigation; Severity of Illness Index; beta-Cyclodextrins
PubMed: 32073124
DOI: 10.1093/ajhp/zxz333 -
The Lancet. Psychiatry Dec 2019
Topics: Drug Combinations; Ketamine; Pregnanolone; United States; United States Food and Drug Administration; beta-Cyclodextrins
PubMed: 31777341
DOI: 10.1016/S2215-0366(19)30445-6 -
Frontiers in Psychiatry 2023Benzodiazepines (BZs) are prescribed as anxiolytics, but their use is limited by side effects including abuse liability and daytime drowsiness. Neuroactive steroids are...
INTRODUCTION
Benzodiazepines (BZs) are prescribed as anxiolytics, but their use is limited by side effects including abuse liability and daytime drowsiness. Neuroactive steroids are compounds that, like BZs, modulate the effects of GABA at the GABA receptor. In a previous study, combinations of the BZ triazolam and neuroactive steroid pregnanolone produced supra-additive (i.e., greater than expected effects based on the drugs alone) anxiolytic effects but infra-additive (i.e., lower than expected effects based on the drugs alone) reinforcing effects in male rhesus monkeys, suggestive of an improved therapeutic window.
METHODS
Female rhesus monkeys (=4) self-administered triazolam, pregnanolone, and triazolam-pregnanolone combinations intravenously under a progressive-ratio schedule. In order to assess characteristic sedative-motor effects of BZ-neuroactive steroid combinations, female rhesus monkeys (n=4) were administered triazolam, pregnanolone, and triazolam-pregnanolone combinations. Trained observers, blinded to condition, scored the occurrence of species-typical and drug-induced behaviors.
RESULTS
In contrast to our previous study with males, triazolam-pregnanolone combinations had primarily supra-additive reinforcing effects in three monkeys but infra-additive reinforcing effects in one monkey. Scores for deep sedation (i.e., defined as atypical loose-limbed posture, eyes closed, does not respond to external stimuli) and observable ataxia (any slip, trip, fall, or loss of balance) were significantly increased by both triazolam and pregnanolone. When combined, triazolam-pregnanolone combinations had supra-additive effects for inducing deep sedation, whereas observable ataxia was attenuated, likely due to the occurrence of robust sedative effects.
DISCUSSION
These results suggest that significant sex differences exist in self-administration of BZ-neuroactive steroid combinations, with females likely to show enhanced sensitivity to reinforcing effects compared with males. Moreover, supra-additive sedative effects occurred for females, demonstrating a higher likelihood of this adverse effect when these drug classes are combined.
PubMed: 37252149
DOI: 10.3389/fpsyt.2023.1142531 -
Journal of Internal Medicine Sep 2023The prevalence of cognitive dysfunction, dementia, and neurodegenerative disorders such as Alzheimer's disease (AD) is increasing in parallel with an aging population.... (Review)
Review
The prevalence of cognitive dysfunction, dementia, and neurodegenerative disorders such as Alzheimer's disease (AD) is increasing in parallel with an aging population. Distinct types of chronic stress are thought to be instrumental in the development of cognitive impairment in central nervous system (CNS) disorders where cognitive impairment is a major unmet medical need. Increased GABAergic tone is a mediator of stress effects but is also a result of other factors in CNS disorders. Positive GABA-A receptor modulating stress and sex steroids (steroid-PAMs) such as allopregnanolone (ALLO) and medroxyprogesterone acetate can provoke impaired cognition. As such, ALLO impairs memory and learning in both animals and humans. In transgenic AD animal studies, continuous exposure to ALLO at physiological levels impairs cognition and increases degenerative AD pathology, whereas intermittent ALLO injections enhance cognition, indicating pleiotropic functions of ALLO. We have shown that GABA-A receptor modulating steroid antagonists (GAMSAs) can block the acute negative cognitive impairment of ALLO on memory in animal studies and in patients with cognitive impairment due to hepatic encephalopathy. Here we describe disorders affected by steroid-PAMs and opportunities to treat these adverse effects of steroid-PAMs with novel GAMSAs.
Topics: Animals; Humans; Aged; Receptors, GABA-A; Neurosteroids; Cognitive Dysfunction; Pregnanolone; Alzheimer Disease; gamma-Aminobutyric Acid
PubMed: 37518841
DOI: 10.1111/joim.13705