flupirtine — not significantly effective (http://www.neurology.org/content/70/15/1272.short)
quinacrine — not effective in human trial (http://www.mrc.ac.uk/Newspublications/News/MRC005691)


“Soluble dimeric prion protein binds PrP(Sc) in vivo and antagonizes prion disease,” Meier et al., Cell (2003)


Conversion of cellular prion protein (PrP(C)) into a pathological conformer (PrP(Sc)) is thought to be promoted by PrP(Sc) in a poorly understood process. Here, we report that in wild-type mice, the expression of PrP(C) rendered soluble and dimeric by fusion to immunoglobulin Fcgamma (PrP-Fc(2)) delays PrP(Sc) accumulation, agent replication, and onset of disease following inoculation with infective prions. In infected PrP-expressing brains, PrP-Fc(2) relocates to lipid rafts and associates with PrP(Sc) without acquiring protease resistance, indicating that PrP-Fc(2) resists conversion. Accordingly, mice expressing PrP-Fc(2) but lacking endogenous PrP(C) are resistant to scrapie, do not accumulate PrP-Fc(2)(Sc), and do not transmit disease to others. These results indicate that various PrP isoforms engage in a complex in vivo, whose distortion by PrP-Fc(2) affects prion propagation and scrapie pathogenesis. The unique properties of PrP-Fc(2) suggest that soluble PrP derivatives may represent a new class of prion replication antagonists.


“Antiprion properties of prion protein-derived cell-penetrating peptides,” Lofgren et al., FASEB J (2008)


In prion diseases, the cellular prion protein (PrP(C)) becomes misfolded into the pathogenic scrapie isoform (PrP(Sc)) responsible for prion infectivity. We show here that peptides derived from the prion protein N terminus have potent antiprion effects. These peptides are composed of a hydrophobic sequence followed by a basic segment. They are known to have cell-penetrating ability like regular cell-penetrating peptides (CPPs), short peptides that can penetrate cellular membranes. Healthy (GT1-1) and scrapie-infected (ScGT1-1) mouse neuronal hypothalamic cells were treated with various CPPs, including the prion protein-derived CPPs. Lysates were analyzed for altered protein levels of PrP(C) or PrP(Sc). Treatment with the prion protein-derived CPPs mouse mPrP(1-28) or bovine bPrP(1-30) significantly reduced PrP(Sc) levels in prion-infected cells but had no effect on PrP(C) levels in noninfected cells. Further, presence of prion protein-derived CPPs significantly prolonged the time before infection was manifested when infecting GT1-1 cells with scrapie. Treatment with other CPPs (penetratin, transportan-10, or poly-L-arginine) or prion protein-derived peptides lacking CPP function (mPrP(23-28,) mPrP(19-30,) or mPrP(23-50)) had no effect on PrP(Sc) levels. The results suggest a mechanism by which the signal sequence guides the prion protein-derived CPP into a cellular compartment, where the basic segment binds specifically to PrP(Sc) and disables formation of prions.


“Glimepiride reduces the expression of PrP(c), prevents PrP(Sc) formation and protects against prion mediated neurotoxicity,” Bate et al., PLoS One (2009)


Treatment with glimepiride, a sulphonylurea approved for the treatment of diabetes mellitus, induced the release of PrPC from the surface of prion-infected neuronal cells. The cell surface is a site where PrPC molecules may be converted to PrPSc and glimepiride treatment reduced PrPSc formation in three prion infected neuronal cell lines (ScN2a, SMB and ScGT1 cells). Glimepiride also protected cortical and hippocampal neurones against the toxic effects of the prion-derived peptide PrP82–146. Glimepiride treatment significantly reduce both the amount of PrP82–146 that bound to neurones and PrP82–146 induced activation of cytoplasmic phospholipase A2 (cPLA2) and the production of prostaglandin E2 that is associated with neuronal injury in prion diseases. Our results are consistent with reports that glimepiride activates an endogenous glycosylphosphatidylinositol (GPI)-phospholipase C which reduced PrPC expression at the surface of neuronal cells. The effects of glimepiride were reproduced by treatment of cells with phosphatidylinositol-phospholipase C (PI-PLC) and were reversed by co-incubation with p-chloromercuriphenylsulphonate, an inhibitor of endogenous GPI-PLC.


Collectively, these results indicate that glimepiride may be a novel treatment to reduce PrPSc formation and neuronal damage in prion diseases.

