Characterisation of the Binding of Cationic Amphiphilic Drugs to Phospholipid Bilayers Using Surface Plasmon Resonance

Nussio, Matthew R.; Sykes, Matthew J.; Miners, John O.; Shapter, Joseph G.

doi:10.1002/cmdc.200600252

Cited by 21 publications

(21 citation statements)

References 22 publications

(37 reference statements)

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“…Eq. 1 is adapted from the Biacore Biaevaluation software and includes two separate binding-affinity terms, K Dlipid and K Dpeptide (30,40) (in the case where drugs were bound to empty liposomes, the K Dpeptide component of Eq. 1 was not used):…”

Section: Methodsmentioning

confidence: 99%

Coexistence of two adamantane binding sites in the influenza A M2 ion channel

Rosenberg

Casarotto

2010

Proc. Natl. Acad. Sci. U.S.A.

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The influenza A virus contains a proton-selective ion channel (M2) that is the target of the adamantane family of drug inhibitors. Two recently published studies relating to adamantane binding of the M2 ion channel using X-ray crystallography and solution NMR have reignited interest in the potential use of adamantanes in combating the spread of influenza A. However, these two studies propose different binding sites for the adamantane drugs with the X-ray M2/ amantadine structure favoring an ion channel pore-binding model and the solution NMR M2/rimantadine structure suggesting the existence of a lipid-facing binding pocket. We conducted a series of surface plasmon resonance (SPR) experiments designed to accurately measure the affinity of amantadine and rimantadine to M2 ion channels embedded in 1,2-dimyristoyl-sn-glycero-phosphocholine (DMPC) liposomes. We find that this class of drug is capable of binding M2 with two different affinities in the order of 10 −4 and 10 −7 M, suggesting that both proposed binding sites are feasible. Furthermore, by examining drug binding to M2 mutant constructs (V27A, S31N, and D44A), it was possible to probe the location of the two binding sites. We show that a high-affinity binding site corresponds to the M2 ion channel pore whereas the secondary, low-affinity binding site can be attributed to the lipid face of the pore. These SPR results are in excellent agreement with the most recent solid-state NMR study of amantadinebound M2 in lipid bilayers and provide independent support that the ion channel pore-binding site is responsible for the pharmacological activity elicited by the adamantane drugs.adamantane drug resistance | surface plasmon resonance | viral ion channel | mutational analysis A s evidenced by recent H5N1 (avian) and H1N1 (swine) influenza outbreaks, the influenza A virus poses an ever-present global threat to human health. One of the initial responses in combating this disease involves the use of antiviral drugs such as the neuraminidase inhibitors oseltamivir and zanamivir. Whereas the efficacy of these drugs is well established, another class of approved antiviral drug known as adamantane M2 inhibitors (amantadine and rimantadine) has in the past been shown to be effective in treating seasonal influenza (1). Unfortunately, the emergence of several adamantane-resistant M2 mutations has severely curtailed the effectiveness of this class of drug to the extent that the Centers for Disease Control have recommended discontinuing its use (2). However, the recent flu outbreaks have demonstrably highlighted the immense benefit of having an effective antiviral treatment at hand, thereby reigniting the search for novel M2 inhibitors (3, 4).The role of the influenza A M2 protein has been well documented. It forms a proton ion channel that operates early in the viral life cycle by facilitating the acidification of the endosomally entrapped virus, thereby enabling release of its RNA genome to allow viral replication (5). The M2 ion channel is 97 amino acids in size but it is t...

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Section: Methodsmentioning

confidence: 99%

Coexistence of two adamantane binding sites in the influenza A M2 ion channel

Rosenberg

Casarotto

2010

Proc. Natl. Acad. Sci. U.S.A.

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“…[11] CADs are able to interact with both the carbon core region and the polar head groups of lipid membranes. [12] Thus, CADs easily interact with biological membranes.…”

Section: Introductionmentioning

confidence: 99%

Identification of Drugs Inducing Phospholipidosis by Novel in vitro Data

et al. 2012

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Drug-induced phospholipidosis (PLD) is a lysosomal storage disorder characterized by the accumulation of phospholipids within the lysosome. This adverse drug effect can occur in various tissues and is suspected to impact cellular viability. Therefore, it is important to test chemical compounds for their potential to induce PLD during the drug design process. PLD has been reported to be a side effect of many commonly used drugs, especially those with cationic amphiphilic properties. To predict drug-induced PLD in silico, we established a high-throughput cell-culture-based method to quantitatively determine the induction of PLD by chemical compounds. Using this assay, we tested 297 drug-like compounds at two different concentrations (2.5 μm and 5.0 μm). We were able to identify 28 previously unknown PLD-inducing agents. Furthermore, our experimental results enabled the development of a binary classification model to predict PLD-inducing agents based on their molecular properties. This random forest prediction system yields a bootstrapped validated accuracy of 86 %. PLD-inducing agents overlap with those that target similar biological processes; a high degree of concordance with PLD-inducing agents was identified for cationic amphiphilic compounds, small molecules that inhibit acid sphingomyelinase, compounds that cross the blood–brain barrier, and compounds that violate Lipinski’s rule of five. Furthermore, we were able to show that PLD-inducing compounds applied in combination additively induce PLD.

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“…24 The label-free technique of surface plasmon resonance has also been applied to detect drug-lipid membrane binding. 25,26 In these studies, liposomes attached to a lipophilic functionalized gold surface were used as model membranes for drug binding.…”

Section: Introductionmentioning

confidence: 99%

High-Throughput Screening of Drug–Lipid Membrane Interactions via Counter-Propagating Second Harmonic Generation Imaging

Nguyen

Conboy

2011

Anal. Chem.

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Here we report the use of counter-propagating second harmonic generation (SHG) to image the interactions between the local anesthetic tetracaine and a multi-component planar supported lipid bilayer array in a label-free manner. The lipid bilayer arrays, prepared using a 3D continuous flow microspotter, allow the effects of lipid phase and cholesterol content on tetracaine binding to be examined simultaneously. SHG images show that tetracaine has a higher binding affinity to liquid-crystalline phase lipids than to solid-gel phase lipids. The presence of 28 mol % cholesterol decreased the binding affinity of tetracaine to bilayers composed of the mixed chain lipid, 1-steroyl-2-oleoyl-sn-glycero-3-phophocholine (SOPC) and the saturated lipids 1,2-dimyristoyl-sn-glycero-3-phophocholine (DMPC) and 1,2-dipamitoyl-sn-glycero-3-phophocholine (DPPC) while having no effect on di-unsaturated 1,2-dioleoyl-sn-glycero-3-phophocholine (DOPC). The maximum surface excess of tetracaine increases with the degree of unsaturation of the phospholipids and decreases with cholesterol in the lipid bilayers. The paper demonstrates that SHG imaging is a sensitive technique that can directly image and quantitatively measure the association of a drug to a multi-component lipid bilayer array, providing a high-throughput means to assess drug-membrane interactions.

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Characterisation of the Binding of Cationic Amphiphilic Drugs to Phospholipid Bilayers Using Surface Plasmon Resonance

Cited by 21 publications

References 22 publications

Coexistence of two adamantane binding sites in the influenza A M2 ion channel

Coexistence of two adamantane binding sites in the influenza A M2 ion channel

Identification of Drugs Inducing Phospholipidosis by Novel in vitro Data

High-Throughput Screening of Drug–Lipid Membrane Interactions via Counter-Propagating Second Harmonic Generation Imaging

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