Mechanism of Inhibition of Viral and Neuronal Pore Loop Ion Channels by the Adamantanes

Essentially all living cells have channel-like proteins embedded in their cell walls. The presence of these channels selectively allow certain classes of charged molecules, or ions, to pass into or out of the cellular interiors, as cell walls by themselves are normally impermeable to such molecules. Channels are often critical components of cells, and opening and closing ion channels is at the centre of many normal cellular processes such as cell fate decisions or intercellular communication. Ion channels are also at the root of some abnormal processes that arise from genetic mutations such as multiple sclerosis, or have functions that are essential for pathogen viability such as in the influenza virus. For this reason, many useful drugs act by forcing ion channels to open or close. One family of drugs that function in this way are the adamantanes. They bind to, and close, a surprisingly wide variety of ion channels, including a proton channel in the flu virus called M2, and calcium channels that are found in humans at neuronal synapses called NMDA receptors, which are involved in memory and learning. For these reasons, adamantanes are prescribed both for the treatment of flu infections, and to alleviate the symptoms of neurodegeneration associated with diseases such as Parkinsons and Alzheimers. Drugs typically interact specifically with proteins such as ion channels at one particular site, and knowing the precise physical properties of those interactions can facilitate design of more specific, and therefore more effective or less toxic drugs. In previous work, we elucidated the physical location at which the adamantanes bind to the flu virus ion channel. Unfortunately, little is known about the same process in the NMDA receptor. We now hope to use what we learned in the M2 case, to understand what physical interactions are needed for binding to the NMDA receptors. In addition, adamantanes such as amantadine and memantine are part of a larger class of potentially therapeutic compounds that bind to the NMDA receptor. Thus, understanding the behaviour of the adamantanes may be broadly applicable for guiding development of more effective drug treatments for neurodegenerative diseases.

Adamantane-based drugs inhibit a wide range of ion channels with different structures and ion selectivities. Historically, they have been used to treat influenza infections because they inhibit the virus M2 proton channel. The spread of resistant virus has, however, rendered the adamantanes essentially useless. Recent experiments in this lab have suggested that the M2 adamantane binding site is composed of functionally important, and therefore highly conserved, residues; there is therefore the possibility that alternative antiviral drugs could be found by targeting this conserved pocket. Thus, Part A of this proposal is aimed at determining the components of M2 that are critical for drug binding, with the ultimate goal of informing the design of next generation antivirals. Adamantanes are also used for the treatment of dementia associated with Alzheimers and Parkinsons diseases. The therapeutic target for this indication is the ion channel domain of the ionotropic NMDA receptor. Although, high-resolution structures of the intact ionotropic NMDA receptor have not been determined due to its size and complexity, the viral ion channel Kcv is a structurally homologous pore-loop ion channel that is also inhibited by the adamantane amantadine. Thus, Kcv is a useful surrogate for studies that focus on adamantane binding and inhibition of this class of ion channels. In Part B and C of the proposal, we will use techniques developed for the investigation of adamantane binding to M2 to establish and exploit an experimental system for the study of adamantane binding to Kcv at the atomic level, and by extension, to the NMDA receptor ion channel. Functional studies using liposomal ion fluxes will validate and extend the structural work. It is anticipated that a detailed characterization of the position and structural and dynamic effects of drug binding within the context of these disparate ion channel structures will provide information for new approaches to drug design for this class of valuable ion channel inhibitors.