Biosensor Analysis of the Interaction between Immobilized Human Serum Albumin and Drug Compounds for Prediction of Human Serum Albumin Binding Levels

Frostell-Karlsson, Åsa; Remaeus, Annika; Roos, Håkan; Andersson, Karl; Borg, Peter; Hämäläinen, M.; Karlsson, Robert

doi:10.1021/jm991174y

Cited by 293 publications

(227 citation statements)

References 34 publications

(37 reference statements)

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“…The relative affinity of these compounds could be compared using this method due to their similar mass and structure. This method is similar to that employed in other studies that determined the relative affinities of related compounds (36). It provides only a relative assessment of binding and should not be directly compared to dissociation constants.…”

Section: Resultsmentioning

confidence: 99%

Affinity-Based Inhibition of β-Amyloid Toxicity

et al. 2002

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Strategies for interfering with protein aggregation are important for elucidating and controlling the pathologies of amyloid diseases. We have previously identified compounds that block the cellular toxicity of the -amyloid peptide, but the relationship between their ability to inhibit toxicity and their affinity for A is unknown. To elucidate this relationship, we have developed an assay capable of measuring the affinities of small molecules for -amyloid peptide. Our approach employs immobilized -amyloid peptide at low density to minimize the problems that arise from variability in the -amyloid aggregation state. We found that low-molecular weight (MW of 700-1700) ligands for -amyloid can be identified readily by using surface plasmon resonance. The best of these bound effectively (K d ∼ 40 µM) to -amyloid. The affinities measured for peptides in the SPR assay correspond to results from A cell toxicity assays. The most potent ligands for immobilized -amyloid are the most potent inhibitors of the neuronal cell toxicity of -amyloid. Compounds with dissocation constants above ∼100 µΜ did not show significant activity in the cell toxicity assays. Our data support the hypothesis that ligands exhibiting greater affinity for the -amyloid peptide are effective at altering its aggregation and inhibiting cell toxicity.

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Section: Resultsmentioning

confidence: 99%

Affinity-Based Inhibition of β-Amyloid Toxicity

et al. 2002

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“…SPR data were double referenced [31][32][33] and analyzed with Biaevaluation software 4.1 (Biacore AB). Sensorgrams were corrected for organic co-solvent effects [35], but as witnessed by others the use of methanol as the organic co-solvent and the use of suitable control surfaces made correction factors negligible [36].…”

Section: Surface Plasmon Resonancementioning

confidence: 99%

Surface Plasmon Resonance Analysis of Antifungal Azoles Binding to CYP3A4 with Kinetic Resolution of Multiple Binding Orientations

