Translational Pharmacology of PRAX-944, a Novel T-Type Calcium Channel Blocker in Development for the Treatment of Essential Tremor.

BACKGROUND: Essential tremor is the most common movement disorder with clear unmet need. Mounting evidence indicates tremor is caused by increased neuronal burst firing and oscillations in cerebello-thalamo-cortical circuitry and may be dependent on T-type calcium channel activity. T-type calcium channels regulate sigma band electroencephalogram (EEG) power during non-rapid eye movement sleep, representing a potential biomarker of channel activity. PRAX-944 is a novel T-type calcium channel blocker in development for essential tremor. OBJECTIVES: Using a rat tremor model and sigma-band EEG power, we assessed pharmacodynamically-active doses of PRAX-944 and their translation into clinically tolerated doses in healthy participants, informing dose selection for future efficacy trials. METHODS: Harmaline-induced tremor and spontaneous locomotor activity were used to assess PRAX-944 efficacy and tolerability, respectively, in rats. Sigma-power was used as a translational biomarker of T-type calcium channel blockade in rats and, subsequently, in a phase 1 trial assessing pharmacologic activity and tolerability in healthy participants. RESULTS: In rats, PRAX-944 dose-dependently reduced tremor by 50% and 72% at 1 and 3 mg/kg doses, respectively, without locomotor side effects. These doses also reduced sigma-power by ~30% to 50% in rats. In healthy participants, sigma-power was similarly reduced by 34% to 50% at 10 to 100 mg, with no further reduction at 120 mg. All doses were well tolerated. CONCLUSIONS: In rats, PRAX-944 reduced sigma-power at concentrations that reduced tremor without locomotor side effects. In healthy participants, comparable reductions in sigma-power indicate that robust T-type calcium channel blockade was achieved at well-tolerated doses that may hold promise for reducing tremor in patients with essential tremor. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

The novel persistent sodium current inhibitor PRAX-562 has potent anticonvulsant activity with improved protective index relative to standard of care sodium channel blockers.

OBJECTIVE: This study investigates the effects of PRAX-562 on sodium current (INa ), intrinsic neuronal excitability, and protection from evoked seizures to determine whether a preferential persistent INa inhibitor would exhibit improved preclinical efficacy and tolerability compared to two standard voltage-gated sodium channel (NaV ) blockers. METHODS: Inhibition of INa  was characterized using patch clamp analysis. The effect on intrinsic excitability was measured using evoked action potentials recorded from hippocampal CA1 pyramidal neurons in mouse brain slices. Anticonvulsant activity was evaluated using the maximal electroshock seizure (MES) model, and tolerability was assessed by measuring spontaneous locomotor activity (sLMA). RESULTS: PRAX-562 potently and preferentially inhibited persistent INa induced by ATX-II or the SCN8A mutation N1768D (half-maximal inhibitory concentration [IC50 ] = 141 and 75 nmol·L-1 , respectively) relative to peak INa tonic/resting block (60× preference). PRAX-562 also exhibited potent use-dependent block (31× preference to tonic block). This profile is considerably different from standard NaV blockers, including carbamazepine (CBZ; persistent INa IC50 = 77 500 nmol·L-1 , preference ratios of 30× [tonic block], less use-dependent block observed at various frequencies). In contrast to CBZ, PRAX-562 reduced neuronal intrinsic excitability with only a minor reduction in action potential amplitude. PRAX-562 (10 mg/kg po) completely prevented evoked seizures without affecting sLMA (MES unbound brain half-maximal efficacious concentration = 4.3 nmol·L-1 , sLMA half-maximal tolerated concentration = 69.7 nmol·L-1 , protective index [PI] = 16×). In contrast, CBZ and lamotrigine (LTG) had PIs of approximately 5.5×, with significant overlap between doses that were anticonvulsant and that reduced locomotor activity. SIGNIFICANCE: PRAX-562 demonstrated robust preclinical anticonvulsant activity similar to CBZ but improved compared to LTG. PRAX-562 exhibited significantly improved preclinical tolerability compared with standard NaV blockers (CBZ and LTG), potentially due to the preference for persistent INa . Preferential targeting of persistent INa may represent a differentiated therapeutic option for diseases of hyperexcitability, where standard NaV blockers have demonstrated efficacy but poor tolerability.

