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The protein which inspired the artwork for our featured structure for April 2021 is the glutamate receptor 2, an important mediator of the synaptic transmissions in the central nervous system, playing a key role in the generation and spread of epileptic seizures.

 

Epilepsy, a burden for many

Epilepsy is a chronic brain disease that affects around 50 million people of all ages worldwide, making it one of the most common neurological diseases globally. People suffering from epilepsy are subject to seizure episodes in which they make involuntary movements and can sometimes lose consciousness. Those seizure episodes result of excessive electrical discharges in different parts of the brain. Although many underlying disease mechanisms can lead to epilepsy, the cause of the disease is still unknown in about 50% of cases globally. Nevertheless, seizures can be controlled. Up to 70% of people living with epilepsy could become seizure-free if they were properly diagnosed and treated.

 

Glutamate receptors, targets for epilepsy therapy

Glutamate receptors are ionotropic transmembrane receptors for the neurotransmitter glutamate, that mediate synaptic transmissions in the central nervous system. Another name for glutamate receptors is AMPA receptors, derived from their ability to be activated by the artificial glutamate analogue AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) which mimics the agonist effects of glutamate.

Glutamate receptors mediate the majority of excitatory neurotransmission, playing a key role in the generation and spread of epileptic seizures, and therefore are promising targets for epilepsy therapy.

 

 

Epilepsy treatment

Treatment can help most people with epilepsy have fewer seizures, and even if some people need treatment for life, some may be able to stop if their seizures disappear over time. Anti-epileptic drugs (AEDs) are the most commonly used treatment for epilepsy. Even if they do not cure epilepsy, they help control seizures in around 7 out of 10 people.

Prior to the 19th century, epilepsy was mostly referred to as the “Sacred Disease�, and its treatment was based on spiritual and supernatural beliefs. Investigation around therapies for epilepsy began in the late 1800s, when electrical stimulation and chemo-convulsants were tested on animals. In the early 1930s, Merritt and Putnam were able to setup a proper methodology allowing them to screen for hundreds of compounds, which allowed them to discover phenytoin. The clinical efficacy of phenytoin was confirmed in 1938, and this drug became the primary treatment for various forms of epilepsy and is still commercially available nowadays.

Phenytoin, as many AEDs, work by changing the levels of chemicals in the patients� brain, tempering with synaptic and non-synaptic receptors such as the glutamate receptor.

 

Glutamate receptor non-competitive antagonists

The most potent and well-tolerated (with fewer side effects) AEDs that act as inhibitors of glutamate receptors act via a non-competitive (negative allosteric) mechanism. Indeed, those drugs function effectively even in the presence of high levels of glutamate.

The first non-competitive glutamate receptor antagonist described was 1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine hydrochloride (GYKI 52466) is an orally-active anticonvulsant, and skeletal muscle relaxant. Even though GYKI 52466 didn’t make its way to being used as a therapeutic drug, it became a prototype for the development of more potent and selective 2,3-benzodiazepines. Besides 2,3-benzodiazepines, other classes of compounds were used as a base for developing therapeutic drugs, such as phthalazine and quinazolinone derivatives.

A number of those compounds have progressed into clinical studies. One of them, Perampanel, was the first antiepileptic drug in the class of selective non-competitive antagonist of glutamate receptors to be approved by the European Commission, and later by the Food and Drug Administration in 2012. It is now used in addition to other drugs to treat partial seizures.

Even if Perampanel was approved for medical use as a safe and effective antiepileptic drug with low incidence of serious adverse effects at low doses, patients taking Perampanel at higher doses do experience worrying side effects, including fatigue, nausea, headache, vertigo, as well as depression and aggression, indicating the need for safer and more efficacious drugs.

 

 

Glutamate receptor 2 and Perampanel

The identification of AEDs has always required the use of seizure models. In the case of the study of glutamate receptor 2 (GluA2), the rat GluA2 and human GluA2 share 99.5% sequence identity, so even if no rat need to be harmed, the rat GluA2 can be studied interchangeably with its human homologue when producing recombinantly expressed protein in structural studies with AEDs.

GluA2’s structure comprises three domains:

• the amino-terminal domain (NH2 domain) and ligand binding domain (LBD), both located outside of the membrane of postsynaptic cells;
• the ion channel, the pore located in the postsynaptic cellsâ€� transmembrane.

GluA2 has an overall Y shape characteristic of the glutamate receptors. It is tetrameric and can form homo-tetramers as well as hetero-tetramers with other glutamate receptor family members. Each GluA2 has four sites to which glutamate can bind, in the LBD of each subunit.

The structure of rat GluA2 with Perampanel (PDB ID ) shows that there are also four sites to which Perampanel can bind, and those sites are located at the interface between the ion channel and linkers connecting it to the LBD.

 

Rat GluA2 (PDB ID ) domains and topology. Left: Cartoon view of the homo-tetramer GluA2 viewed parallel to the membrane, with NH2-domain, LDB and ion channel highlighted in orange, blue and pink respectively. Inner and outer sides of the membrane are indicated by parallel grey bars.

 

Rat GluA2 (PDB ID ) tetramer and Perampanel binding site. Left: Cartoon view of the homo-tetramer GluA2 viewed parallel to the membrane, each subunit in a different colour. Right: Top view of the ion channel, each subunit in a different colour. The four Perampanel molecules are highlighted in spheres.

When glutamate binds, the extracellular loops between the LBD and the ion channel move towards each other, opening the pore. Perampanel binding at the interface between the LBD and the ion channel likely immobilizes the domains relative to each other, preventing the transmission of conformational changes necessary for the ion channel opening and therefore stabilizing the GluA2 closed state.

Perampanel is one drug among many for treating epilepsy, and gaining a structural understanding of how AEDs carry out their non-competitive inhibition will aid the development of improved drugs targeting glutamate receptors.

Deborah Harrus

 

About the artwork

“For my artwork, I have etched the protein for Glutamate Receptor 2 for Antiepileptic Drugs to create a piece of art on epilepsy. I decided to recreate the 3D image of the protein as the filament of the lightbulb; the protein represents how the medication is fighting the seizures that are triggered by light. The lightbulb is switched off because the protein for the antiepileptic drug is preventing the seizures from happening therefore removing the danger that the lightbulb imposes. Photosensitive epilepsy is not common however it still infiltrates people’s lives and prevents them from experiences that happen every day.�

 Esther Robson, a student from Leys (Cambridge, UK).

 View the artwork in the .

 

Structures mentioned in this article

Structure of the AMPA subtype ionotropic glutamate receptor GluA2 in complex with non-competitive inhibitor Perampanel:

 

Sources

Perampanel (PDBe)