Glycosylphosphatidylinositol (GPI)-anchoring of calcium channel alpha2delta subunits: function and pharmacology
Lead Research Organisation:
University College London
Department Name: Neuroscience Physiology and Pharmacology
Abstract
In patients, damage to peripheral nerves results from many different diseases, and can result in chronic pain that is poorly treated by most of the commonly used pain killers. Common conditions such as diabetes and recurrent herpes zoster infection (shingles), may result in such chronic pain. These conditions are termed post-herpetic neuralgia and diabetic neuropathy. Moreover, treatment with certain forms of cytotoxic chemotherapy can also cause nerve damage, which may be irreversible. These types of pain due to nerve damage are grouped under the umbrella term ?neuropathic pain?. At their mildest, the symptoms would be tingling in the extremities, and at their worst they can involve stabbing spontaneous pain, or pain resulting from light touch. A drug called gabapentin and its newer relative pregabalin (initially developed for the treatment of epilepsy) are effective for many patients in the alleviation of this type of neuropathic pain.
Recent research has shown that gabapentin and pregabalin have their effect by binding to a protein (termed alpha2delta) that makes up part of a complex called a voltage-gated calcium channel. These calcium channels are present in all brain cells, and are essential for communication between them. Furthermore, the amount of this protein in pain-sensing nerves is increased several-fold in experimental models of neuropathic pain.
Our study aims to examine some of the fundamental properties of these alpha2delta proteins, as we have recently discovered an exciting and novel aspect of their structure. We now hope to understand at the molecular level, how these alpha2delta proteins work, and how the binding of gabapentin (and related drugs) to these proteins interferes with their correct function. The more we know about the properties of these proteins, the better we will be able to understand how drugs such as gabapentin have their action, which is an essential first step in the development of novel and potentially more effective drugs in this class. This is important as not all patients obtain relief of neuropathic pain symptoms by taking gabapentin, and the reason for this is not well understood. We therefore hope that our research will aid in the development of novel drugs, that might have the potential to be more effective in certain patients or in certain conditions.
Recent research has shown that gabapentin and pregabalin have their effect by binding to a protein (termed alpha2delta) that makes up part of a complex called a voltage-gated calcium channel. These calcium channels are present in all brain cells, and are essential for communication between them. Furthermore, the amount of this protein in pain-sensing nerves is increased several-fold in experimental models of neuropathic pain.
Our study aims to examine some of the fundamental properties of these alpha2delta proteins, as we have recently discovered an exciting and novel aspect of their structure. We now hope to understand at the molecular level, how these alpha2delta proteins work, and how the binding of gabapentin (and related drugs) to these proteins interferes with their correct function. The more we know about the properties of these proteins, the better we will be able to understand how drugs such as gabapentin have their action, which is an essential first step in the development of novel and potentially more effective drugs in this class. This is important as not all patients obtain relief of neuropathic pain symptoms by taking gabapentin, and the reason for this is not well understood. We therefore hope that our research will aid in the development of novel drugs, that might have the potential to be more effective in certain patients or in certain conditions.
Technical Summary
Voltage-gated calcium channels are essential for numerous processes in excitable cells, including transmitter release from neurons. The alpha2delta subunit is an accessory subunit of voltage-gated calcium channels, and four alpha2delta genes have been cloned. The alpha2delta subunit proteins are important for three main reasons:
Firstly, they are essential for the correct trafficking and functioning of voltage-gated calcium channels, and may also have other roles, such as cell adhesion.
Secondly, many forms of damage to peripheral sensory nerves, both in patients and animal models, may result in long-lasting neuropathic pain, associated with altered expression of many genes in damaged dorsal root ganglion (DRG) neurons, one of which is alpha2delta-1, which is strongly up-regulated.
Thirdly, alpha2delta-1 and alpha2delta-2 are the therapeutically-relevant binding sites for the gabapentinoid drugs, gabapentin and pregabalin, which are effective in the treatment of certain forms of epilepsy and in neuropathic pain.
