Antibodies can be raised that specifically target misfolded prion proteins, leaving native proteins free to carry on their normal functions. This is the conclusion of a paper which appeared in yesterday's Nature Medicine online.
Misfolded proteins have been implicated in the etiology of a host of neurodegenerative diseases, including Alzheimer's, Parkinson's and prion protein diseases such as Creutzfeld-Jacob disease (CJD). The article, reported by Neil Cashman at the University of Toronto, together with a host of collaborators in Canada, the U.S. and the U.K., raises hopes that antibodies which recognize such proteins may someday be used to diagnose and treat these disorders.
Prion proteins usually come in two flavors, normal (cellular) and infectious. The latter tend to aggregate, suggesting that the protein has undergone a conformational change that exposes amino acids not normally found on the protein surface. With this in mind, first author Eustache Paramithiotis and colleagues asked if such conformational changes also expose new epitopes, or short stretches of amino acids that could serve as antibody binding sites. To test this hypothesis, the authors exposed cellular prion protein (PrPC) to low pH, a treatment that promotes aggregation, then looked for changes in the amino acid composition on the surface of the protein.
By measuring the fluorescence of the protein at low pH, Paramithiotis found an increase in the number of tyrosine residues that are exposed to solution. Armed with this, and with the knowledge that the majority of PrPC tyrosines exist in tandem, the authors immunized rabbits with a tyrosine-tyrosine-arginine tripeptide, which they coupled to the highly antigenic carrier, keyhole limpet hemocyanin. The authors then purified immunoglobulins from the rabbits and tested them against brain homogenates from mice suffering from the prion disease scrapie. One polyclonal antibody preparation precipitated infectious prion (PrPSc), but failed to precipitate cellular prion.
Buoyed by this proof-of-principle experiment, Paramithiotis and colleagues then generated mouse monoclonal antibodies to the tripeptide. Of 10 such antibodies tested, all of them bound specifically to PrPSc from mouse brain homogenates, but more importantly, they also bound specifically to PrPSc in human brain samples taken from sufferers of CJD and Gerstmann-Straussler-Scheinker syndrome, another prion disease. Also, in a testament to the specificity of the antibodies, the authors found that the monoclonals immunoprecipitated PrPSc, but not PrP, from infected mouse spleen, an organ known to harbor over a thousandfold more of the cellular prion.
This data comes hot on the heels of a report in Nature last March (see ARF related news story) which showed that passive immunization of mice with antibodies that recognize both cellular and infectious prion protects the animals against scrapie. The therapeutic efficacy of these new PrPSc selective antibodies remains to be determined.—Tom Fagan
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- Paramithiotis E, Pinard M, Lawton T, LaBoissiere S, Leathers VL, Zou WQ, Estey LA, Lamontagne J, Lehto MT, Kondejewski LH, Francoeur GP, Papadopoulos M, Haghighat A, Spatz SJ, Head M, Will R, Ironside J, O'Rourke K, Tonelli Q, Ledebur HC, Chakrabartty A, Cashman NR. A prion protein epitope selective for the pathologically misfolded conformation. Nat Med. 2003 Jul;9(7):893-9. PubMed.