Evoked potentials as a translatable biomarker to track functional remyelination.

Enhancing remyelination is a key therapeutic strategy for demyelinating diseases such as multiple sclerosis. To achieve this goal, a central challenge is being able to quantitatively and longitudinally track functional remyelination, especially with translatable biomarkers that can be performed in both preclinical models and in the clinic. We developed the methodology to stably measure multi-modal sensory evoked potentials from the skull surface over the course of months in individual mice and applied it to a genetic mouse model of oligodendrocyte ablation and demyelination. We found that auditory and somatosensory evoked potential latencies reliably increased over time during the early phase of the model and recovered spontaneously and almost completely during a later phase. Histological examination supported the interpretation that the evoked potential latency changes dynamically reflect changes in CNS myelination. Specifically, we found reduction of myelination in corresponding brain regions at the time that sensory evoked potentials were maximally impacted. Importantly, we also found that myelination levels recovered when evoked potential latencies recovered. Other changes known to associate with demyelination were also observed at the time of delayed evoked potentials, including the emergence of white matter vacuoles and increased markers for activated microglia and macrophages; these changes also fully reversed by the time that evoked potentials recovered. Our results support the hypothesis that skull-surface recorded evoked potential latencies can dynamically track CNS myelination changes. The methods developed here allow for longitudinally tracking functional myelination changes in vivo in preclinical rodent models with a quantitative biomarker that can also be applied clinically and will facilitate translational development of CNS remyelinating therapies.

Preclinical to Human Translational Pharmacology of the Novel M1 Positive Allosteric Modulator MK-7622.

The current standard of care for treating Alzheimer's disease is acetylcholinesterase inhibitors, which nonselectively increase cholinergic signaling by indirectly enhancing activity of nicotinic and muscarinic receptors. These drugs improve cognitive function in patients, but also produce unwanted side effects that limit their efficacy. In an effort to selectively improve cognition and avoid the cholinergic side effects associated with the standard of care, various efforts have been aimed at developing selective M1 muscarinic receptor activators. In this work, we describe the preclinical and clinical pharmacodynamic effects of the M1 muscarinic receptor-positive allosteric modulator, MK-7622. MK-7622 attenuated the cognitive-impairing effects of the muscarinic receptor antagonist scopolamine and altered quantitative electroencephalography (qEEG) in both rhesus macaque and human. For both scopolamine reversal and qEEG, the effective exposures were similar between species. However, across species the minimum effective exposures to attenuate the scopolamine impairment were lower than for qEEG. Additionally, there were differences in the spectral power changes produced by MK-7622 in rhesus versus human. In sum, these results are the first to demonstrate translation of preclinical cognition and target modulation to clinical effects in humans for a selective M1 muscarinic receptor-positive allosteric modulator.

Tau Antibody Targeting Pathological Species Blocks Neuronal Uptake and Interneuron Propagation of Tau in Vitro.

The clinical progression of Alzheimer disease (AD) is associated with the accumulation of tau neurofibrillary tangles, which may spread throughout the cortex by interneuronal tau transfer. If so, targeting extracellular tau species may slow the spreading of tau pathology and possibly cognitive decline. To identify suitable target epitopes, we tested the effects of a panel of tau antibodies on neuronal uptake and aggregation in vitro. Immunodepletion was performed on brain extract from tau-transgenic mice and postmortem AD brain and added to a sensitive fluorescence resonance energy transfer-based tau uptake assay to assess blocking efficacy. The antibodies reduced tau uptake in an epitope-dependent manner: N-terminal (Tau13) and middomain (6C5 and HT7) antibodies successfully prevented uptake of tau species, whereas the distal C-terminal-specific antibody (Tau46) had little effect. Phosphorylation-dependent (40E8 and p396) and C-terminal half (4E4) tau antibodies also reduced tau uptake despite removing less total tau by immunodepletion, suggesting specific interactions with species involved in uptake. Among the seven antibodies evaluated, 6C5 most efficiently blocked uptake and subsequent aggregation. More important, 6C5 also blocked neuron-to-neuron spreading of tau in a unique three-chamber microfluidic device. Furthermore, 6C5 slowed down the progression of tau aggregation even after uptake had begun. Our results imply that not all antibodies/epitopes are equally robust in terms of blocking tau uptake of human AD-derived tau species.