et al. 2006

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The heme-containing Cytochrome P450s (CYPs) are a major enzymatic determinant of drug clearance and drug-drug interactions. The CYP3A4 isoform is inhibited by antifungal imidazoles or triazoles, which form low spin heme iron complexes via formation of a nitrogen-ferric iron coordinate bond. However, CYP3A4 also slowly oxidizes the antifungal itraconazole (ITZ) at a site that is ∼ 25 Å from the triazole nitrogens, suggesting that large antifungal azoles can adopt multiple orientations within the CYP3A4 active site. Here, we report a surface plasmon resonance (SPR) analysis with kinetic resolution of two binding modes of ITZ, and the related drug ketoconazole (KTZ). SPR reveals a very slow off-rate for one binding orientation. Multiphasic binding kinetics are observed and one of the two binding components resolved by curve-fitting exhibits 'equilibrium overshoot'. Pre-loading of CYP3A4 with the heme ligand imidazole abolishes this component of the antifungal azole binding trajectories, and it eliminates the conspicuously slow off-rate. The fractional populations of CYP3A4 complexes corresponding to different drug orientations can be manipulated by altering the duration of the pulse of drug exposure. UV-vis difference absorbance titrations yield low spin spectra and K D values that are consistent with the high affinity complex resolved by SPR. These results demonstrate that ITZ and KTZ bind in multiple orientations, including a catalytically productive mode and a slowly-dissociating inhibitory mode. Most importantly, they provide the first example of an SPR-based method for the kinetic characterization of drug binding to any human CYP, including mechanistic insight not available from other methods. Keywords protein-ligand interactions; tight binding inhibitors; drug metabolism; drug interactionsThe hepatic and intestinal heme-containing Cytochrome P450s (CYPs) 1 oxidize most drugs, and thus play a critical role in their metabolism and disposition [1][2][3]. Among the various human CYP isoforms, CYP3A4 contributes most to drug metabolism. CYP3A4 exhibits complex allosteric kinetics, which may result from drug-drug interactions on a single CYP molecule through multiple drug binding, as suggested by steady state kinetic behavior [4][5][6], large active sites observed in crystal structures [7][8][9][10] and spectroscopic studies [11]. Complex kinetics † This work was supported by NIH grants GM32165 (WMA, NI, KLK) and GM07750 (JTP). * Corresponding author: Tel: (206) FAX: (206) [12][13][14][15], and equilibrium optical titrations that rely on the inhibitor-or substrate-dependent changes in ferric spin state equilibrium [16][17][18]. The complexity of CYP kinetics severely impedes prediction of drug clearance and drug-drug interactions [12][13][14][15]19]. Clearly, new methods are required to elucidate the molecular basis of CYP allosterism and complex inhibitor or substrate binding.The antifungal azoles, including ketoconazole (KTZ) and itraconazole (ITZ), historically have been considered to be potent inh...

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“…This methodology results particularly promising for their potential application in drug protein binding screening and in kinetics investigations. [22][23][24][25][26] The present review essentially focuses on the application of the use of both immobilized albumin (biochromatography) and CD on the same protein-drug system in solution, largely on the basis of our previous publications. These methodologies have been shown to be efficient in characterizing the binding sites on the protein and to give evidence of the changes in the binding properties of the protein arising by interaction between different ligands.…”

Section: Current Approaches For the Study Of The Binding Properties Omentioning

confidence: 99%

“…This methodology is well suited for screening assay, because of the miniaturization of the system and the short time for the analysis that facilitates its automatization. [22][23][24][25][26] Moreover, with optical biosensors, neither of the interacting molecules needs to be labeled, and the response obtained is directly proportional to the mass of the analyte that binds to the surface. Therefore, the ligand binding interaction occurs in a condition as close as possible to its native state.…”

Section: Biorecognition On Surface Immobilized Hsamentioning

confidence: 99%

Drug binding to human serum albumin: Abridged review of results obtained with high‐performance liquid chromatography and circular dichroism

2006

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The drug binding to plasma and tissue proteins are fundamental factors in determining the overall pharmacological activity of a drug. Human serum albumin (HSA), together with a 1 -acid glycoprotein (AGP), are the most important plasma proteins, which act as drug carriers, with drug pharmacokinetic implications, resulting in important clinical impacts for drugs that have a relatively narrow therapeutic index. This review focuses on the combination of biochromatography and circular dichroism as an effective approach for the characterization of albumin binding sites and their enantioselectivity. Furthermore, their applications to the study of changes in the binding properties of the protein arising by the reversible or covalent binding of drugs are discussed, and examples of physiological relevance reported. Perspectives of these studies reside in supporting the development of new drugs, which require miniaturization to facilitate the screening of classes of compounds for their binding to the target protein, and a deeper characterization of the mechanisms involved in the molecular recognition processes.

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Biosensor Analysis of the Interaction between Immobilized Human Serum Albumin and Drug Compounds for Prediction of Human Serum Albumin Binding Levels

Cited by 293 publications

References 34 publications

Affinity-Based Inhibition of β-Amyloid Toxicity

Affinity-Based Inhibition of β-Amyloid Toxicity

Surface Plasmon Resonance Analysis of Antifungal Azoles Binding to CYP3A4 with Kinetic Resolution of Multiple Binding Orientations

Drug binding to human serum albumin: Abridged review of results obtained with high‐performance liquid chromatography and circular dichroism

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