Clinical targeting of HIV capsid protein with a long-acting small molecule.

Oral antiretroviral agents provide life-saving treatments for millions of people living with HIV, and can prevent new infections via pre-exposure prophylaxis1-5. However, some people living with HIV who are heavily treatment-experienced have limited or no treatment options, owing to multidrug resistance6. In addition, suboptimal adherence to oral daily regimens can negatively affect the outcome of treatment-which contributes to virologic failure, resistance generation and viral transmission-as well as of pre-exposure prophylaxis, leading to new infections1,2,4,7-9. Long-acting agents from new antiretroviral classes can provide much-needed treatment options for people living with HIV who are heavily treatment-experienced, and additionally can improve adherence10. Here we describe GS-6207, a small molecule that disrupts the functions of HIV capsid protein and is amenable to long-acting therapy owing to its high potency, low in vivo systemic clearance and slow release kinetics from the subcutaneous injection site. Drawing on X-ray crystallographic information, we designed GS-6207 to bind tightly at a conserved interface between capsid protein monomers, where it interferes with capsid-protein-mediated interactions between proteins that are essential for multiple phases of the viral replication cycle. GS-6207 exhibits antiviral activity at picomolar concentrations against all subtypes of HIV-1 that we tested, and shows high synergy and no cross-resistance with approved antiretroviral drugs. In phase-1 clinical studies, monotherapy with a single subcutaneous dose of GS-6207 (450 mg) resulted in a mean log10-transformed reduction of plasma viral load of 2.2 after 9 days, and showed sustained plasma exposure at antivirally active concentrations for more than 6 months. These results provide clinical validation for therapies that target the functions of HIV capsid protein, and demonstrate the potential of GS-6207 as a long-acting agent to treat or prevent infection with HIV.

A highly potent long-acting small-molecule HIV-1 capsid inhibitor with efficacy in a humanized mouse model.

People living with HIV (PLWH) have expressed concern about the life-long burden and stigma associated with taking pills daily and can experience medication fatigue that might lead to suboptimal treatment adherence and the emergence of drug-resistant viral variants, thereby limiting future treatment options1-3. As such, there is strong interest in long-acting antiretroviral (ARV) agents that can be administered less frequently4. Herein, we report GS-CA1, a new archetypal small-molecule HIV capsid inhibitor with exceptional potency against HIV-2 and all major HIV-1 types, including viral variants resistant to the ARVs currently in clinical use. Mechanism-of-action studies indicate that GS-CA1 binds directly to the HIV-1 capsid and interferes with capsid-mediated nuclear import of viral DNA, HIV particle production and ordered capsid assembly. GS-CA1 selects in vitro for unfit GS-CA1-resistant capsid variants that remain fully susceptible to other classes of ARVs. Its high metabolic stability and low solubility enabled sustained drug release in mice following a single subcutaneous dosing. GS-CA1 showed high antiviral efficacy as a long-acting injectable monotherapy in a humanized mouse model of HIV-1 infection, outperforming long-acting rilpivirine. Collectively, these results demonstrate the potential of ultrapotent capsid inhibitors as new long-acting agents for the treatment of HIV-1 infection.

Modeling Nonlinear Synaptic Dynamics: A Laguerre-Volterra Network Framework for Improved Computational Efficiency in Large Scale Simulations.

Synapses are key components in signal transmission in the brain, often exhibiting complex non-linear dynamics. Yet, they are often crudely modelled as linear exponential equations in large-scale neuron network simulations. Mechanistic models that use detailed channel receptor kinetics more closely replicate the nonlinear dynamics observed at synapses, but use of such models are generally restricted to small scale simulations due to their computational complexity. Previously, we have developed an ``input-output'' (IO) synapse model using the Volterra functional series to estimate nonlinear synaptic dynamics. Here, we present an improvement on the IO synapse model using the extbf{Laguerre-Volterra network (LVN) framework. We demonstrate that utilization of the LVN framework helps reduce memory requirements and improves the simulation speed in comparison to the previous iteration of the IO synapse model. We present results that demonstrate the accuracy, memory efficiency, and speed of the LVN model that can be extended to simulations with large numbers of synapses. Our efforts enable complex nonlinear synaptic dynamics to be modelled in large-scale network models, allowing us to explore how synaptic activity may influence network behavior and affects memory, learning, and neurodegenerative diseases.

Development of a detailed model of calcium dynamics at the postsynaptic spine of an excitatory synapse.

