Femtomole immunodetection of synthetic and endogenous amyloid-β oligomers and its application to Alzheimer’s disease drug candidate screening
Alzheimer’s disease (AD) is a fatal, progressive dementia for which there is no cure and for which a molecular basis has yet to be established. However, considerable evidence suggests that AD is linked to neurotoxic assemblies of the 42-amino-acid peptide amyloid β (Aβ). There is now a clear body of evidence that shows this neurotoxicity resides not only in insoluble fibrils of Aβ but also in soluble Aβ ADDLs (Aβ-derived diffusible ligands) and larger protofibrils. Further, anti-Aβ antibodies have been reported to reverse memory failure in human amyloid precursor protein (hAPP)-expressed transgenic mice in a manner that suggests symptom reversal is attributable to targeting of ADDLs. Clearly, a search for drugs targeting the assembly of these soluble Aβ species represents a new and potentially important approach to the treatment of AD. In this work we describe the development of a dot-blot immunoassay to measure ADDL at the femtomole level, its use in defining the time course of ADDL formation, and its use in determining the presence of ADDLs in the hAPP transgenic mouse brain. Discussion of a protocol to screen agents for inhibition of neurotoxic ADDL formation both in vivo and in vitro is also presented. The methods are suitable for screening combinatorial libraries and, importantly, provide the potential for simultaneous information on candidate transport across the blood-brain barrier.
Index EntriesAmyloid-β Alzheimer’s disease oligomers cyclodextrin drug screening ADDLs
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- Podlisny M. B., Walsh D. M., Amarante P., Ostaszewski B. L., Stimson E. R., Maggio J. E., et al. (1998) Oligomerization of endogenous and synthetic amyloid beta-protein at nanomolar levels in cell culture and stabilization of monomer by Congo red. Biochemistry 37, 3602–3611.PubMedCrossRefGoogle Scholar
- Small G. W., Rabins P. V., Barry P. P., Buckholtz N. S., DeKosky S. T., Ferris S. H., et al. (1997) Diagnosis and treatment of Alzheimer disease and related disorders. Consensus statement of the American Association for Geriatric Psychiatry, the Alzheimer’s Association, and the American Geriatrics Society. JAMA 278, 1363–1371.PubMedCrossRefGoogle Scholar
- Terry R. D., ed. (1999) The neuropathology of Alzheimer disease and the structural basis of its cognitive alterations, in Alzheimer Disease, Terry, Lippincott Williams & Wilkins, Philadelphia, PA, pp. 187–206.Google Scholar
- Walsh D. M., Lomakin A., Benedek G. B., Condron M. M., and Teplow D. B. (1997) Amyloid beta-protein fibrillogenesis. Detection of a protofibrillar intermediate. J. Biol. Chem. 272, 22,364–22,372.Google Scholar