Jun 272008
 
ResearchBlogging.orgAs I mentioned in my post on the recent paper by Kukar et al., the disruption of amyloid plaques has been an ongoing focus in Alzheimer’s disease research. However, plaques and inclusions are of concern in many diseases, and as a result there is a great deal of interest in finding molecules that can either dissociate, or prevent the formation of, amyloids of many different kinds of proteins. In the most recent issue of Nature Structural and Molecular Biology there is a paper suggesting that (-)-epigallocatechin gallate (EGCG) may be able to interfere with multiple proteins that form β-rich aggregates.

EGCG is a chemical found in green tea (although it is doubtful you could realistically drink enough green tea to absorb the concentrations used in this study). Previous studies had suggested that it altered the aggregation behavior of α-synuclein (αS) and huntingtin. So, Ehrnhoefer et al. use highly purified EGCG in a number of experiments to determine what effect it had on αS and the amyloid-β peptide (Aβ) (1). What these proteins have in common, besides the fact that their aggregation is associated with disease, is that the single proteins take on a β-strand structure that assembles into fibrils.

Ehrnhoefer et al. find that EGCG interferes with some aspect of this process, reducing the formation of the fibrils while inducing the formation of some alternate oligomeric structure. In the case of αS, the result is a spherical oligomer (Figure 1), although the gel filtration results indicate that a large spectrum of oligomeric states is formed at lower concentrations (the trace suggests that these oligomeric forms are interconverting during elution). NMR and other data show that EGCG associates directly, but non-specifically, with the protein backbone, and that the compound has strongest affinity for the C-terminus of αS, which may play a role in preventing aggregation. The EGCG-treated oligomers had reduced β-strand content (as assayed by CD). Treatment with EGCG appeared to reduce αS toxicity in cultured cells, although this was measured strictly in terms of cell death.

Similar results were seen with Aβ—addition of EGCG reduced the formation of fibrils and the toxicity of amyloids towards cultured cells. Again, the oligomers formed in the presence of EGCG could be quite large, and took a spherical shape.

Based on these data, the authors propose that EGCG binds preferentially to unfolded proteins and interferes with the formation of regular β-strand structure. The EGCG-bound proteins are unable to form fibrils, and therefore EGCG oligomers compete with fibrils for monomers, slowing the formation of the latter. The net effect is to divert these unfolded proteins out of amyloidogenic pathways and into alternate oligomeric structures, which appear to be nontoxic, or at least less toxic.

Can EGCG or a derivative be turned into a drug to treat Alzheimer’s disease, or a general treatment for amyloidoses? This is an uncertain proposition. As the authors of a commentary (2) in the same issue of NSMB note, EGCG’s nonspecific assault on amyloids may damage some normal structures built on this architecture. Moreover, because EGCG seems to bind unfolded regions nonspecifically, it has the potential to interfere with any of the numerous signaling proteins that possess such regions. The potential for side effects is very high, and the continued viability of cultured cells in the presence of EGCG, while reassuring, is not a particular reason to believe the compound is safe at high concentrations in the human nervous system.

The promiscuity of EGCG’s interactions with unfolded regions poses another problem, in that all these proteins will act to interfere with EGCG’s action on its intended target. Fairly high ratios of EGCG were necessary in these assays, and they mostly involved purified proteins. In vivo, all unfolded proteins will act to titrate EGCG out of plasma, meaning that significant quantities of this (or any other non-specific molecule like it) would need to be used in order to achieve the desired effect. This again raises the likelihood of side effects.

Of course, the most severe complication arises from the nature of amyloidoses themselves. Although the obvious presence of plaques and inclusions naturally leads us to suspect that these aggregates are the agents causing the disease, increasing evidence suggests that amyloids are merely the endpoints of some other process that is the actual culprit. In the case of Alzheimer’s disease, for instance, a recent article in Nature Medicine has provided very strong evidence that soluble Aβ dimers are the dominant contributors to Alzheimer’s pathophysiology (go check out Ashutosh’s excellent discussion of this article over at The Curious Wavefunction for more information). Now, given that these dimers do eventually form amyloid, it seems likely that they have β structure in their pathogenic form, which EGCG will probably disrupt, but this is not guaranteed. Small molecules like EGCG that prevent deposition into amyloid may actually exacerbate the problems they are meant to solve. Further research is needed to establish that the EGCG oligomers of αS and Aβ are not toxic in vivo.

Diseases associated with protein aggregation continue to pose a challenge precisely because we have such a poor handle on their pathogenesis. Ehrnhoefer et al. clearly demonstrate that EGCG possesses the ability to alter the behavior of amyloidogenic unfolded proteins. While that may imply that it has promise as a broad-spectrum drug to attack these diseases, the promiscuity of its action is a cause for concern from the perspective of dosage and side effects. And, because soluble oligomers may well be the pathogenic species in many (if not all) of these diseases, our optimism about this approach must be tempered with an awareness that the actual effect of EGCG may be to enhance, rather than diminish, the toxicity of the relevant protein targets.

1. Ehrnhoefer, D.E., Bieschke, J., Boeddrich, A., Herbst, M., Masino, L., Lurz, R., Engemann, S., Pastore, A., Wanker, E.E. (2008). EGCG redirects amyloidogenic polypeptides into unstructured, off-pathway oligomers. Nature structural & molecular biology, 15(6), 558-566. DOI: 10.1038/nsmb.1437

2. Roberts, B.E., Shorter, J. (2008). Escaping amyloid fate. Nature Structural & Molecular Biology, 15(6), 544-546. DOI: 10.1038/nsmb0608-544

  2 Responses to “EGCG disrupts amyloid oligomers”

  1. This explains why my tubulin assay wasn't working the other day when i spilled green tea all over the spec… oh shit, perhaps i've said too much….

  2. Thanks for the plug. It just keeps on getting more and more interesting. Recently I have started thinking of AD as a possible evolutionary adaptation against pathogens. Will pen my thoughts on this sometime.

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