Tau: From research to clinical development.

Alzheimer's Association Research Roundtable Fall 2015-Tau: From research to clinical development. Tau pathology is recognized as the key driver of disease progression in Alzheimer's and other neurodegenerative diseases. Although this makes tau an attractive target for the development of novel diagnostic and therapeutic strategies, the mechanisms underlying the onset and progression of tau-related neurotoxicity remain elusive. Recent strides in the development of sophisticated preclinical models and the emergence of tau PET imaging and fluid biomarkers provide new opportunities to increase our understanding of tau biology, overcome translational challenges, and accelerate the advancement of tau therapeutics from bench to bedside. With this in mind, the Alzheimer's Association convened a Research Roundtable in October 2015, bringing together experts from academia, industry, and regulatory agencies to discuss the latest understanding of tau pathogenic pathways and review the evolution of tau therapeutics and biomarkers currently in development. The meeting provided a forum to share experience and expertise with the common goal of advancing the discovery and development of new treatment strategies and expediting the design and implementation of efficient clinical trials.

Use of structured expert judgment to forecast invasions by bighead and silver carp in Lake Erie.

Identifying which nonindigenous species will become invasive and forecasting the damage they will cause is difficult and presents a significant problem for natural resource management. Often, the data or resources necessary for ecological risk assessment are incomplete or absent, leaving environmental decision makers ill equipped to effectively manage valuable natural resources. Structured expert judgment (SEJ) is a mathematical and performance-based method of eliciting, weighting, and aggregating expert judgments. In contrast to other methods of eliciting and aggregating expert judgments (where, for example, equal weights may be assigned to experts), SEJ weights each expert on the basis of his or her statistical accuracy and informativeness through performance measurement on a set of calibration variables. We used SEJ to forecast impacts of nonindigenous Asian carp (Hypophthalmichthys spp.) in Lake Erie, where it is believed not to be established. Experts quantified Asian carp biomass, production, and consumption and their impact on 4 fish species if Asian carp were to become established. According to experts, in Lake Erie Asian carp have the potential to achieve biomass levels that are similar to the sum of biomasses for several fishes that are harvested commercially or recreationally. However, the impact of Asian carp on the biomass of these fishes was estimated by experts to be small, relative to long term average biomasses, with little uncertainty. Impacts of Asian carp in tributaries and on recreational activities, water quality, or other species were not addressed. SEJ can be used to quantify key uncertainties of invasion biology and also provide a decision-support tool when the necessary information for natural resource management and policy is not available.

Identification of a methoxynaphthalene scaffold as a core replacement in quinolizidinone amide M(1) positive allosteric modulators.

A series of methoxynaphthalene amides were prepared and evaluated as alternatives to quinolizidinone amide M1 positive allosteric modulators. A methoxy group was optimal for M1 activity and addressed key P-gp issues present in the aforementioned quinolizidinone amide series.

Using structured expert judgment to assess invasive species prevention: Asian carp and the Mississippi-Great Lakes hydrologic connection.

Recently, authors have theorized that invasive species prevention is more cost-effective than control in protecting ecosystem services. However, quantification of the effectiveness of prevention is rare because experiments at field scales are expensive or infeasible. We therefore used structured expert judgment to quantify the efficacy of 17 proposed strategies to prevent Asian carp invasion of the Laurentian Great Lakes via the hydrologic connection between the Mississippi and Great Lakes watersheds. Performance-weighted expert estimates indicated that hydrologic separation would prevent 99% (95,100; median, 5th and 95th percentiles) of Asian carp access, while electric and acoustic-bubble-strobe barriers would prevent 92% (85,95) and 92% (75,95), respectively. For all other strategies, estimated effectiveness was lower, with greater uncertainty. When potential invasions by other taxa are considered, the effectiveness of hydrologic separation increases relative to strategies that are effective primarily for fishes. These results could help guide invasive species management in many waterways globally.

