Evaluation of Quinacrine Treatment for Prion Diseases

ABSTRACT Based on in vitro observations in scrapie-infected neuroblastoma cells, quinacrine has recently been proposed as a treatment for Creutzfeldt-Jakob disease (CJD), including a new variant CJD which is linked to contamination of food by the bovine spongiform encephalopathy (BSE) agent. The present study investigated possible mechanisms of action of quinacrine on prions. The ability of quinacrine to interact with and to reduce the protease resistance of PrP peptide aggregates and PrPres of human and animal origin were analyzed, together with its ability to inhibit the in vitro conversion of the normal prion protein (PrPc) to the abnormal form (PrPres). Furthermore, the efficiencies of quinacrine and chlorpromazine, another tricyclic compound, were examined in different in vitro models and in an experimental murine model of BSE. Quinacrine efficiently hampered de novo generation of fibrillogenic prion protein and PrPres accumulation in ScN2a cells. However, it was unable to affect the protease resistance of preexisting PrP fibrils and PrPres from brain homogenates, and a “curing” effect was obtained in ScGT1 cells only after lengthy treatment. In vivo, no detectable effect was observed in the animal model used, consistent with other recent studies and preliminary observations in humans. Despite its ability to cross the blood-brain barrier, the use of quinacrine for the treatment of CJD is questionable, at least as a monotherapy. The multistep experimental approach employed here could be used to test new therapeutic regimes before their use in human trials.

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Inhibition of Protease-Resistant Prion Protein Accumulation In Vitro by Curcumin

Journal of Virology ◽

10.1128/jvi.77.9.5499-5502.2003 ◽

2003 ◽

Vol 77 (9) ◽

pp. 5499-5502 ◽

Cited By ~ 74

Author(s):

Byron Caughey ◽

Lynne D. Raymond ◽

Gregory J. Raymond ◽

Laura Maxson ◽

Jay Silveira ◽

...

Keyword(s):

Prion Protein ◽

Inhibitory Concentration ◽

Transmissible Spongiform Encephalopathy ◽

Neuroblastoma Cells ◽

In Vivo Studies ◽

Spongiform Encephalopathy ◽

Protein Accumulation ◽

The Brain

ABSTRACT Inhibition of the accumulation of protease-resistant prion protein (PrP-res) is a prime strategy in the development of potential transmissible spongiform encephalopathy (TSE) therapeutics. Here we show that curcumin (diferoylmethane), a major component of the spice turmeric, potently inhibits PrP-res accumulation in scrapie agent-infected neuroblastoma cells (50% inhibitory concentration, ∼10 nM) and partially inhibits the cell-free conversion of PrP to PrP-res. In vivo studies showed that dietary administration of curcumin had no significant effect on the onset of scrapie in hamsters. Nonetheless, other studies have shown that curcumin is nontoxic and can penetrate the brain, properties that give curcumin advantages over inhibitors previously identified as potential prophylactic and/or therapeutic anti-TSE compounds.

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PrP P102L and Nearby Lysine Mutations Promote Spontaneous In Vitro Formation of Transmissible Prions

Journal of Virology ◽

10.1128/jvi.01276-17 ◽

2017 ◽

Vol 91 (21) ◽

Cited By ~ 6

Author(s):

Allison Kraus ◽

Gregory J. Raymond ◽

Brent Race ◽

Katrina J. Campbell ◽

Andrew G. Hughson ◽

...

Keyword(s):

