A variety of approaches are beginning to home in on Aβ oligomers as drug targets, and some of them were showcased at the International Conference on Alzheimer’s Disease held 11-16 July in Vienna, Austria. Take Dieter Willbold’s D3 peptide, for example. Willbold is heading institutes at the University of Düsseldorf and the Forschungszentrum Jülich, near Düsseldorf, Germany (see ARF companion story) on oligomer/aggregate detection). In Vienna, he introduced the field to a stable peptide that turns toxic oligomers into an apparently innocuous non-amyloid material and appears to work in mouse models.

Called D3, the peptide came out of a method called mirror-image phage display. In essence, this is an elegant way of obtaining D-enantiomeric peptides that bind to a target of choice, in this case Aβ oligomers. D-peptides are chiral opposites of the more common L-peptides. They appeal to drug developers because unlike L-peptides, they withstand protease degradation in the blood, stomach, and intestine. With mirror-image phage display, scientists screen a large library of phages displaying L-peptides with the D- version of their favorite drug target, and then synthesizes a mirror image of the screen’s best hit. That nets a D-peptide sticking to the L version of the target (for a detailed description of this method and other examples of its use in medical research, see free review).

Willbold’s institutional website features a concise description of two of the peptides obtained in this way. At ICAD, Willbold focused on the 12-mer D3. He first showed using a thioflavin T assay that D3 inhibits Aβ42 fibril formation in vitro. Then he showed that increasing doses of it restored cell viability in the presence of otherwise toxic Aβ42 using an MTT toxicity assay (Shearman et al., 1994). In mice expressing both the APPswe and PS1DE9 mutations, D3 infused directly into the brain with a micropump, or dripped into the drinking water, reduced amyloid deposits in the brain (van Groen et al., 2008) and restored a water maze learning impairment in these mice.

Willbold wondered, How might this work? To address this question, the Jülich team then performed size exclusion chromatography as described previously by Lars Lannfelt and colleagues at Uppsala University in Sweden (Johannson et al., 2006). Lannfelt’s group has poured much effort into optimizing biochemical methods to separate different forms of Aβ in human samples. When Willbold’s team compared the fractions that came off a size exclusion column loaded with either an Aβ sample or the same Aβ sample with D3, the scientists noticed that in the latter, the protofibril fraction was gone. But the monomer fraction was no bigger. “So where was the Aβ? We checked the centrifugation pellet, and there it was,” Willbold told a reporter. The D3 peptide had precipitated the oligomers/protofibrils. With density gradient centrifugation, the result was similar. This method separates Aβ species in a sample by size, and they can then be lined up on SDS PAGE. On the PAGE, the Aβ/D3 sample was missing the middle fractions representing oligomers/protofibrils; most of the Aβ showed up in the highest-molecular-weight fractions.

Precipitation of Aβ in the brain would sound alarming to anyone even vaguely aware of AD research, but this material is different, Willbold emphasized. It is not an amyloid. Besides being non-toxic to cells and the mice, at least, it does not bind thioflavin T, nor does it form fibers or fibrils in the electron microscope, Willbold reported. In a dynamic light-scattering experiment (a biophysical method to measure the size distribution of particles in a given sample), Aβ stayed at one constant size, whereas the Aβ/D3 sample separated after about seven minutes into larger particles of varying sizes. Aileen Funke, a group leader in Willbold's lab, added that in an in-vitro seeding assay, the Aβ-D3 complex was unable to seed fibrillization. In light of these findings, the D3 peptide cannot be considered an example of what were previously dubbed “plaque busters,” Willbold said. Some of these compounds have given rise to products that are themselves either toxic and/or amyloidogenic. “We do not know exactly yet what D3 does to the protofibrils, but the resulting complex is neither toxic nor amyloidogenic,” Willbold said.

This is not the only study targeting oligomers with the help of D-amino acids. At the AD/PD conference last March in Prague, Anat Frydman-Marom won a Young Investigator Award for her work on a β-sheet breaking di-peptide that does much the same thing to Aβ oligomers in vitro and in an APP transgenic mouse. According to a recent article, this di-peptide comprises a D-isomer and a non-chiral amino acid, and behaves like a safe, orally available small-molecule drug (see Frydman-Marom et al., 2009).—Gabrielle Strobel.


No Available Comments

Make a Comment

To make a comment you must login or register.


News Citations

  1. Vienna: New Tack to See Amyloid Oligomers in Body Fluids

Paper Citations

  1. . Inhibition of PC12 cell redox activity is a specific, early indicator of the mechanism of beta-amyloid-mediated cell death. Proc Natl Acad Sci U S A. 1994 Feb 15;91(4):1470-4. PubMed.
  2. . Reduction of Alzheimer's disease amyloid plaque load in transgenic mice by D3, A D-enantiomeric peptide identified by mirror image phage display. ChemMedChem. 2008 Dec;3(12):1848-52. PubMed.
  3. . Physiochemical characterization of the Alzheimer's disease-related peptides A beta 1-42Arctic and A beta 1-42wt. FEBS J. 2006 Jun;273(12):2618-30. PubMed.
  4. . Cognitive-performance recovery of Alzheimer's disease model mice by modulation of early soluble amyloidal assemblies. Angew Chem Int Ed Engl. 2009;48(11):1981-6. PubMed.

External Citations

  1. free review
  2. institutional website

Further Reading