The Anti-prion Activity of Congo Red

175

225

Prion proteins are key molecules in transmissible spongiform encephalopathies (TSEs), but the precise mechanism of the conversion from the cellular form (PrP C ) to the scrapie form (PrP Sc ) is still unknown. Here we discovered a chemical chaperone to stabilize the PrP C conformation and identified the hot spots to stop the pathogenic conversion. We conducted in silico screening to find compounds that fitted into a ''pocket'' created by residues undergoing the conformational rearrangements between the native and the sparsely populated high-energy states (PrP*) and that directly bind to those residues. Forty-four selected compounds were tested in a TSE-infected cell culture model, among which one, 2-pyrrolidin-1-yl-N-[4-[4-(2-pyrrolidin-1-yl-acetylamino)-benzyl]-phenyl]-acetamide, termed GN8, efficiently reduced PrP Sc . Subsequently, administration of GN8 was found to prolong the survival of TSE-infected mice. Heteronuclear NMR and computer simulation showed that the specific binding sites are the A-S2 loop (N159) and the region from helix B (V189, T192, and K194) to B-C loop (E196), indicating that the intercalation of these distant regions (hot spots) hampers the pathogenic conversion process. Dynamics-based drug discovery strategy, demonstrated here focusing on the hot spots of PrP C , will open the way to the development of novel anti-prion drugs.anti-prion compound ͉ binding sites ͉ chemical chaperone ͉ dynamicsbased drug discovery ͉ transmissible spongiform encephalopathy

Section: Discussionmentioning

confidence: 99%

Hot spots in prion protein for pathogenic conversion

Kuwata

Nishida

Matsumoto

et al. 2007

175

225

“…Several mechanisms may lead to PrP Sc depletion: prevention of PrP C to PrP Sc conversion by stabilization of PrP C , interference with the interaction of PrP C and PrP Sc (36), or sequestration and/or reversion of PrP Sc to a protease-sensitive state (33). In addition, abrogation of PrP C synthesis or prevention of transport to the cell surface could interrupt prion propagation.…”

Section: Discussionmentioning

confidence: 99%

Scrapie prion protein accumulation by scrapie-infected neuroblastoma cells abrogated by exposure to a prion protein antibody

Enari

Flechsig²,

Weissmann³

2001

402

312

Exposure of susceptible neuroblastoma N2a cells to mouse scrapie prions leads to infection, as evidenced by the continued presence of the scrapie form of the prion protein (PrP Sc ) and infectivity after 300 or more cell doublings. We find that exposure to phosphatidylinositol-specific phospholipase C (PIPLC) or to the monoclonal anti-prion protein (PrP) antibody 6H4 not only prevents infection of susceptible N2a cells but also cures chronically scrapie-infected cultures, as judged by the long-term abrogation of PrP Sc accumulation after cessation of treatment. A nonpassaged, stationary infected culture rapidly loses PrP Sc when exposed to the antibody or PIPLC, indicating that the PrP Sc level is determined by steady state equilibrium between formation and degradation, and that depletion of the cellular form of PrP can interrupt the propagation of PrP Sc . These findings encourage the belief that passive immunization may provide a therapeutic approach to prion disease.

“…Values were expressed as percentage of signal intensity of samples nontreated with tetracyclines, and the significance of difference was assessed by Dunnett's test. Control experiments included (i) the incubation of samples with 1 M to 1 mM gentamicin instead of tetracycline compounds, (ii) the addition of tetracycline or doxycycline to the samples immediately before proteinase K digestion, and (iii) the treatment of proteinase K-digested samples with 3 M guanidine isothiocyanate, pH 2.5, for 10 min, followed by the addition of BSA to a final concentration of 200 g͞ml and methanol precipitation, before SDS͞PAGE and immunoblot (34).…”

Section: In Vitro Conversion Of Protease-resistant Prp To a Protease-mentioning

confidence: 99%

Tetracyclines affect prion infectivity

Forloni¹,

Iussich²,

Awan³

et al. 2002

179

102

Prion diseases are transmissible neurodegenerative disorders of humans and animals for which no effective treatment is available. Conformationally altered, protease-resistant forms of the prion protein (PrP) termed PrP Sc are critical for disease transmissibility and pathogenesis, thus representing a primary target for therapeutic strategies. Based on previous findings that tetracyclines revert abnormal physicochemical properties and abolish neurotoxicity of PrP peptides in vitro, we tested the ability of these compounds to interact with PrP Sc from patients with the new variant of Creutzfeldt-Jakob disease (vCJD) and cattle with bovine spongiform encephalopathy (BSE). The incubation with tetracycline hydrochloride or doxycycline hyclate at concentrations ranging from 10 M to 1 mM resulted in a dose-dependent decrease in protease resistance of PrP Sc . This finding prompted us to investigate whether tetracyclines affect prion infectivity by using an animal model of disease. Syrian hamsters were injected intracerebrally with 263K scrapie-infected brain homogenate that was coincubated with 1 mM tetracycline hydrochloride, 1 mM doxycycline hyclate, or vehicle solution before inoculation. Hamsters injected with tetracycline-treated inoculum showed a significant delay in the onset of clinical signs of disease and prolonged survival time. These effects were paralleled by a delay in the appearance of magnetic-resonance abnormalities in the thalamus, neuropathological changes, and PrP Sc accumulation. When tetracycline was preincubated with highly diluted scrapie-infected inoculum, one third of hamsters did not develop disease. Our data suggest that these well characterized antibiotics reduce prion infectivity through a direct interaction with PrP Sc and are potentially useful for inactivation of BSE-or vCJD-contaminated products and prevention strategies.