We have found that these drugs do not cause acute block of calcium channels, but rather inhibit trafficking of the alpha2-delta subunits both in vitro and in vivo. We have further shown in vitro that trafficking of CaValpha1 subunits is also inhibited. They therefore represent an example of a novel mechanism of action of small molecule therapeutic agents, to inhibit ion channel trafficking.
It is therefore imperative that we understand better the mechanisms of alpha2delta processing in cells. Our recent novel and exciting finding that alpha2delta subunits are not, as previously thought, transmembrane proteins, but rather are attached to the external leaflet of the plasma-membrane by a glycosylphosphatidylinositol (GPI)-lipid-anchor, in a similar way to prion-proteins, has opened a new route to understanding their mechanism of action. GPI-anchored proteins are strongly localised in membrane microdomains termed lipid-rafts.
We will now examine the trafficking and localization of alpha2delta subunits in cell lines, polarized cells and neurons, in relation to their GPI-anchoring. We will examine the functional consequences and the biochemistry of GPI-anchoring, particularly of the major brain isoforms alpha2delta -1 and alpha2delta-3. We will also examine the consequences of the potential interaction of alpha2delta subunits with other lipid raft-associated proteins, such as prion proteins. Finally, a major objective is to examine the mechanism of action of gabapentinoid drugs in relation to the GPI-anchoring of alpha2delta-1 and -2, since such proteins utilise specific trafficking pathways.
This research has the potential to elucidate the mechanism of action of alpha2delta subunits, and the gabapentinoid drugs which bind to them.
Firstly, they are essential for the correct trafficking and functioning of voltage-gated calcium channels, and may also have other roles, such as cell adhesion.
Secondly, many forms of damage to peripheral sensory nerves, both in patients and animal models, may result in long-lasting neuropathic pain, associated with altered expression of many genes in damaged dorsal root ganglion (DRG) neurons, one of which is alpha2delta-1, which is strongly up-regulated.
Thirdly, alpha2delta-1 and alpha2delta-2 are the therapeutically-relevant binding sites for the gabapentinoid drugs, gabapentin and pregabalin, which are effective in the treatment of certain forms of epilepsy and in neuropathic pain.
We have found that these drugs do not cause acute block of calcium channels, but rather inhibit trafficking of the alpha2-delta subunits both in vitro and in vivo. We have further shown in vitro that trafficking of CaValpha1 subunits is also inhibited. They therefore represent an example of a novel mechanism of action of small molecule therapeutic agents, to inhibit ion channel trafficking.
It is therefore imperative that we understand better the mechanisms of alpha2delta processing in cells. Our recent novel and exciting finding that alpha2delta subunits are not, as previously thought, transmembrane proteins, but rather are attached to the external leaflet of the plasma-membrane by a glycosylphosphatidylinositol (GPI)-lipid-anchor, in a similar way to prion-proteins, has opened a new route to understanding their mechanism of action. GPI-anchored proteins are strongly localised in membrane microdomains termed lipid-rafts.
We will now examine the trafficking and localization of alpha2delta subunits in cell lines, polarized cells and neurons, in relation to their GPI-anchoring. We will examine the functional consequences and the biochemistry of GPI-anchoring, particularly of the major brain isoforms alpha2delta -1 and alpha2delta-3. We will also examine the consequences of the potential interaction of alpha2delta subunits with other lipid raft-associated proteins, such as prion proteins. Finally, a major objective is to examine the mechanism of action of gabapentinoid drugs in relation to the GPI-anchoring of alpha2delta-1 and -2, since such proteins utilise specific trafficking pathways.
This research has the potential to elucidate the mechanism of action of alpha2delta subunits, and the gabapentinoid drugs which bind to them.
Organisations
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ORCID iD |
| Annette Dolphin (Principal Investigator) |