Novel glycosaminoglycan precursors as anti-amyloid agents part II.
In vivo amyloids consist of two classes of constituents. The first is the disease defining protein, e.g., A beta in Alzheimer's disease. The second is a set of common structural components that usually are the building blocks of basement membrane (BM), a tissue structure that serves as a scaffold onto which cells normally adhere. In vitro binding interactions between one of these BM components and amyloidogenic proteins rapidly change the conformation of the amyloidogenic protein into amyloid fibrils. The offending BM component is a heparan sulfate (HS) proteoglycan (HSPG), part of which is protein and the remainder a specific linear polysaccharide, which is the portion responsible for binding, and imparting the typical amyloid structure, to the amyloid precursor protein/peptide. Our past work has demonstrated that agents that inhibit the binding between HS and the amyloid precursor are effective anti-amyloid compounds both in vitro and in vivo. The present work is concerned with the design and synthesis of modified sugar precursors of HS, which, when incorporated into the polysaccharide, will alter its structure so that it loses its amyloid precursor protein/peptide-binding and fibril-inducing properties. As part of our continuing study, since our previous report, 17 additional compounds have been designed and synthesized based primarily on the known steps involved in HS biosynthesis. In addition to the 4 reported last year, 10 more have been assessed in tissue culture for their inhibitory effect on heparan sulfate synthesis, and one of these has been assessed for its AA-amyloid inhibitory properties. The majority of the novel sugars are analogues of N-acetylglucosamine. They have been modified either at the 4-OH, 3-OH, or 2-N positions. The majority of the 2-N analogues provide data suggesting that hepatocyte N-demethylases remove the N-substituents converting the 2-N analogues into the natural sugar, a process that dilutes the D-[3H] glucosamine tracer used to track heparan sulfate synthesis and thereby gives the impression that biosynthetic inhibition is occurring. To date 3-deoxy analogues have failed to affect heparan sulfate synthesis significantly. Compounds incorporating the 3,4-dideoxy structural feature are currently being assessed. Using primary hepatocyte cultures, we reported previously that a 4-deoxy analogue is incorporated into HS and terminates its elongation. From the 4-deoxy series, one of the compounds has now been assessed in an in vivo model of AA-amyloid induction. This 4-deoxy analogue inhibited splenic AA amyloid deposition by at least 50%, and liver AA amyloid deposition by 85% when measured as amyloid/unit area of tissue. Furthermore, the spleen weights of the treated group were 1/2-1/3 of that in the untreated group indicating that the total splenic amyloid was 1/4-1/6 of that in the untreated group. The results provide further evidence that heparan sulfate is a critical factor in amyloidogenesis and modifications of sugar precursors of heparan sulfate synthesis may provide leads for therapeutic intervention in amyloidogenesis.
Kisilevsky
Robert
R
Department of Pathology, Queen's University, Kingston, Ontario, Canada. kisilevsky@cliff.path.queensu.ca
Szarek
Walter A
WA
D000544:Alzheimer Disease / Q000378:metabolism*; D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000378:metabolism*; D000818:Animals; D018929:Cell Culture Techniques; D006025:Glycosaminoglycans / Q000235:genetics / Q000378:metabolism*; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism; D006493:Heparin / Q000031:analogs & derivatives* / Q000378:metabolism; D006497:Heparan Sulfate / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011509:Proteoglycans / Q000378:metabolism; D013154:Spleen / Q000378:metabolism
D016229:Amyloid beta-Peptides; D006025:Glycosaminoglycans; D019812:Heparan Sulfate Proteoglycans; D006493:Heparin; D006497:Heparan Sulfate; D011509:Proteoglycans; C036090:heparin proteoglycan