Prion Protein ◽

Prion Diseases ◽

Transmissible Spongiform Encephalopathy ◽

Clinical Signs ◽

Protein Aggregates ◽

Structural Features ◽

Spongiform Encephalopathy ◽

Specific Sequence

ABSTRACT Accumulation of fibrillar protein aggregates is a hallmark of many diseases. While numerous proteins form fibrils by prion-like seeded polymerization in vitro, only some are transmissible and pathogenic in vivo. To probe the structural features that confer transmissibility to prion protein (PrP) fibrils, we have analyzed synthetic PrP amyloids with or without the human prion disease-associated P102L mutation. The formation of infectious prions from PrP molecules in vitro has required cofactors and/or unphysiological denaturing conditions. Here, we demonstrate that, under physiologically compatible conditions without cofactors, the P102L mutation in recombinant hamster PrP promoted prion formation when seeded by minute amounts of scrapie prions in vitro. Surprisingly, combination of the P102L mutation with charge-neutralizing substitutions of four nearby lysines promoted spontaneous prion formation. When inoculated into hamsters, both of these types of synthetic prions initiated substantial accumulation of prion seeding activity and protease-resistant PrP without transmissible spongiform encephalopathy (TSE) clinical signs or notable glial activation. Our evidence suggests that PrP's centrally located proline and lysine residues act as conformational switches in the in vitro formation of transmissible PrP amyloids. IMPORTANCE Many diseases involve the damaging accumulation of specific misfolded proteins in thread-like aggregates. These threads (fibrils) are capable of growing on the ends by seeding the refolding and incorporation of the normal form of the given protein. In many cases such aggregates can be infectious and propagate like prions when transmitted from one individual host to another. Some transmitted aggregates can cause fatal disease, as with human iatrogenic prion diseases, while other aggregates appear to be relatively innocuous. The factors that distinguish infectious and pathogenic protein aggregates from more innocuous ones are poorly understood. Here we have compared the combined effects of prion seeding and mutations of prion protein (PrP) on the structure and transmission properties of synthetic PrP aggregates. Our results highlight the influence of specific sequence features in the normally unstructured region of PrP that influence the infectious and neuropathogenic properties of PrP-derived aggregates.

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Synergistic and strain-specific effects of bovine spongiform encephalopathy and scrapie prions in the cell-free conversion of recombinant prion protein

Journal of General Virology ◽

10.1099/vir.0.81590-0 ◽

2006 ◽

Vol 87 (12) ◽

pp. 3753-3761 ◽

Cited By ~ 13

Author(s):

Martin Eiden ◽

Gottfried J. Palm ◽

Winfried Hinrichs ◽

Ulrich Matthey ◽

Ralph Zahn ◽

...

Keyword(s):

Bovine Spongiform Encephalopathy ◽

Prion Protein ◽

De Novo ◽

Proteinase K ◽

Spongiform Encephalopathy ◽

Cooperative Action ◽

Specific Effects ◽

Scrapie Strains ◽

Recombinant Prion Protein

This study describes the conversion of murine PrPC by PrPSc from three different mouse scrapie strains (ME7, 87V and 22A) and from a mouse-passaged bovine spongiform encephalopathy (BSE) strain (BSE/Bl6). This was demonstrated by a modified, non-radioactive, cell-free conversion assay using bacterial prion protein, which was converted into a proteinase K (PK)-resistant fragment designated PrPres. Using this assay, newly formed PrPres could be detected by an antibody that discriminated de novo PrPres and the original PrPSc seed. The results suggested that PrPres formation occurs in three phases: the first 48 h when PrPres formation is delayed, followed by a period of substantially accelerated PrPres formation and a plateau phase when a maximum concentration of PrPres is reached after 72 h. The conversion of prokaryotically expressed PrPC by ME7 and BSE prions led to unglycosylated, PK-digested, abnormal PrPres fragments, which differed in molecular mass by 1 kDa. Therefore, prion strain phenotypes were retained in the cell-free conversion, even when recombinant PrPC was used as the substrate. Moreover, co-incubation of ME7 and BSE prions resulted in equal amounts of both ME7- and BSE-derived PrPres fragments (as distinguished by their different molecular sizes) and also in a significantly increased total amount of de novo-generated PrPres. This was found to be more than twice the amount of either strain when incubated separately. This result indicates a synergistic effect of both strains during cell-free conversion. It is not yet known whether such a cooperative action between BSE and scrapie prions also occurs in vivo.

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ALT neuroblastoma chemoresistance due to telomere dysfunction–induced ATM activation is reversible with ATM inhibitor AZD0156

Science Translational Medicine ◽

10.1126/scitranslmed.abd5750 ◽

2021 ◽

Vol 13 (607) ◽

pp. eabd5750

Author(s):

Balakrishna Koneru ◽

Ahsan Farooqi ◽

Thinh H. Nguyen ◽

Wan Hsi Chen ◽

Ashly Hindle ◽

...

