For the past decade, Alzheimer researchers have gradually built the argument that small species of the amyloid-β peptide might be harming neurons in ways quite separate from the damage done by fibrillized forms. In short, their slogan is, “It’s Not the Plaques, Stupid!” This research addresses a lingering disconnect between the distribution of plaque pathology and that of cognitive dysfunction in AD, but it has been vexed by the problem that Aβ oligomers were maddeningly elusive. After all, what’s not physically in hand or visible to the eye won’t easily persuade skeptical colleagues. A visit to this year’s 35th Annual Conference of the Society for Neuroscience, held November 12-16 in Washington, D.C., however, made clear that this field is moving rapidly. While there are no breakthroughs to report, the large number of presentations on Aβ oligomers and the frequency with which they come up in conversation with scientists make clear that oligomeric Aβ, also called ADDLs, has become the object of intense investigation.
A major difficulty has been the challenge of obtaining sufficiently pure forms of Aβ for quantitative study. By now, several labs have developed better isolation protocols and tools to characterize various forms of Aβ oligomer. It’s too soon to offer a general comparison between them, and an undisputed lead contender has not yet emerged either for the title of most important pathogenic form, or most attractive therapeutic lead. Yet this area has produced a number of intriguing candidates worth knowing about, and academic labs as well as biotechnology and pharmaceutical companies are exploring experimental vaccines against “their” oligomers. Below are selected summaries of a few such studies. Readers who were at the conference are cordially invited to share their notes with the Alzforum community and write comments on presentations missing from this report.
A group not previously covered in this news section that pursues Aβ oligomer research is Heinz Hillen’s and Ulrich Ebert’s at Abbott GmbH in Ludwigshafen, Germany. Stefan Barghorn presented results of a stable Aβ42 oligomer, which this group calls “globulomer” and believes to be pathogenic. This globulomer joins diffusible oligomers called ADDLs first discovered by Bill Klein (Lambert et al., 1998), and secreted cell-based oligomers studied by Dominic Walsh and Dennis Selkoe (Podlisny et al., 1998) in a growing group of suspects for cognitive deficits in AD. (See also section on Ashe below.) The Abbott poster describes how Hillen and colleagues generated a defined oligomer from synthetic Aβ and fatty acids, and then assessed its possible relevance in vivo. They came up with a 60 kD ball of 12 Aβ molecules in which the middle amino acid stretches (20-42) turn inward to form a hydrophobic ball and the N-termini stick out. Monoclonal (8F5) and polyclonal (5598) antibodies generated against the globulomer bind a structural epitope on it, but not APP, Aβ monomers, or fibrils. The monoclonal antibody stains plaques in sections of human AD and mice overexpressing mutant APP (Tg2576). The scientists report that they have detected the Aβ globulomer epitope in the soluble fraction of the cortex of three AD patients and of Tg2576 mice, but that it is absent from human CSF. Similar to anti-ADDL antibodies, the globulomer antibody binds to processes of cultured hippocampal neurons, but not to glia. This work appeared in print last month (see Barghorn et al., 2005).
Looking for a functional effect of this preparation in brain proved tricky when it turned out that the globulomer does not penetrate tissue well, probably because it is too sticky to diffuse far beyond the site of injection. However, when applied instead to rat brain slices, the globulomer completely blocked long-term potentiation, the scientists reported. Ebert said his group has no evidence that the 60 kD globulomer forms in the human AD brain exactly as the 12mer proposed on the poster. After all, the ingredients of synthetic Aβ and fatty acids sound a bit like a recipe for detergent micelles. Instead, Ebert and colleagues propose that in a person’s aging brain, Aβ might oligomerize only part of the way toward the full globulomer, and perhaps does so in response to intrinsic or environmental risk factors, such as age- or diet-related alterations in membrane composition.
Ebert and Barghorn emphasize two points of their study. First, the structural epitope provides a starting point for immunotherapy. Abbott is pursuing an immunotherapy approach about which the scientists stayed mum. Second, the globulomer is not a way station on a stepwise polymerization of monomer-dimer-oligomer-protofibril-fibril. Instead, they associate on a separate track, of which the globulomer would be a stable product. Some unknown factor may cause a conformational switch that drives Aβ down this pathway, Ebert said.