A 3D individual-based aquatic transport model for the assessment of the potential dispersal of planktonic larvae of an invasive bivalve.

The unwanted impacts of non-indigenous species have become one of the major ecological and economic threats to aquatic ecosystems worldwide. Assessing the potential dispersal and colonization of non-indigenous species is necessary to prevent or reduce deleterious effects that may lead to ecosystem degradation and a range of economic impacts. A three dimensional (3D) numerical model has been developed to evaluate the local dispersal of the planktonic larvae of an invasive bivalve, Asian clam (Corbicula fluminea), by passive hydraulic transport in Lake Tahoe, USA. The probability of dispersal of Asian clam larvae from the existing high density populations to novel habitats is determined by the magnitude and timing of strong wind events. The probability of colonization of new near-shore areas outside the existing beds is low, but sensitive to the larvae settling velocity ws. High larvae mortality was observed due to settling in unsuitable deep habitats. The impact of UV-radiation during the pelagic stages, on the Asian clam mortality was low. This work provides a quantification of the number of propagules that may be successfully transported as a result of natural processes and in function of population size. The knowledge and understanding of the relative contribution of different dispersal pathways, may directly inform decision-making and resource allocation associated with invasive species management.

Selective activation of the M1 muscarinic acetylcholine receptor achieved by allosteric potentiation.

The forebrain cholinergic system promotes higher brain function in part by signaling through the M(1) muscarinic acetylcholine receptor (mAChR). During Alzheimer's disease (AD), these cholinergic neurons degenerate, therefore selectively activating M(1) receptors could improve cognitive function in these patients while avoiding unwanted peripheral responses associated with non-selective muscarinic agonists. We describe here benzyl quinolone carboxylic acid (BQCA), a highly selective allosteric potentiator of the M(1) mAChR. BQCA reduces the concentration of ACh required to activate M(1) up to 129-fold with an inflection point value of 845 nM. No potentiation, agonism, or antagonism activity on other mAChRs is observed up to 100 microM. Furthermore studies in M(1)(-/-) mice demonstrates that BQCA requires M(1) to promote inositol phosphate turnover in primary neurons and to increase c-fos and arc RNA expression and ERK phosphorylation in the brain. Radioligand-binding assays, molecular modeling, and site-directed mutagenesis experiments indicate that BQCA acts at an allosteric site involving residues Y179 and W400. BQCA reverses scopolamine-induced memory deficits in contextual fear conditioning, increases blood flow to the cerebral cortex, and increases wakefulness while reducing delta sleep. In contrast to M(1) allosteric agonists, which do not improve memory in scopolamine-challenged mice in contextual fear conditioning, BQCA induces beta-arrestin recruitment to M(1), suggesting a role for this signal transduction mechanism in the cholinergic modulation of memory. In summary, BQCA exploits an allosteric potentiation mechanism to provide selectivity for the M(1) receptor and represents a promising therapeutic strategy for cognitive disorders.

The control of an invasive bivalve, Corbicula fluminea, using gas impermeable benthic barriers in a large natural lake.

Anoxia can restrict species establishment in aquatic systems and the artificial promotion of these conditions can provide an effective control strategy for invasive molluscs. Low abundances (2-20 m(-2)) of the nonnative bivalve, Asian clam (Corbicula fluminea), were first recorded in Lake Tahoe, CA-NV in 2002 and by 2010 nuisance-level population densities (>10,000 m(-2)) were observed. A non-chemical control method using gas impermeable benthic barriers to reduce dissolved oxygen (DO) concentrations available to C. fluminea was tested in this ultra-oligotrophic natural lake. In 2009, the impact of ethylene propylene diene monomer (EPDM) sheets (9 m(2), n = 6) on C. fluminea beds was tested on 1-7 day intervals over a 56 day period (August-September). At an average water temperature of 18 °C, DO concentrations under these small barriers were reduced to zero after 72 h resulting in 100 % C. fluminea mortality after 28 days. In 2010, a large EPDM barrier (1,950 m(2)) was applied to C. fluminea populations for 120 days (July-November). C. fluminea abundances were reduced over 98 % after barrier removal, and remained significantly reduced (>90 %) 1 year later. Non-target benthic macroinvertebrate abundances were also reduced, with variable taxon-specific recolonization rates. High C. fluminea abundance under anoxic conditions increased the release of ammonium and soluble reactive phosphorus from the sediment substrate; but levels of unionized ammonia were low at 0.004-0.005 mg L(-1). Prolonged exposure to anoxia using benthic barriers can provide an effective short term control strategy for C. fluminea.