Keyword(s):

Cell Lines ◽

Ataxia Telangiectasia ◽

De Novo ◽

Clinical Stage ◽

Neuroblastoma Cell ◽

Neuroblastoma Cells ◽

Telomere Dysfunction ◽

Neuroblastoma Cell Lines

Cancers overcome replicative immortality by activating either telomerase or an alternative lengthening of telomeres (ALT) mechanism. ALT occurs in ~25% of high-risk neuroblastomas, and progression in patients with ALT neuroblastoma during or after front-line therapy is frequent and often fatal. Temozolomide + irinotecan is commonly used as salvage therapy for neuroblastoma. Patient-derived cell lines and xenografts established from patients with relapsed ALT neuroblastoma demonstrated de novo resistance to temozolomide + irinotecan [SN-38 in vitro, P < 0.05; in vivo mouse event-free survival (EFS), P < 0.0001] vs. telomerase-positive neuroblastomas. We observed that ALT neuroblastoma cells manifested constitutive ataxia-telangiectasia mutated (ATM) activation due to spontaneous telomere dysfunction which was not observed in telomerase-positive neuroblastoma cells. We demonstrated that induction of telomere dysfunction resulted in ATM activation that, in turn, conferred resistance to temozolomide + SN-38 (4.2-fold change in IC50, P < 0.001). ATM knockdown (shRNA) or inhibition using a clinical-stage small-molecule inhibitor (AZD0156) reversed resistance to temozolomide + irinotecan in ALT neuroblastoma cell lines in vitro (P < 0.001) and in four ALT xenografts in vivo (EFS, P < 0.0001). AZD0156 showed modest to no enhancement of temozolomide + irinotecan activity in telomerase-positive neuroblastoma cell lines and xenografts. Ataxia telangiectasia and Rad3 related (ATR) inhibition using AZD6738 did not enhance temozolomide + SN-38 activity in ALT neuroblastoma cells. Thus, ALT neuroblastoma chemotherapy resistance occurs via ATM activation and is reversible with ATM inhibitor AZD0156. Combining AZD0156 with temozolomide + irinotecan warrants clinical testing for neuroblastoma.

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Prion protein as a target for therapeutic interventions

Pure and Applied Chemistry ◽

10.1351/pac200476050915 ◽

2004 ◽

Vol 76 (5) ◽

pp. 915-920 ◽

Cited By ~ 4

Author(s):

P. P. Liberski

Keyword(s):

Prion Protein ◽

Phenotypic Expression ◽

Therapeutic Interventions ◽

Transmissible Spongiform Encephalopathies ◽

Spongiform Encephalopathy ◽

Public Health Perspective ◽

Spongiform Encephalopathies ◽

Mad Cow Disease

Transmissible spongiform encephalopathies (TSEs), currently known as prion diseases, are neurodegenerative disorders of the central nervous system (CNS) caused by an elusive infectious agent called “prion” (proteinaceous infectious particle). These dis orders include: kuru, Creutzfeldt –Jakob disease (CJD) and its variant (vCJD), Gerstmann–Sträussler–Scheinker (GSS) disease and fatal familial insomnia (FFI) in humans, scrapie in sheep and goats, bovine spongiform encephalopathy (BSE) or mad cow disease, and chronic wasting disease (CWD) in cervids. According to the widely accepted “prion hypothesis”, prion is an aggregate of the abnormal isoform of prion protein (PrPSc). Prion protein is a cell-derived glycoprotein (this normal isoform is called PrPc) encoded by a gene on chromosome 20 in humans (PRNP). In familial forms of TSEs, mutations within the ORF of PRNP are linked to the phenotypic expression of the disease. TSEs are important from public health perspective, and “mad cow disease has created the greatest threat to the safety of human food supply in modern times. vCJD threatens the safety of the blood supply worldwide”. Thus, to search for effective therapy is more than an urgent task. In TSEs, aggregates of PrPSc accumulate in the brain in a form of plaques, or synaptic deposits. The conversion of PrPc into PrPSc and subsequent deposits of PrPSc are targets for therapeutic interventions. These include: tricyclic compounds—acridine and phenothiazine derivatives; quinacrine; anti-PrPSc antibodies; dendrimers; polyethylene antibiotics (amphotericin B, MS-8209); pentosan polysulfate; and dextran sulfate. All these compounds are active in many in vitro and in vivo assays, but not proved definitely active in humans. Thus, albeit interesting and promising, the chemotherapy of TSEs is still in the infant phase.

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Characterization and polyanion-binding properties of purified recombinant prion protein

Biochemical Journal ◽

10.1042/bj3420605 ◽

1999 ◽

Vol 342 (3) ◽

pp. 605-613 ◽

Cited By ~ 20

Author(s):

Debbie B. BRIMACOMBE ◽

Alan D. BENNETT ◽

Fred S. WUSTEMAN ◽

Andrew C. GILL ◽

Janine C. DANN ◽

...

