A cationic tetrapyrrole inhibits toxic activities of the cellular prion protein

Massignan, Tania; Cimini, Sara; Stincardini, Claudia; Cerovic, Milica; Vanni, Ilaria; Elezgarai, Saioa R.; Moreno, José A.; Stravalaci, Matteo; Negro, Alessandro; Sangiovanni, Valeria; Restelli, Elena; Riccardi, Geraldina; Gobbi, Marco; Castilla, Joaquı́n; Borsello, Tiziana; Nonno, Romolo; Biasini, Emiliano

doi:10.1038/srep23180

Cited by 35 publications

(43 citation statements)

References 63 publications

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“…1, C and D). The effect of TMPyP in the DBCA was recently reported by Massignan and colleagues (35). Given this correlation between the inhibitory activity of compounds in ScN2a cells and in the DBCA, we decided to use the DBCA as the read-out in a high-throughput screen to see if we could discover previously unknown molecules that might also have anti-prion activity.…”

Section: Resultsmentioning

confidence: 99%

Identification of Anti-prion Compounds using a Novel Cellular Assay

Imberdis

Heeres

Yueh

et al. 2016

Journal of Biological Chemistry

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Edited by F. Anne StephensonPrion diseases are devastating neurodegenerative disorders with no known cure. One strategy for developing therapies for these diseases is to identify compounds that block conversion of the cellular form of the prion protein (PrP C ) into the infectious isoform (PrP Sc ). Most previous efforts to discover such molecules by high-throughput screening methods have utilized, as a read-out, a single kind of cellular assay system: neuroblastoma cells that are persistently infected with scrapie prions. Here, we describe the use of an alternative cellular assay based on suppressing the spontaneous cytotoxicity of a mutant form of PrP (⌬105-125). Using this assay, we screened 75,000 compounds, and identified a group of phenethyl piperidines (exemplified by LD7), which reduces the accumulation of PrP Sc in infected neuroblastoma cells by >90% at low micromolar doses, and inhibits PrP Sc -induced synaptotoxicity in hippocampal neurons. By analyzing the structure-activity relationships of 35 chemical derivatives, we defined the pharmacophore of LD7, and identified a more potent derivative. Active compounds do not alter total or cell-surface levels of PrP C , and do not bind to recombinant PrP in surface plasmon resonance experiments, although at high concentrations they inhibit PrP Sc -seeded conversion of recombinant PrP to a misfolded state in an in vitro reaction (RTQuIC). This class of small molecules may provide valuable therapeutic leads, as well as chemical biological tools to identify cellular pathways underlying PrP Sc metabolism and PrP C function.Prion diseases are fatal neurodegenerative disorders that are due to the conversion of a normal, neuronal glycoprotein (PrP C ) 3 into an infectious isoform (PrP Sc ) that propagates itself by an autocatalytic templating process (1, 2). In addition to their intrinsic interest to biologists, prion diseases are of enormous medical and public health concern. A global epidemic of bovine spongiform encephalopathy, a prion disease of cattle, emerged in the 1980s and 1990s, resulting in contamination of food supplies and transmission of the disease to a small number of human beings (3, 4). Prion contamination has also increased the risk of blood transfusions, and organ transplants (5). Most recently, a prion-like process has been found to play a role in the CNS dissemination of misfolded proteins in more common neurodegenerative disorders, including Alzheimer's disease, Parkinson's disease, and tauopathies, and there is even evidence that these diseases can be spread between individuals by iatrogenic means (6, 7). There are currently no cures for prion diseases. A great deal of effort has been invested over the past 25 years in identifying compounds that block the conversion of PrP C into PrP Sc as a therapeutic strategy. Most of these efforts have used as a readout a single kind of cellular assay system: N2a neuroblastoma cells that are chronically infected with scrapie prions (designated ScN2a cells). N2a cells are one of the few cell lines capable...