Postsynaptic calcium dynamics play a critical role in synaptic plasticity, but are often difficult to measure in experimental protocols due to their relatively fast rise and decay times, and the small spine dimensions. To circumvent these limitations, we propose to develop a computational model of calcium dynamics in the postsynaptic spine. This model integrates the main elements that participate in calcium concentration influx, efflux, diffusion and buffering. These consist of (i) spine geometry; (ii) calcium influx through NMDA receptors and voltage-dependent calcium channels (VDCC); (iii) calcium efflux with plasma membrane calcium pumps (PMCA) and sodium-calcium exchangers (NCX); (iv) intracellular calcium stores; and (v) calcium buffers. We herein present computational results we obtained and compare them with experimentally measured data, thereby validating the proposed model. Overall the development of such postsynaptic calcium model may help us better understand the intricacies of interplay between the different elements that shape calcium dynamics and impact synaptic plasticity in normal functions and pathologies. This model also constitutes a first step in the development of a nonlinear input-output calcium dynamics model for multi-scale, large scale neuronal simulations.

Volterra representation enables modeling of complex synaptic nonlinear dynamics in large-scale simulations.

Chemical synapses are comprised of a wide collection of intricate signaling pathways involving complex dynamics. These mechanisms are often reduced to simple spikes or exponential representations in order to enable computer simulations at higher spatial levels of complexity. However, these representations cannot capture important nonlinear dynamics found in synaptic transmission. Here, we propose an input-output (IO) synapse model capable of generating complex nonlinear dynamics while maintaining low computational complexity. This IO synapse model is an extension of a detailed mechanistic glutamatergic synapse model capable of capturing the input-output relationships of the mechanistic model using the Volterra functional power series. We demonstrate that the IO synapse model is able to successfully track the nonlinear dynamics of the synapse up to the third order with high accuracy. We also evaluate the accuracy of the IO synapse model at different input frequencies and compared its performance with that of kinetic models in compartmental neuron models. Our results demonstrate that the IO synapse model is capable of efficiently replicating complex nonlinear dynamics that were represented in the original mechanistic model and provide a method to replicate complex and diverse synaptic transmission within neuron network simulations.

Synaptic Efficacy as a Function of Ionotropic Receptor Distribution: A Computational Study.

Glutamatergic synapses are the most prevalent functional elements of information processing in the brain. Changes in pre-synaptic activity and in the function of various post-synaptic elements contribute to generate a large variety of synaptic responses. Previous studies have explored postsynaptic factors responsible for regulating synaptic strength variations, but have given far less importance to synaptic geometry, and more specifically to the subcellular distribution of ionotropic receptors. We analyzed the functional effects resulting from changing the subsynaptic localization of ionotropic receptors by using a hippocampal synaptic computational framework. The present study was performed using the EONS (Elementary Objects of the Nervous System) synaptic modeling platform, which was specifically developed to explore the roles of subsynaptic elements as well as their interactions, and that of synaptic geometry. More specifically, we determined the effects of changing the localization of ionotropic receptors relative to the presynaptic glutamate release site, on synaptic efficacy and its variations following single pulse and paired-pulse stimulation protocols. The results indicate that changes in synaptic geometry do have consequences on synaptic efficacy and its dynamics.

The volterra functional series is a viable alternative to kinetic models for synaptic modeling--calibration and benchmarking.

Synaptic transmission is governed by a series of complex and highly nonlinear mechanisms and pathways in which the dynamics have a profound influence on the overall signal sent to the postsynaptic cell. In simulation, these mechanisms are often represented through kinetic models governed by state variables and rate law equations. Calculations of such ordinary differential equations (ODEs) in kinetic models can be computationally intensive, and although algorithms have been optimally developed to handle ODEs efficiently, simulation of numerous, large and complex kinetic models requires a prohibitively large amount of computational power. Here we present an alternative representation of ionotropic glutamatergic receptors AMPAr and NMDAr kinetic models consisting of input-output surrogates of the receptor models which can capture the nonlinear dynamics seen in the kinetic models. We benchmark this Input-Output (IO) synapse model and compare it with kinetic receptor models to evaluate the simulation time required when using either synapse model, as well as the number of time steps each model needs for simulation. While remaining faithful to the original dynamics of the model, our results indicate that the IO synapse model requires less simulation time than the kinetic models under conditions which elicit normal physiological responses, thereby improving computational efficiency while preserving the complex non-linear dynamics of the receptors. These IO surrogates therefore constitute an appealing alternative to kinetic models in large scale networks simulations.

A comparison between direct and indirect measurements of neurotransmitter vesicle release dynamics: a computational study.