Fused heterocyclic M1 positive allosteric modulators.

Fused aromatics such as naphthalene were identified as highly potent and CNS penetrant M(1) positive allosteric modulators during an SAR study to replace the phenyl B-ring linkage.

Quinolizidinone carboxylic acid selective M1 allosteric modulators: SAR in the piperidine series.

SAR study of the piperidine moiety in a series of quinolizidinone carboxylic acid M(1) positive allosteric modulators was examined. While the SAR was generally flat, compounds were identified with high CNS exposure to warrant additional in vivo evaluation.

Discovery of the First Orally Available, Selective KNa1.1 Inhibitor: In Vitro and In Vivo Activity of an Oxadiazole Series.

The gene KCNT1 encodes the sodium-activated potassium channel KNa1.1 (Slack, Slo2.2). Variants in the KCNT1 gene induce a gain-of-function (GoF) phenotype in ionic currents and cause a spectrum of intractable neurological disorders in infants and children, including epilepsy of infancy with migrating focal seizures (EIMFS) and autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE). Effective treatment options for KCNT1-related disease are absent, and novel therapies are urgently required. We describe the development of a novel class of oxadiazole KNa1.1 inhibitors, leading to the discovery of compound 31 that reduced seizures and interictal spikes in a mouse model of KCNT1 GoF.

A stochastic model for annual reproductive success.

Demographic stochasticity can have large effects on the dynamics of small populations as well as on the persistence of rare genotypes and lineages. Survival is sensibly modeled as a binomial process, but annual reproductive success (ARS) is more complex and general models for demographic stochasticity do not exist. Here we introduce a stochastic model framework for ARS and illustrate some of its properties. We model a sequence of stochastic events: nest completion, the number of eggs or neonates produced, nest predation, and the survival of individual offspring to independence. We also allow multiple nesting attempts within a breeding season. Most of these components can be described by Bernoulli or binomial processes; the exception is the distribution of offspring number. Using clutch and litter size distributions from 53 vertebrate species, we demonstrate that among-individual variability in offspring number can usually be described by the generalized Poisson distribution. Our model framework allows the demographic variance to be calculated from underlying biological processes and can easily be linked to models of environmental stochasticity or selection because of its parametric structure. In addition, it reveals that the distributions of ARS are often multimodal and skewed, with implications for extinction risk and evolution in small populations.

Heterocyclic fused pyridone carboxylic acid M(1) positive allosteric modulators.

The phenyl ring in a series of quinolone carboxylic acid M(1) positive allosteric modulators was replaced with a variety of heterocycles in order to reduce protein plasma binding and enhance CNS exposure.

Hydroxy cycloalkyl fused pyridone carboxylic acid M(1) positive allosteric modulators.

Incorporation of hydroxycycloalkyl fused pyridone carboxylic acids in lieu of quinolone carboxylic acids enhance free fraction without increased susceptibility to P-glycoprotein transport.

N-heterocyclic derived M1 positive allosteric modulators.

Replacement of a phenyl ring with N-linked heterocycles in a series of quinolone carboxylic acid M1 positive allosteric modulators was investigated. In particular, a pyrazole derivative exhibited improvements in potency, free fraction, and CNS exposure.

Parallel synthesis of N-biaryl quinolone carboxylic acids as selective M(1) positive allosteric modulators.

An iterative analog library synthesis approach was employed in the exploration of a quinolone carboxylic acid series of selective M(1) positive allosteric modulators, and strategies for improving potency and plasma free fraction were identified.