Targeting early AD oligomers with a simple dipeptide

A couple of days ago, an American man in Switzerland became the first human being to have his assisted suicide filmed and broadcasted on television. Craig Ewert was a 59 year old university professor who had been diagnosed with motor neuron disease in 2006. He had been given 3-5 years to live, but the disease took its terrible toll in just a few months. By the end of 2006 Ewert decided he had had enough. He contacted Dignitas, a Swiss organization started by a lawyer that has helped literally hundreds of people to peacefully end their lives. On a quiet, sunny morning, Ewert sipped a lethal cocktail of barbiturates completely of his own volition, and then bit on a switch that stopped his ventilator, again of his own volition. The film crew was allowed to film the entire operation except for after he passed away, when his wife wanted a few minutes of silence to herself.

Assisted suicide is an immensely controversial topic laden with moral issues. I have thought about it and discussed it with my friends many times, and I usually find myself ending up in favour of it at least in some cases where death is inevitable and the quality of life is going to assuredly become worse. Motor neuron disease is one such ailment. Alzheimer's disease is another. I personally don't think there's any other disease as cruel as AD, with loved ones trying to grasp at a person's identity as it slips away all too slowly and painfully. To me AD seems to be one of those unambiguous cases where a person who is still coherent should be able to exercise his right to die a dignified death. As horrible as it is to watch a loved one decide to willingly end his or her life, it is usually infinitely worse to watch them fade away into a wilderness of silence.

But the insidious pain of AD also makes the search for AD therapies a particularly compelling and desperate one. 15 million people are afflicted with AD, and in 20 years that burden is supposed to double. Part of the trouble for targeting AD is a still incomplete understanding of its molecular basis. While the initial amyloid hypothesis that posits the formation of insoluble amyloid protein fibrils is still very much relevant, much less is known about its exact relation to the disease, whether as a cause or symptom. The most important recent finding with respect to this hypothesis is that it's not insoluble aggregates but soluble oligomers that are the toxic species.

A recent overview in Nature (The Plaque Plan, Nature, November 13 2008) talked about the disappointments and ambiguities facing researchers in AD. Amyloid burden does not always correlate with cognitive impairment. In addition, two big clinical trials designed to target the formation of amyloid have both ended in failure and confusion. It's back to the drawing board for many embattled scientists. A major problem inherent in testing AD therapies is the late stage at which they inevitably have to be initiated in the absence of an early test for diagnosis. Thus disease progression is already advanced and perhaps that is the reason the therapies don't work as planned. The future should be as much focused on early brain and cerebrospinal fluid imaging as on new therapies. And of course, we are still in the rudimentary stages of achieving that supreme goal of understanding how memories are formed and stored.

At any rate, the midnight oil keeps burning and the search goes one, and Ehud Gazit from Tel Aviv University has published a paper in Angewandte Chemie that features a simple dipeptide for targeting and disassembling early soluble amyloid fibrils. The dipeptide is D-Trp-Aib, consisting of D-tryptophan and the unnatural amino acid amino isobutyric acid. This amino acid is rather unique and has been extensively studied because it seems to lend pronounced alpha helicity to peptides. The running hypothesis in its inclusion in the inhibitor is that it is a ß-breaker, an amino acid that helps to break the ß sheets that are a signature of amyloid. The work also focuses on the central region of the Aß (1-42) amyloid peptide that forms its aromatic core. The peptide sequence here is KLVFFAE, and the central Phe residues are thought to strongly influence the aggregation of the peptide. Thus the use of the D-Trp; apparently it would help to interfere in aromatic assembly. In addition it would also escape degradation by ubiquitous proteases.

Being a simple uncapped dipeptide, the blood-brain barrier permeability of this molecule is admittedly lousy (It has a depressing logP value of -0.8). Strategies could probably be worked up for enhancing delivery by chemical modification. However, other tests that the authors used attest to its efficacy as a amyloid-interfering agent. The compound inhibits the oligomerization of the amyloid oligomers as shown in a SDS-PAGE gel; it inhibited oligomers that can be stabilized using SDS. Interestingly its inhibition ability decreases at intermediate concentration and then goes up again. The compound also dissociated amyloid fibrils as shown in fluorescence assays with thioflavin-T. Lastly and more importantly, it seems to demonstrate long-term potentiation (LTP) in mice that may lead to an increase in cognitive retention. The molecule also showed good bioavailability and low toxicity.

In addition the researchers also did a NMR study on the interaction of D-Trp-Aib with the core of amyloid Aß (1-42). Using TOCSY and other spectra they detected differences in the chemical shifts of alpha protons of some key amino acids. Using NOE information they have also proposed a family of solution structures representing the bound conformation of the dipeptide with the sequence. I am more skeptical about this result. Small sequences of peptides rapidly interconvert in solution and give averaged NMR signals. The binding of the peptide to the sequence is probably an on-off event in which it preferentially stabilizes a particular conformation. A more detailed structural study combined with kinetics experiments would shed light on the key binding events.

How far such therapies would take us in tackling AD in human beings remains to be seen; in any case, maybe the trials should begin soon.

Other AD-related posts: water in amyloid, amyloid dimers as possible culprits, "seminal" truths about amyloid, and amyloid as a possible window into historical pathogen wars

References:
Anat Frydman-Marom, Meirav Rechter, Irit Shefler, Yaron Bram, Deborah E. Shalev, Ehud Gazit (2008). Cognitive-Performance Recovery of Alzheimer's Disease Model Mice by Modulation of Early Soluble Amyloidal Assemblies Angewandte Chemie International Edition DOI: 10.1002/anie.200802123