Faculty Bibliography

We have reported that an amyloid-beta derivative, K6Abeta1-30-NH2 reduces amyloid burden in mice to a similar extent as previously shown for Abeta1-42 (Am J Pathol 159:439-47,2001). This derivative may be a safer alternative to Alzheimer's vaccination with Abeta1-42 because it has a low beta-sheet content while maintaining the main antigenic sites of Abeta. To determine the in vivo effect of other derivatives with similar in vitro properties, we immunized Tg2576 mice with Abeta1-30-NH2, in which amino acids 18 and 19 were substituted with glutamate (Abeta1-30E18E19). In a parallel study, mice were immunized with K6Abeta1-30E18E19. Freund's adjuvant was used to allow a comparison with our findings with K6Abeta1-30-NH2. Antibody titers were detectable, but much lower than we had observed for K6Abeta1-30-NH2 or Abeta1-42, indicating that the central hydrophobic region of Abeta may have an epitope important for modulating humoral response. Cognitive performance was assessed in a radial arm maze before sacrifice at 19-21 months. Control Tg mice had more errors than their wild-type littermates (p<0.01), and the Abeta1-30E18E19-treated mice (p<0.05). Mice receiving K6Abeta1-30E18E19 also performed better than their Tg controls (p<0.05). Histologically, no difference was observed in brain amyloid plaque burden in 6E10 stained brain sections from the Abeta1-30E18E19-vaccinated mice, compared to vehicle treated mice. Furthermore, amyloid burden did not correlate with cognitive performance. Analysis of plaque burden in the K6Abeta1-30E18E19-immunized mice is underway, as well as measurements of brain levels of Abeta to determine if these values will provide a better correlation with cognitive performance. A robust antibody response and a diminished plaque burden may not be necessary for a therapeutic effect of Abeta derived vaccines

The prion protein (PrP) is a copper binding protein; however, the role of copper in prion infection is unclear. Under some conditions copper facilitates refolding of denatured PrPSc into a protease resistant and infectious form. Hence copper may enhance the infectivity of the prion protein. To determine the feasibility of copper targeted therapy for prion disease, we treated mice (n=10 per group) with d-penicillamine (d-PEN; 100 mg/kg, i.p.), immediately following scrapie inoculation (139A strain, i.p.). Subsequent drug injections were daily, five days per week. d-PEN delayed the onset of prion disease in the mice (p=0.002). The effect was more pronounced at the 1000-fold dilution of agent (d-PEN=179 +- 3 days, VEH=165 +- 4, p=0.006), but a trend for a delay was observed at the 10-fold dilution (d-PEN=153 +- 2, VEH=146 +- 3, p=0.1). As expected, d-PEN reduced brain copper levels (p<0.01) by 26% (10-fold dil.; p=0.04) and 32% (1000-fold dil.; p=0.02), compared to control animals. Brain levels of iron and zinc were not reduced. To further support the notion that the therapeutic effect of d-PEN was mediated through its copper chelating properties, brain homogenates from terminally ill 139A infected mice were incubated with copper and d-PEN. Following a 72 h incubation, copper sulfate increased aggregation of the prion protein in a dose dependent manner, resulting in an enhanced resistance to proteinase K. This effect was counteracted by co-incubation with d-PEN. These findings support the proposed in vivo effect of d-PEN in delaying the onset of prion disease in these mice. Copper chelator-based therapy may benefit those incubating prion disease but this approach may be more effective at higher doses and/or in a multi-targeted combinational therapy

The outbreak of new variant Creutzfeldt-Jakob disease has raised the specter of a potentially large population being at risk to develop this prionosis. None of the prionoses currently have an effective treatment. Recently, vaccination has been shown to be effective in mouse models of another neurodegenerative condition, namely Alzheimer's disease. Here we report that vaccination with recombinant mouse prion protein delays the onset of prion disease in mice. Vaccination was performed both before peripheral prion exposure and after exposure. A delay in disease onset was seen in both groups, but was more prolonged in animals immunized before exposure. The increase in the incubation period closely correlated with the anti-prion protein antibody titer. This promising finding suggests that a similar approach may work in humans or other mammalian species at risk for prion disease