Presynaptic vesicular release of neurotransmitters is a stochastic process involving complex mechanisms triggered by an elevation of calcium concentration. The mechanisms behind neurotransmitters release play a critical role in synaptic function and plasticity. Understanding its properties, both in term of its dynamics and its underlying mechanisms, may therefore help further our understanding of synaptic plasticity. However, measuring vesicle release dynamics is experimentally challenging. One experimental protocol used to determine the dynamic properties of vesicle release is to measure postsynaptic current. However, this method inherently not only captures properties of the release itself, but also the contributions from the postsynaptic receptors. Here we propose to use a synapse simulation platform known as EONS/RHENOMS to capture the functional properties of vesicle release, separate from the dynamics known to be associated with postsynaptic receptors, and compare the results with those determined experimentally. We find that despite attempts to reduce interference of postsynaptic dynamics, the receptor channel properties, particularly desensitization, may influence the overall measured results significantly. Re-estimating release rate by taking into account the contributions of postsynaptic receptors may give further insight into release dynamics and further our overall understanding on synaptic plasticity.

Modeling of the nervous system: from modulation of glutamatergic and gabaergic molecular dynamics to neuron spiking activity.

One of the fundamental characteristics of the brain is its hierarchical and temporal organization: scales in both space and time must be considered to fully grasp the system's underlying mechanisms and their impact on brain function. Complex interactions taking place at the molecular level regulate neuronal activity that further modifies the function of millions of neurons connected by trillions of synapses, ultimately giving rise to complex function and behavior at the system level. Likewise, the spatial complexity is accompanied by a complex temporal integration of events taking place at the microsecond scale leading to slower changes occurring at the second, minute and hour scales. These integrations across hierarchies of the nervous system are sufficiently complex to have impeded the development of routine multi-level modeling methodologies. The present study describes an example of our multiscale efforts to rise from the biomolecular level to the neuron level. We more specifically describe how we integrate biomolecular mechanisms taking place at glutamatergic and gabaergic synapses and integrate them to study the impact of these modifications on spiking activity of a CA1 pyramidal cell in the hippocampus.

Integrated multiscale modeling of the nervous system: predicting changes in hippocampal network activity by a positive AMPA receptor modulator.

One of the fundamental characteristics of the brain is its hierarchical organization. Scales in both space and time that must be considered when integrating across hierarchies of the nervous system are sufficiently great as to have impeded the development of routine multilevel modeling methodologies. Complex molecular interactions at the level of receptors and channels regulate activity at the level of neurons; interactions between multiple populations of neurons ultimately give rise to complex neural systems function and behavior. This spatial complexity takes place in the context of a composite temporal integration of multiple, different events unfolding at the millisecond, second, minute, hour, and longer time scales. In this study, we present a multiscale modeling methodology that integrates synaptic models into single neuron, and multineuron, network models. We have applied this approach to the specific problem of how changes at the level of kinetic parameters of a receptor-channel model are translated into changes in the temporal firing pattern of a single neuron, and ultimately, changes in the spatiotemporal activity of a network of neurons. These results demonstrate how this powerful methodology can be applied to understand the effects of a given local process within multiple hierarchical levels of the nervous system.

Modeling of the nervous system: from molecular dynamics and synaptic modulation to neuron spiking activity.

The brain is a perfect example of an integrated multi-scale system, as the complex interactions taking place at the molecular level regulate neuronal activity that further modifies the function of millions of neurons connected by trillions of synapses, ultimately giving rise to complex function and behavior at the system level. Likewise, the spatial complexity is accompanied by a complex temporal integration of events taking place at the microsecond scale leading to slower changes occurring at the second, minute and hour scales. In the present study we illustrate our approach to model and simulate the spatio-temporal complexity of the nervous system by developing a multi-scale model integrating synaptic models into the neuronal and ultimately network levels. We apply this approach to a concrete example and demonstrate how changes at the level of kinetic parameters of a receptor model are translated into significant changes in the firing of a pyramidal neuron. These results illustrate the abilities of our modeling approach and support its direct application to the evaluation of the effects of drugs, from functional target to integrated system.

Paired-pulse stimulation at glutamatergic synapses - pre- and postsynaptic components.

Paired-pulse stimulation is a standard protocol that has been used for decades to characterize dynamic systems: the differences in responses to two sequential identical stimuli as a function of inter-stimulus interval provide quantitative information on the dynamics of the system. In neuroscience, the paired-pulse protocol is also widely used at multiple levels of analysis, from behavioral conditioning to synaptic plasticity, and in particular to define the biomolecular mechanism of learning and memory. In a system as small and complex as synapses, it is extremely challenging - if not impossible - to experimentally gain access to the multitude of possible readouts. In the present study, we first introduce a computational synaptic modeling platform that we developed and calibrated based on experimental data from both our laboratories and a variety of publications. We then show how this platform allows not only to replicate experimental data, but also to go beyond technological boundaries and investigate the main parameters responsible for regulation of synaptic transmission and plasticity. The results provide critical information regarding the respective role of various subsynaptic processes and of their interactions. Additionally, this approach can strengthen our understanding of potential dysfunctions (pathologies) and suggest potential approaches to re-establish normal function.