Keyword(s):

Prion Protein ◽

Mammalian Cells ◽

Metal Ion ◽

Transmissible Spongiform Encephalopathy ◽

Spongiform Encephalopathy ◽

Divalent Metal Ions ◽

Dissociation Kinetics ◽

Recombinant Prion Protein

Certain polysulphated polyanions have been shown to have prophylactic effects on the progression of transmissible spongiform encephalopathy disease, presumably because they bind to prion protein (PrP). Until now, the difficulty of obtaining large quantities of native PrP has precluded detailed studies of these interactions. We have over-expressed murine recombinant PrP (recPrP), lacking its glycophosphoinositol membrane anchor, in modified mammalian cells. Milligram quantities of secreted, soluble and partially glycosylated protein were purified under non-denaturing conditions and the identities of mature-length aglycosyl recPrP and two cleavage fragments were determined by electrospray MS. Binding was assessed by surface plasmon resonance techniques using both direct and competitive ligand-binding approaches. recPrP binding to immobilized polyanions was enhanced by divalent metal ions. Polyanion binding was strong and showed complex association and dissociation kinetics that were consistent with ligand-directed recPrP aggregation. The differences in the binding strengths of recPrP to pentosan polysulphate and to other sulphated polyanions were found to parallel their in vivo anti-scrapie and in vitro anti-scrapie-specific PrP formation potencies. When recPrP was immobilized by capture on metal-ion chelates it was found, contrary to expectation, that the addition of polyanions promoted the dissociation of the protein.

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Mutated but Not Deleted Ovine PrPCN-Terminal Polybasic Region Strongly Interferes with Prion Propagation in Transgenic Mice

Journal of Virology ◽

10.1128/jvi.02805-15 ◽

2015 ◽

Vol 90 (3) ◽

pp. 1638-1646 ◽

Cited By ~ 4

Author(s):

Manal Khalifé ◽

Fabienne Reine ◽

Sophie Paquet-Fifield ◽

Johan Castille ◽

Laetitia Herzog ◽

...

Keyword(s):

Amino Acid ◽

Prion Protein ◽

Major Effect ◽

Cellular Prion Protein ◽

Spongiform Encephalopathy ◽

Structural Variations ◽

Prion Propagation ◽

Polybasic Region

ABSTRACTMammalian prions are proteinaceous infectious agents composed of misfolded assemblies of the host-encoded, cellular prion protein (PrP). Physiologically, the N-terminal polybasic region of residues 23 to 31 of PrP has been shown to be involved in its endocytic trafficking and interactions with glycosaminoglycans or putative ectodomains of membrane-associated proteins. Several recent reports also describe this PrP region as important for the toxicity of mutant prion proteins and the efficiency of prion propagation, bothin vitroandin vivo. The question remains as to whether the latter observations made with mouse PrP and mouse prions would be relevant to other PrP species/prion strain combinations given the dramatic impact on prion susceptibility of minimal amino acid substitutions and structural variations in PrP. Here, we report that transgenic mouse lines expressing ovine PrP with a deletion of residues 23 to 26 (KKRP) or mutated in this N-terminal region (KQHPH instead of KKRPK) exhibited a variable, strain-dependent susceptibility to prion infection with regard to the proportion of affected mice and disease tempo relative to findings in their wild-type counterparts. Deletion has no major effect on 127S scrapie prion pathogenesis, whereas mutation increased by almost 3-fold the survival time of the mice. Deletion marginally affected the incubation time of scrapie LA19K and ovine bovine spongiform encephalopathy (BSE) prions, whereas mutation caused apparent resistance to disease.IMPORTANCERecent reports suggested that the N-terminal polybasic region of the prion protein could be a therapeutic target to prevent prion propagation or toxic signaling associated with more common neurodegenerative diseases such as Alzheimer's disease. Mutating or deleting this region in ovine PrP completes the data previously obtained with the mouse protein by identifying the key amino acid residues involved.

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Down-Regulation of Phosphoribosyl Pyrophosphate Synthetase 1 Inhibits Neuroblastoma Cell Proliferation

Cells ◽

10.3390/cells8090955 ◽

2019 ◽

Vol 8 (9) ◽

pp. 955

Author(s):

Jifu Li ◽

Junhong Ye ◽

Shunqin Zhu ◽

Hongjuan Cui

Keyword(s):