The recent termination of a Phase II clinical trial in which volunteers with Alzheimer's disease (AD) were vaccinated with Amyloid-beta (AP)1-42, has cast doubt on the feasibility of this therapeutic approach. While the exact reasons for the cerebral inflammation in these patients is being determined, it is difficult to evaluate the cause of these adverse effects. The most likely reasons are Abeta1-42 toxicity and/or autoimmunity. Abeta vaccination approaches are based on the hypothesis that Abeta deposition and toxicity are central to the pathogenesis of AD. Therefore, it is counterintuitive to use the whole Abeta peptide for human vaccination. Abeta1-40/42 is a major plaque component that forms inflammatory/toxic fibrils as observed in many in vitro and in vivo studies. Furthermore, numerous studies have shown that Abeta1-40/42 bidirectionally crosses the blood-brain barrier (BBB) in experimental animals. Additionally, in vitro and in vivo studies indicate that minute amounts of Abeta1-42 may seed fibril/amyloid formation. The elderly, a target population for AD therapy, often have a poor immune response to vaccines, which enhances the gravity of these safety concerns. In these patients with an attenuated immune reaction, injected Abeta1-42 may initiate and/or enhance congophilic angiopathy, which eventually may result in reduced cerebral blood flow and/or intracerebral bleeding. Abeta1-42 may also cross the BBB and once within the brain parenchyma it may contribute to plaque formation and/or co-deposit on plaques. Together, these effects within blood vessels and/or brain parenchyma may actually enhance the progression of AD. Given the potential serious side effects of Abeta1-42 vaccination, it is safer to use immunogenic Abeta derivatives, which are less likely to be toxic. The main immunogenic epitopes of Abeta1-42 are contained within the first 30 amino acids of the peptide. Taking this into account, we have developed soluble antigenic Abeta derivatives, which are nonfibrillogenic and nontoxic in human cell culture. Our prototype peptide, K6Abeta1-30-NHz, diminishes amyloid burden to a similar extent as reported for Abeta1-42. Additionally, ramified IL-1beta-positive microglia as well as phagocytes, associated with the Abeta plaques, were absent in the immunized mice, indicating reduced inflammation in these animals at the time point examined. Autoimmunity may be the culprit if follow-up studies reveal that the brain inflammation is related to antibody interactions with AD and/or amyloid precursor protein (APP). In such a scenario, any vaccination approach targeting A(3 can have similar consequences, although preventive treatment initiated prior to amyloid deposition may not result in these adverse reactions. T-cell-related autoimmunity may also be involved and can be expected to be less with Abeta derivatives not containing certain T-cell epitopes. An alternative to the active vaccination approach is passive immunization, which is associated with a lower risk of irreversible autoimmunity. This approach may also be used in patients with a muted immune response to the vaccine. However, in a chronic disease such as AD repeated antibody injections may lead to an anti-idiotype response and the resulting serum immune complexes can cause vasculitis and/or glomerulonephritis. Reduction of soluble Abeta within the peripheral system may be a critical part of the pathway that reduces cerebral plaque burden in Tg mice and ultimately in AD patients. Overall, the use of nontoxic A(3 derivatives and/or antibodies with very limited access into the CNS, such as IgM, may prove to have reduced si Any therapeutic approach will be more effective when used prophylactically because of neuronal loss and increased amyloid burden in the later stages of AD. Reversal of clinical symptoms cannot be expected and early diagnosis of AD may be needed for effective therapy. (C) 2002 Wiley-Liss, Inc

Recent reports indicate that amyloid-beta (Abeta) vaccine-based therapy for Alzheimer's disease (AD) may be on the horizon. There are, however, concerns about the safety of this approach. Immunization with Abeta1-42 may not be appropriate in humans because it crosses the blood-brain barrier, can seed fibril formation, and is highly fibrillogenic. Abeta1-42 fibrils can in turn cause inflammation and neurotoxicity. This issue is of a particular concern in the elderly who often do not mount an adequate immune response to vaccines. Our findings show that vaccination with nonamyloidogenic/nontoxic Abeta derivative may be a safer therapeutic approach to impede the progression of Abeta-related histopathology in AD. Although the site of action of the anti-Abeta antibodies has been suggested to be within the brain, peripheral clearance of Abeta may have a greater role in reducing cerebral amyloid plaques in these animals and eventually in AD patients. Antibodies in general are predominantly found outside the central nervous system (CNS) and will, therefore, primarily clear systemic Abeta compared to brain Abeta. This disruption of the equilibrium between central and peripheral Abeta should then result in efflux of Abeta out of the brain, and subsequent removal of plaques. Abeta therapy can be targeted to the periphery, which may result in fewer CNS side effects, such as inflammation. Future Abeta derived vaccines should include T(h) epitopes, carriers and/or lipid moieties to enhance antibody production in the elderly, the population predominantly affected by AD

To date, transmissibility of amyloid diseases has not been thoroughly investigated. Although only some of these conformational disorders are considered infectious, all amyloid diseases could be infectious under certain conditions. For transmissibility, endogenous expression of an amyloidogenic peptide required, as well as the presence of an inoculum that is rich in amyloid fibrils and/or their precursors. Notably, administration of one type of amyloid might result in deposition of a different amyloid. Various cofactors could be essential for transmission - some might chaperone the amyloid peptides and/or fibrils, thereby directly facilitating their propagation; others might indirectly stabilize and/or increase levels of conformers with a high beta-sheet content. It is possible that these chaperones are induced by inflammation, which itself can lead to secondary amyloidosis. Thus, amyloid-related therapeutic approaches should not be based on administration of amyloidogenic peptides in conjunction with an inflammatory stimulus, such as in a recently halted clinical trial for Alzheimer's disease