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Molecular imaging of intracellular drug-membrane aggregate formation.

ABSTRACT: Clofazimine is a lipophilic antibiotic with an extremely long pharmacokinetic half-life associated with the appearance of crystal-like drug inclusions, in vivo. Here, we studied how clofazimine accumulates inside cells in the presence of supersaturating, extracellular concentrations of the drug (in the range of physiological drug concentrations). Based on a combination of molecular imaging, biochemical analysis and electron microscopy techniques, clofazimine mass increased inside cells in vitro, over a period of several days, with discrete clofazimine inclusions forming in the cytoplasm. These inclusions grew in size, number and density, as long as the drug-containing medium was replenished. With Raman confocal microscopy, clofazimine's spectral signature in these inclusions resembled that of amorphous clofazimine precipitates and was unlike that of clofazimine crystals. Additional experiments revealed that clofazimine first accumulated in mitochondria, with ensuing changes in mitochondrial structure and function. In turn, the degenerating organelles coalesced, fused with each other and condensed to form prominent drug-membrane aggregates (dubbed autophagosome-like drug inclusions or "aldis"). Like clofazimine, it is possible that intracellular drug-membrane aggregate formation is a common phenomenon underlying the reported phenotypic effects of many other small molecule drugs.

SUBMITTER: Baik J

PROVIDER: S-EPMC3185106 | biostudies-literature | 2011 Oct

REPOSITORIES: biostudies-literature

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Molecular imaging of intracellular drug-membrane aggregate formation.

Baik Jason J Rosania Gus R GR

Molecular pharmaceutics 20110812 5

Clofazimine is a lipophilic antibiotic with an extremely long pharmacokinetic half-life associated with the appearance of crystal-like drug inclusions, in vivo. Here, we studied how clofazimine accumulates inside cells in the presence of supersaturating, extracellular concentrations of the drug (in the range of physiological drug concentrations). Based on a combination of molecular imaging, biochemical analysis and electron microscopy techniques, clofazimine mass increased inside cells in vitro, ...[more]

PMID: 21800872

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Project description:As an important neuropathological hallmark of Alzheimer's disease (AD), the oligomerization of amyloid-β (Aβ) peptides has been intensively investigated in both theoretical and experimental studies. However, the oligomerization space in terms of the kinetics, molecular mechanism, and oligomer structures remains mysterious to us. An equation that can quantitatively describe the time it takes for Aβ oligomers to appear in the human brain at a given Aβ monomer concentration is extremely vital for us to understand the development and disease progression of AD. In this study, we utilized molecular dynamics (MD) simulations to investigate the oligomerization of Aβ42 peptides at five different monomer concentrations. We have elucidated the formation pathways of Aβ tetramers, characterized the oligomer structures, estimated the oligomerization time for Aβ dimers, trimers, and tetramers, and for the first-time derived equations that could quantitatively describe the relationship between the oligomerization time and the monomer concentration. Applying these equations, our prediction of oligomerization time agrees well with the experimental and clinical findings, in spite of the limitations of our oligomerization simulations. We have found that the Aβ oligomerization time depends on the monomer concentration by a power of -2.4. The newly established equations will enable us to quantitatively estimate the risk score of AD, which is a function of age. Moreover, we have identified the most dominant pathway of forming Aβ tetramers, probably the most important and toxic Aβ oligomer. Our results have shown that the structures of Aβ42 dimer, trimer, and tetramer, which are distinguishable from each other, depend on the monomer concentration at which the oligomers form. Representative oligomer structures, which can serve as potential drug targets, have been identified by clustering analysis. The MD sampling adequacy has been validated by the excellent agreement between the calculated and measured collisional cross section (CCS) parameters (the prediction errors are within 2%). In a conclusion, this study provides the kinetics and structure basis for developing inhibitors to decelerate the Aβ oligomerization process.

Dataset Information

Molecular imaging of intracellular drug-membrane aggregate formation.

Publications

Molecular imaging of intracellular drug-membrane aggregate formation.

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