The precise cellular location in which PrPC is converted to PrPSc remains controversial with advocates for the endosomal recycling compartment [48], [49]. However, anti-PrP antibodies reduced PrPSc formation, suggesting that conversion occurs either at the cell surface, or after PrPC has been internalised from the cell surface [13], [15], [50]. Such observations indicate that surface expression of PrPC is a prerequisite for PrPSc formation and that glimepiride reduced PrPSc formation by shedding PrPC from the cell surface. Our finding that glimepiride treatment released PrP molecules from ScGT1 cells raised concerns that glimepiride might cause the release of PrPSc and facilitate its spread throughout the brain. However, all PrP released from cells within 1 hour of treatment was sensitive to digestion with proteinase K. Although the presence of proteinase K sensitive PrPSc is well documented [51], the released PrP was also sensitive to digestion with thermolysin which has been reported to digest PrPC but not PrPSc [29]. Collectively these results indicate that the PrP released from ScGT1 cells was PrPC. The longer term effects of glimepiride treatment showed that twice daily treatment for 7 days caused a dose-dependent reduction in the PrPSc content of ScGT1, ScN2a and SMB cells. It also reduced the amount of PrPSc released into supernatants over this period, excluding the possibility that the reduction in cell-associated PrPSc was due to glimepiride induced the release of PrPSc from cells. Our findings are consistent with the hypothesis that glimepiride acts by limiting the supply of PrPC to cellular sites that are essential for PrPSc formation.

These results suggest that PrPSc is formed from a subset of PrPC molecules that recycle to and from the cell surface. Perhaps more significantly they indicate that the intracellular PrPC molecules were poor substrates for conversion to PrPSc. In addition, the repopulation of surface PrPC in glimepiride treated cells was from newly synthesised PrPC rather than from the intracellular pool, as it was delayed by the inclusion of the protein synthesis inhibitor cycloheximide. We conclude that there are at least 2 pools of PrPC: one that consists of PrPC molecules that recycle to and from the cell surface and are susceptible to conversion to PrPSc and another pool of PrPC molecules that are mostly intracellular and are not readily converted to PrPSc suggesting that they follow different trafficking pathways. Such results are consistent with reports that altering the trafficking of PrPC alters PrPSc formation [52], [53].

The unregulated activation of PLA2 is recognized as a key event in some neurodegenerative diseases [41], [42] and is the first step in the production of eicosanoids, docosanoids and platelet activating factors, high concentrations of which can cause glial activation, synapse damage and neuronal death. Furthermore, PLA2 plays a critical role in neurotoxicity caused by PrP peptides [55]. Our experiments showed that PrP82–146 activated cPLA2 in neurones and increased PGE2 production, a marker of PLA2 activation that is increased in scrapie infected mice [56], [57] and in the cerebrospinal fluid of patients with CJD [58], [59]. Pre-treatment with glimepiride significantly reduced PrP82–146 induced activation of cPLA2 and the production of PGE2. Although the precise mechanism is not clear, the activation of cPLA2 occurs in cholesterol-sensitive lipid rafts [60], [61]. Since glimepiride induced digestion of GPI anchored proteins affects membrane cholesterol in adipocytes [62] our results are consistent with the hypothesis that glimepiride modifies lipid rafts required for cPLA2 activation. Other studies suggest that the neurotoxicity of PrP peptides is through the amplication of PrPC associated signalling pathways [63] which may be downregulated following glimepiride treatment.

Prion infection increased cholesterol in cell membranes [32]. Since the insulin receptor is found within lipid rafts [64] and insulin signalling is cholesterol dependent [65], [66], prion infection induced changes in cell cholesterol may modify insulin signalling. This is consistent with observations that prion infection affects insulin and insulin-like growth factor receptors in cell lines [67], [68] and that scrapie infection induced diabetes mellitus in hamsters is directly damaging the central nervous system, without affecting the pancreas [69]. Thus, glimepiride treatment may also reverse prion-induced effects on insulin signalling.


“2-Aminothiazoles as therapeutic leads for prion disease,” Gallardo-Godoy et. al., Journal of Medical Chemistry (2011)

2-Aminothiazoles are a new class of small molecules with antiprion activity in prion-infected neuroblastoma cell lines (J. Virol. 2010, 84, 3408). We report here structure-activity studies undertaken to improve the potency and physiochemical properties of 2-aminothiazoles, with a particular emphasis on achieving and sustaining high drug concentrations in the brain. The results of this effort include the generation of informative structure-activity relationships (SAR) and the identification of lead compounds that are orally absorbed and achieve high brain concentrations in animals. The new aminothiazole analogue (5-methylpyridin-2-yl)-[4-(3-phenylisoxazol-5-yl)-thiazol-2-yl]-amine (27), for example, exhibited an EC(50) of 0.94 μM in prion-infected neuroblastoma cells (ScN2a-cl3) and reached a concentration of ∼25 μM in the brains of mice following three days of oral administration in a rodent liquid diet. The studies described herein suggest 2-aminothiazoles as promising new leads in the search for effective therapeutics for prion diseases.


