A Therapeutic Strategy for Treatment-Resistant Status Epilepticus

As early as secondary school, pupils are taught that particles, such as ions and molecules, passively diffuse from areas of high concentration to areas of low concentration. They are also taught that opposite electric charges attract each other. These two concepts, when combined in a neuroscientific context, form the basis of the electrochemical gradient, which is of significance in relation to neuronal activity. An understanding of the biophysics of the neuron will aid in understanding the topic of the current article: in the field of preclinical research, a newly-developed compound, OV350, was used by Jarvis and colleagues (2023) to treat benzodiazepine-resistant status epilepticus (SE) in mice. The present article will provide an overview of neuronal electrophysiology, before divining into SE and the rationale behind OV350 as a potential therapeutic strategy for this disorder.

Electrical Activity and the Neuron

At resting state, a neuron is more negatively charged on the inside than it is on the outside. This has been known from voltage measurements carried out by Hodgkin and Huxley (1945) on the axon of the Loligo forbesii squid (Figure 1). For a neuron to be active or fire, the neuronal membrane must be depolarized. Depolarization occurs, for example, when positively-charged ions, such as Na+, flow into the cell. Instead, a neuronal membrane is hyperpolarized when the membrane potential (or voltage) becomes more negative. This happens when negatively-charged ions, such as Cl­-, flow into the cell, among other events. The neuronal membrane itself is a barrier to ionic flow: ions flow across membranes through leak ion channels, voltage-gated ion channels, and neurotransmitter-gated ion channels, among other structures.

Figure 1: Loligo forbesii. The axon of this squid contributed to our current understanding of the biophysical properties of the neuronal membrane (Merculiano, 1896, Table 8).

Ionic flow across the neuronal membrane is influenced by the electrochemical driving force. This force is, in turn, influenced by two factors, which are concentration gradients and membrane potential: ions will passively diffuse from an environment where they are at high concentration, to an environment where they are at low concentration; positively-charged ions will flow to a negatively-charged environment, and vice versa. Neurons possess homeostatic mechanisms, e.g., transmembranous protein transporters, in place which regulate intraneuronal ionic concentrations, the values of which influence neuronal activity. Certain disorders are characterized by pathological neuronal activity; a common example of such a disorder is epilepsy.

Status Epilepticus

Epilepsy is a disorder characterized by recurrent seizures. According to the International League Against Epilepsy, “an epileptic seizure is a transient occurrence of signs and/or symptoms due to abnormal excessive or synchronous neuronal activity in the brain” (Fisher et al., 2005, p. 471). The most severe manifestation of epilepsy is SE. Historically, SE has been recognised as prolonged seizures that occur long enough, i.e., 30 minutes, to cause brain damage (Seinfeld et al., 2016). It is responsible for approximately 10% of epilepsy-related deaths (Trinka et al., 2023). The first-line medications for the treatment of early SE (seizure duration exceeding five minutes) are benzodiazepines, namely diazepam, lorazepam, and midazolam (Kim et al., 2021). These are GABA-A receptor potentiators. GABA-A receptors themselves are postsynaptic neurotransmitter-gated ion channels (Figure 2). Of note, γ-aminobutyric acid (GABA), the neurotransmitter that binds to these receptors, is the main inhibitory neurotransmitter of the nervous system.

When GABA binds to a GABA-A receptor, the GABA-A receptor undergoes structural changes resulting in the opening of the channel in the neuronal membrane. GABA-A receptor channels are permeable to Cl- ions, which flow into neurons. This ionic flow causes the hyperpolarization of neuronal membranes and renders neurons less excitable to stimuli. Benzodiazepines bind to GABA-A receptors at a site that is separate from that for GABA. By doing so, benzodiazepines increase the affinity of GABA-A receptors for GABA and, in turn, the opening frequency of the channels (Bianchi et al., 2009).

Figure 2: Cryo-electron microscopy maps of the GABAA receptor, the therapeutic target of some anti-epileptic drugs, e.g. benzodiazepines. Modified from Phulera et al. (2018, Figure 1).

About two-thirds of patients with early or stage I SE respond favourably to benzodiazepines, and about 40% of established or stage II SE cases are resistant to benzodiazepine treatment (Rollo et al., 2023). Of note, mortality in SE has been associated with drug resistance (Trinka et al., 2023). Therefore, benzodiazepine resistance presents a significant challenge to SE treatment.

KCC2 and OV350

KCC2 is a chloride-potassium co-transporter protein in the neuronal membrane that acts to reset the intracellular concentration of Cl- ions by pushing these ions out of the cell (Figure 3) (Liu et al., 2020). By doing so, KCC2 regulates the concentration gradient of Cl- ions across the neuronal membrane and, hence, the electrochemical driving force for the flow of these ions into the cell. In principle, if the extrusion of Cl- ions is dysregulated, the driving force for the flow of these ions through GABAARs can weaken, or its direction can even be reversed. Under KCC2 disruption, GABA can have a depolarizing effect on neurons (Hübner et al., 2001). In the context of epilepsy, previous research has shown that low KCC2 activity impacts the severity of SE (Silayeva et al., 2015), and potentiating its activity has been regarded as a potential therapeutic strategy (Moore et al., 2018). Furthermore, it has also been seen that KCC2 activity can improve benzodiazepine efficacy against sustained diazepam-resistant seizures (Cheung et al., 2022).

Jarvis and colleagues (2023) developed a compound, OV350, that activates KCC2. OV350 reduced intracellular Cl- concentrations in mouse hippocampal neurons in vitro, and also limited neuronal hyperexcitability in brain slices (Figure 3). Subcutaneous injections of OV350 and diazepam were effective in arresting kainic acid-induced, benzodiazepine-refractory SE and limiting neuronal injury in mice. As the authors discuss in their article, there are certain limitations tied to this research. The authors did not establish the effective dose for the OV350 compound, which would be the dose at which the compound arrests seizures with only minimal adverse events. Their observations were made on a single mouse model of SE, the kainic acid model. The findings will also have to be validated in additional models, e.g. lithium-pilocarpine model. Whether this compound can prevent the emergence of spontaneous or recurrent seizures is also something that merits investigation. Nonetheless, the current findings have lent further support for the role of KCC2 activation in the treatment of SE. Furthermore, they point to the potential of polytherapy as an intervention for treatment-resistant SE.

Figure 3: Co-administration of diazepam and OV350 reduced intracellular Cl- ion concentration, arrested benzodiazepine-resistant SE, and limited neuronal injury (Jarvis et al., 2023).

Conclusion

Status epilepticus (SE) is characterised by prolonged epileptic seizures, with the potential for brain damage and even death. Benzodiazepines act to inhibit pathological neuronal excitability in epilepsy, by potentiating GABAA­R activation and the subsequent flow of negatively-charged ions into neurons. KCC2 has emerged as a promising therapeutic strategy for benzodiazepine-resistant SE, given its role in resetting the concentration gradient of Cl- ions across the neuronal membrane. Jarvis et al. (2023) developed a KCC2 activator, OV350, which arrested benzodiazepine-refractory SE and limited neuronal injury in mice when it was co-administered with diazepam. It should be noted that there is a plethora of biological mechanisms in neurons that either favour or hinder neuronal excitability. Kim et al. (2021) support that early combinations of anti-epileptic drugs with different mechanisms of action could be beneficial in SE treatment. However, as the authors also point out, evidence for rational polytherapies against SE is still lacking. With their work, Jarvis et al. (2023) have offered needed insights into the potential of polytherapy for SE.

Bibliographic references

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