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Section: Resultsmentioning

confidence: 99%

Identification of Anti-prion Compounds using a Novel Cellular Assay

Imberdis

Heeres

Yueh

et al. 2016

Journal of Biological Chemistry

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“…As expected, cells exposed to zeocin alone showed a strong decrease in viability (<70 %, not shown). Conversely, co‐treatment with the porphyrin Fe III ‐TMPyP, previously shown to inhibit the activity of ΔCR PrP in the DBCA, abrogated the effect (not shown). Among the different compounds, we identified eight molecules ( 34 , 38 , 39, 43 , 46, 50 , 53 and 54 ) capable of rescuing at least 20 % of antibiotic‐induced cell death (Figure ).…”

Section: Figurementioning

confidence: 80%

“…We have previously described the DBCA, a cellular assay based on the intrinsic cytotoxic effects of PrP molecules carrying mutations in the central region of the protein . Multiple previous studies indicated that this assay is a useful tool to study PrP‐mediated neurotoxic pathways, as well as to identify and characterize novel anti‐prion compounds . Here, we used the DBCA to screen a representative diverse set of small molecules from an original collection of chemicals developed at EDASA Scientific.…”

Section: Figurementioning

confidence: 99%

“…To rule out the possibility that the rescuing effects shown by compound 55 in the different DBCA experiments were due to an interaction of the molecule with zeocin, we performed the assay in two modified versions. In contrast to mutant PrP, expression of WT PrP does not confer hypersensitivity to zeocin . Therefore, we performed the DBCA in HEK293 cells stably expressing WT PrP, with increasing concentrations of zeocin (0–2000 μg mL −1 ), in presence or absence of molecule 55 (Figure S7).…”

Section: Figurementioning

confidence: 99%

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A Small‐Molecule Inhibitor of Prion Replication and Mutant Prion Protein Toxicity

et al. 2017

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Into the fold: Prion diseases are neurodegenerative disorders characterized by the accumulation in the brain of a self‐replicating, misfolded isoform (PrPSc) of the cellular prion protein (PrPC). No therapies are available for these pathologies. We capitalized on previously described cell‐based assays to screen a library of small molecules, and identified 55, a compound capable of counteracting both prion replication and toxicity. Compound 55 may represent the starting point for the development of a completely new class of therapeutics for prion diseases.

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“…Here we have examined the effects on α‐synuclein of a tetrapyrrole that was initially discovered in a screen for inhibitors of prion disease, iron(III) meso‐ tetra ( N ‐methyl‐4‐pyridyl‐prophine) (Figure ), denoted hereafter as Fe‐TMPyP. Fe‐TMPyP, which was first proposed to act by stabilizing the natively structured C‐terminal domain of PrP, via specific binding with a stoichiometry of 1, is effective at inhibiting a broad range of prion strains, unlike other anti‐prion compounds . However, it was later found to have high affinity also for the C‐terminal domain when unfolded, with its binding preventing the formation of stable misfolded aggregates .…”

Section: Introductionmentioning

confidence: 99%

Characterizing the inhibition of α‐synuclein oligomerization by a pharmacological chaperone that prevents prion formation by the protein PrP

Dong

Garen

Mercier³

et al. 2019

Protein Science

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Aggregation of the disordered protein α‐synuclein into amyloid fibrils is a central feature of synucleinopathies, neurodegenerative disorders that include Parkinson's disease. Small, pre‐fibrillar oligomers of misfolded α‐synuclein are thought to be the key toxic entities, and α‐synuclein misfolding can propagate in a prion‐like way. We explored whether a compound with anti‐prion activity that can bind to unfolded parts of the protein PrP, the cyclic tetrapyrrole Fe‐TMPyP, was also active against α‐synuclein aggregation. Observing the initial stages of aggregation via fluorescence cross‐correlation spectroscopy, we found that Fe‐TMPyP inhibited small oligomer formation in a dose‐dependent manner. Fe‐TMPyP also inhibited the formation of mature amyloid fibrils in vitro, as detected by thioflavin T fluorescence. Isothermal titration calorimetry indicated Fe‐TMPyP bound to monomeric α‐synuclein with a stoichiometry of 2, and two‐dimensional heteronuclear single quantum coherence NMR spectra revealed significant interactions between Fe‐TMPyP and the C‐terminus of the protein. These results suggest commonalities among aggregation mechanisms for α‐synuclein and the prion protein may exist that can be exploited as therapeutic targets.

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A cationic tetrapyrrole inhibits toxic activities of the cellular prion protein

Cited by 35 publications

References 63 publications

Identification of Anti-prion Compounds using a Novel Cellular Assay

Identification of Anti-prion Compounds using a Novel Cellular Assay

A Small‐Molecule Inhibitor of Prion Replication and Mutant Prion Protein Toxicity

Characterizing the inhibition of α‐synuclein oligomerization by a pharmacological chaperone that prevents prion formation by the protein PrP

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