If pathogenic Aβ oligomers are more than unstable transition states on the way to fibrillization, then it should be possible to isolate them. Indeed, some researchers are beginning to do just that. Karen Hsiao Ashe at the University of Minnesota Medical School, Minneapolis, described the isolation of a pathogenic12mer of Aβ her lab calls Aβ*. At a satellite symposium held by the Alzheimer Research Consortium the day before the SfN conference itself, Ashe described this work as part of her group’s longstanding effort to identify forms of Aβ and tau that cause the cognitive deficit in AD, which Ashe believes to be separate from the deficit that follows structural damage, that is, plaques, tangles, dystrophic neurites, gliosis, and synapse and neuronal loss. To that end, Ashe’s group tried to isolate the molecule that causes the behavioral deficits described in the Tg2576 mice she generated in 1996. Setting a strategy that evokes Robert Koch’s postulates of criteria an agent must fulfill to be considered the cause of a disease, Ashe told the audience she aimed to first identify the causative molecule, then purify it out of brain of impaired mice, and then inoculate young, healthy animals to see if it prompts the memory deficit.
To do this, the researchers first characterized the behavioral deficit of the Tg2576 mice. The mice’s performance declined at 6 months of age to a stable level and then dipped down further at 15 months. The scientists decided to isolate a form of Aβ that tracks this performance and, therefore, searched for one whose level rises at 6 months and then stays unchanged for about 8 months. This ruled out all published species of Aβ, including ADDLs, whose levels rise in this period and do not correlate with cognitive performance, said Ashe. For a while, the project was stuck. But in 2002, Sylvain Lesné joined the lab. He developed a fractionation procedure that pulled, out of a soluble fraction enriched for extracellular components, a highly stable 12mer whose time course over the mice’s lifespan fit the bill. It is a 56 kD complex, confirmed by mass spectrometry master Austin Yang at University of Southern California (see ARF SfN story on tau) to contain a form of Aβ. For the third step of testing its effect in healthy animals, Ashe collaborated with Michela Gallagher at Johns Hopkins University in Baltimore, Maryland, a noted rat behavioral neurobiologist. Full results are not available yet but, in short, injection of Aβ*56 purified from impaired Tg2576 mice into young healthy rats left memory acquisition unchanged but blocked memory retention, Ashe reported. Ongoing steps include a project asking whether the presence of this substance distinguished between people who died with AD from those who died without the disease, and a project focusing on the interaction of Aβ*56 with its receptor.
“This is the first time that an agent that disrupts memory has been purified from the brain of an AD model,” Ashe said. “Because it is the agent that causes the cognitive deficit, this brings us a step closer to develop antibodies that could be specifically used as a diagnostic,” she added. This presentation raised intense interest among the audience. Among the follow-up questions was one about what might make the concentration of one species of Aβ remain stable over 10 months while most other species increase. This work is now in press at Nature. —Gabrielle Strobel.
See also Part 2 of this news update.
- SfN: Return of the Other—Tau Is Back, Part 3
- SfN: Amyloid Oligomers—Not So Elusive, After All? Part 2
- Lambert MP, Barlow AK, Chromy BA, Edwards C, Freed R, Liosatos M, Morgan TE, Rozovsky I, Trommer B, Viola KL, Wals P, Zhang C, Finch CE, Krafft GA, Klein WL. Diffusible, nonfibrillar ligands derived from Abeta1-42 are potent central nervous system neurotoxins. Proc Natl Acad Sci U S A. 1998 May 26;95(11):6448-53. PubMed.
- Podlisny MB, Walsh DM, Amarante P, Ostaszewski BL, Stimson ER, Maggio JE, Teplow DB, Selkoe DJ. Oligomerization of endogenous and synthetic amyloid beta-protein at nanomolar levels in cell culture and stabilization of monomer by Congo red. Biochemistry. 1998 Mar 17;37(11):3602-11. PubMed.
- Barghorn S, Nimmrich V, Striebinger A, Krantz C, Keller P, Janson B, Bahr M, Schmidt M, Bitner RS, Harlan J, Barlow E, Ebert U, Hillen H. Globular amyloid beta-peptide oligomer - a homogenous and stable neuropathological protein in Alzheimer's disease. J Neurochem. 2005 Nov;95(3):834-47. PubMed.
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