Cell Proliferation ◽

De Novo ◽

Neuroblastoma Cell ◽

Neuroblastoma Cells ◽

Phosphoribosyl Pyrophosphate ◽

Down Regulation ◽

Nucleotide Synthesis ◽

Phosphoribosyl Pyrophosphate Synthetase

Phosphoribosyl pyrophosphate synthetase 1 (PRPS1) is a key enzyme in de novo nucleotide synthesis and nucleotide salvage synthesis pathways that are critical for purine and pyrimidine biosynthesis. Abnormally high expression of PRPS1 can cause many diseases, including hearing loss, hypotonia, and ataxia, in addition to being associated with neuroblastoma. However, the role of PRPS1 in neuroblastoma is still unclear. In this study, we found that PRPS1 was commonly expressed in neuroblastoma cells and was closely related to poor prognosis for cancer. Furthermore, down-regulation of PRPS1 inhibited neuroblastoma cell proliferation and tumor growth in vitro and in vivo via disturbing DNA synthesis. This study provides new insights into the treatment of neuroblastoma patients and new targets for drug development.

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Inactivation of Animal and Human Prions by Hydrogen Peroxide Gas Plasma Sterilization

Infection Control and Hospital Epidemiology ◽

10.1086/598342 ◽

2009 ◽

Vol 30 (8) ◽

pp. 769-777 ◽

Cited By ~ 41

Author(s):

C. Rogez-Kreuz ◽

R. Yousfi ◽

C. Soufflet ◽

I. Quadrio ◽

Z.-X. Yan ◽

...

Keyword(s):

Hydrogen Peroxide ◽

Prion Protein ◽

Medical Devices ◽

Transmissible Spongiform Encephalopathies ◽

Spongiform Encephalopathy ◽

Plasma Sterilization ◽

Gas Plasma ◽

Resistant Form

Prions cause various transmissible spongiform encephalopathies. They are highly resistant to the chemical and physical decontamination and sterilization procedures routinely used in healthcare facilities. The decontamination procedures recommended for the inactivation of prions are often incompatible with the materials used in medical devices. In this study, we evaluated the use of low-temperature hydrogen peroxide gas plasma sterilization systems and other instrument-processing procedures for inactivating human and animal prions. We provide new data concerning the efficacy of hydrogen peroxide against prions from in vitro or in vivo tests, focusing on the following: the efficiency of hydrogen peroxide sterilization and possible interactions with enzymatic or alkaline detergents, differences in the efficiency of this treatment against different prion strains, and the influence of contaminating lipids. We found that gaseous hydrogen peroxide decreased the infectivity of prions and/or the level of the protease-resistant form of the prion protein on different surface materials. However, the efficiency of this treatment depended strongly on the concentration of hydrogen peroxide and the delivery system used in medical devices, because these effects were more pronounced for the new generation of Sterrad technology. The Sterrad NX sterilizer is 100% efficient (0% transmission and no protease-resistant form of the prion protein signal detected on the surface of the material for the mouse-adapted bovine spongiform encephalopathy 6PB1 strain and a variant Creutzfeldt-Jakob disease strain). Thus, gaseous or vaporized hydrogen peroxide efficiently inactivates prions on the surfaces of medical devices.

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In-vitro- und In-vivo-Wirkung von Schilddrüsenhormon auf das Wachstum von Neuroblastomzellen

Nuklearmedizin ◽

10.1055/s-0038-1629520 ◽

1990 ◽

Vol 29 (03) ◽

pp. 120-124

Author(s):

R. P. Baum ◽

E. Rohrbach ◽

G. Hör ◽

B. Kornhuber ◽

E. Busse

Keyword(s):

Cell Differentiation ◽

Neuroblastoma Cells ◽

Control Group ◽

Initial Value ◽

Initial Rise ◽

Biphasic Effect ◽

Contrast Microscopy ◽

Morphological Sign

The effect of triiodothyronine (T3) on the differentiation of cultured neuroblastoma (NB) cells was studied after 9 days of treatment with a dose of 10-4 M/106 cells per day. Using phase contrast microscopy, 30-50% of NB cells showed formation of neurites as a morphological sign of cellular differentiation. The initial rise of the mitosis rate was followed by a plateau. Changes in cyclic nucleotide content, in the triphosphates and in the activity of the enzyme ornithine decarboxylase (ODC) were assessed in 2 human and 2 murine cell lines to serve as biochemical parameters of the cell differentiation induced by T3. Whereas the cAMP level increased significantly (3 to 7 fold compared with its initial value), the cGMP value dropped to 30 to 50% of that of the control group. ATP and GTP increased about 200%, the ODC showed a decrease of about 50%. The present studies show a biphasic effect of T3 on neuroblastoma cells: the initial rise of mitotic activity is followed by increased cell differentiation starting from day 4 of the treatment.

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