“Styryl based and tricyclic coumpounds as potential anti-prion agents,” Chung et. al., Plos One (2011)

We used a well characterized tissue culture model of prion infection, where mouse neuroblastoma cells (N2a) were infected with 22L PrPSc, to screen compounds for anti-prion activity. In a prior study we designed a library of styryl based, potential imaging compounds which were selected for high affinity binding to Alzheimer’s disease β-amyloid plaques and good blood-brain barrier permeability. In the current study we screened this library for activity in the N2a/22L tissue culture system. We also tested the anti-prion activity of two clinically used drugs, trimipramine and fluphenazine, in the N2a/22L system. These were selected based on their structural similarity to quinacrine, which was previously reported to have anti-prion activity. All the compounds were also screened for toxicity in tissue culture and their ability to disaggregate amyloid fibrils composed of PrP and β-amyloid synthetic peptides in vitro. Two of the imaging agents, 23I and 59, were found to be both effective at inhibiting prion infection in N2a/22L tissue culture and to be non-toxic. These two compounds, as well as trimipramine and fluphenazine were evaluated in vivo using wild-type CD-1 mice infected peripherally with 139A PrPSc. All four agents significantly prolonged the asymptomatic incubation period of prion infection (p<0.0001 log-rank test), as well as significantly reducing the degree of spongiform change, astrocytosis and PrPSc levels in the brains of treated mice. These four compounds can be considered, with further development, as candidates for prion therapy.


“Unswitched immunoglobulin M response prolongs mouse survival in prion disease,” Tayebi et. al., Journal of General Virology (2009)

At present, there is no effective therapy for any of the neurodegenerative amyloidoses, despite renewed efforts to identify compounds active against the various implicated pathogenetic molecules. We have screened a library of 2960 natural and synthetic compounds in two cell lines chronically infected with mouse prions, and have identified eight new inhibitors of prion replication in vitro. They belong to two distinct chemical families that have not previously been recognised as effective in the field of transmissible spongiform encephalopathies: seven are 3-aminosteroids and one is a derivative of erythromycin A with an oxime functionality. Our results suggest that these aminosteroids inhibit prion replication by triggering a common target, possibly implicated in the regulatory pathways of cellular prion protein metabolism. Furthermore, using a quantitative approach for the study of protein stability, it was shown that the erythromycin A derivative altered prion protein stability by direct interaction. Such direct targeting of this amyloid precursor might provide new clues for the understanding of prion diseases and, more importantly, help to define new molecules that are active against prion diseases.


“ST1859 reduces prion infectivity and increases survival in experimental scrapie,” Columbo et. al., Archives of Virology

On the basis of the structural homologies between ST1859 (1[(2-hydroxy-1-naphtyl)methyl]-2-naphthol) and the anti-prion agents and its anti-amyloidogenic activity, we tested whether this molecule altered the biochemical properties of aggregates formed in vitro by synthetic prion peptides and affected prion infectivity in experimental scrapie. Co-incubation of ST1859 with the peptides PrP 106–126 and PrP 82–146 reduced their fibrillogenic capacity and their resistance to digestion with protease K. Hamsters inoculated with the ST1859-treated homogenate showed a significant delay in the onset of clinical signs of disease and longer survival. Survival was also significantly longer in infected hamsters treated peripherally with ST1859 for the whole post-inoculation period until the onset of clinical symptoms. Similar results were found with the analogue ST1745. Our data indicate that ST1859 reduces prion infectivity and can exert a therapeutic effect in experimental scrapie.

favorable BBB passage


“Anti-prion activity of protein-bound polysaccharide K in prion-infected cells and animals,” Hamanaka et. al., Biochemical and Biophysical Research Communications (2011)

Protein-bound polysaccharide K (PSK) is a clinical immunotherapeutic agent that exhibits various biological activities, including anti-tumor and anti-microbial effects. In the present study, we report on the anti-prion activity of PSK. It inhibited the formation of protease-resistant abnormal prion protein in prion-infected cells without any apparent alterations in either the normal prion protein turnover or the autophagic function in the cells. Its anti-prion activity was predominantly composed of the high molecular weight component(s) of the protein portion of PSK. A single subcutaneous dose of PSK slightly but significantly prolonged the survival time of peritoneally prion-infected mice, but PSK-treated mice produced neutralizing antibodies against the anti-prion activity of PSK. These findings suggest that PSK is a new anti-prion substance that may be useful in elucidating the mechanism of prion replication, although the structure of the anti-prion component(s) of PSK requires further evaluation.

► Our findings provide the first evidence of protein-bound polysaccharide K (PSK) as a new type of anti-prion compound. ► K PSK has anti-prion activity in vitro and in vivo. ► High molecular weight protein component(s) of PSK mainly cause anti-prion activity. ► PSK may be useful in elucidating the mechanism of prion replication.


“Intraventricular pentosan polysulfate in human prion diseases: an observational study in the UK,” Bone et. al., European Journal of Neurology (2008)

This observational study assessed the effect of continuous intraventricular infusion of pentosan polysulphate (PPS) in seven patients at different clinical centres in the UK.  Complications of intraventricular catheterization were frequent. PPS was well-tolerated over a wide dose range (11-110 microg/kg/day) during the 6-month study. Four patients were assessed for the entire study period: one remained stable, two showed minimal deterioration and one progressed significantly. Mean survival of all patients was longer than reported values for natural history of specific prion disorders. Possible reasons for these findings are explored.

Commentary from MRC:

  • PPS does not stop the progression of vCJD and other prion diseases.  Loss of brain function continues after treatment has started and, where measured by a series of brain scans, loss of brain tissue also continued.
  • The drug itself does not seem to carry a risk of serious side effects from prolonged usage at the modest doses given although one patient had seizures and two others evidence of bleeding into the brain.
  • Surgical complications of intraventricular catheter and pump placement occurred.
  • There remains uncertainty as to what precise dose should be administered to individuals.
  • Some of the patients treated with PPS appear to have survived for long periods.  However, it cannot be concluded that the treatment has had a beneficial effect, because it was impossible to make direct comparison with similar but untreated patients.  It is also very difficult to determine exactly when the disease starts and this obviously affects the estimation of survival time.


“Mechanistic insights into the cure of prion disease by novel antiprion compounds,” Webb et al., Journal of Virology (2007)


Congo red in an azo dye that has been used for many years to detect abnormal prion protein in the brains of diseased patients or animals.  Congo red has little therapeutic potential for the treatment of these diseases due to toxicity and poor permeation of the blood-brain barrier.  We have prepared two Congo red derivatives, designed without these liabilities, with potent activity in cellular models of prion disease.  One of these compounds cured cells of the transmissible agent.  The mechanism of action oft these compounds is possibly multifactorial.  The high affinity of Congo red derivatives, including compounds that are ineffective and are effective at the cure of prion disease, for abnormally folded prion protein suggests that the amyloidophylic property of there derivatives is not as critical to the mechanism of as other effects.  Congo red derivatives that are effective at the cure of prion disease increased the degradation of abnormal PrP by the proteasome.  Therefore, the principal mechanism of action of the Congo red analogues was to prevent inhibition of proteasomal activity by Prp(Sc).

In summary, we have shown that two Congo red derivatives (termed WSP677 and WSP740) are able to cause a persistent clearing of PK-resistant protein from scrapie-infected cells.  We also showed that WSP740 could eliminate the agent of disease transmission…. The mechanism of action of the compounds potentially involves the binding of PrP aggregates and increased activity of the proteasome in breaking them downAntioxidant activity of some of the compounds could also play a role but would be secondary to other factors…. We anticipate that the present work will facilitate further studies to determine if WSP740 is also an effective antiprion agent in vivo.


“HEPES inhibits the conversion of prion protein in cell culture,” Delmouly et al., J Gen Virol (2011)


HEPES is a well-known buffering reagent used in cell-culture medium. Interestingly, this compound is also responsible for significant modifications of biological parameters such as uptake of organic molecules, alteration of oxidative stress mechanisms or inhibition of ion channels. While using cell-culture medium supplemented with HEPES on prion-infected cells, it was noticed that there was a significant concentration-dependent inhibition of accumulation of the abnormal isoform of the prion protein (PrP(Sc)). This effect was present only in live cells and was thought to be related to modification of the PrP environment or biology. These results could modify the interpretation of cell-culture assays of prion therapeutic agents, as well as of previous cell biology results obtained in the field using HEPES buffers. This inhibitory effect of HEPES could also be exploited to prevent contamination or propagation of prions in cell culture.