title: A Fluorescent Oligothiophene-Bis-Triazine ligand interacts with PrP fibrils and detects SDS-resistant oligomers in human prion diseases.
abstract: BACKGROUND
  Prion diseases are characterized by the accumulation in the central nervous system of an abnormally folded isoform of the prion protein, named PrP(Sc). Aggregation of PrP(Sc) into oligomers and fibrils is critically involved in the pathogenesis of prion diseases. Oligomers are supposed to be the key neurotoxic agents in prion disease, so modulation of prion aggregation pathways with small molecules can be a valuable strategy for studying prion pathogenicity and for developing new diagnostic and therapeutic approaches. We previously identified thienyl pyrimidine compounds that induce SDS-resistant PrP(Sc) (rSDS-PrP(Sc)) oligomers in prion-infected samples.
  
  
  RESULTS
  Due to the low effective doses of the thienyl pyrimidine hits, we synthesized a quaterthiophene-bis-triazine compound, called MR100 to better evaluate their diagnostic and therapeutic potentials. This molecule exhibits a powerful activity inducing rSDS-PrP(Sc) oligomers at nanomolar concentrations in prion-infected cells. Fluorescence interaction studies of MR100 with mouse PrP fibrils showed substantial modification of the spectrum, and the interaction was confirmed in vitro by production of rSDS-oligomer species upon incubation of MR100 with fibrils in SDS-PAGE gel. We further explored whether MR100 compound has a potential to be used in the diagnosis of prion diseases. Our results showed that: (i) MR100 can detect rSDS-oligomers in prion-infected brain homogenates of various species, including human samples from CJD patients; (ii) A protocol, called "Rapid Centrifugation Assay" (RCA), was developed based on MR100 property of inducing rSDS-PrP(Sc) oligomers only in prion-infected samples, and avoiding the protease digestion step. RCA allows the detection of both PK-sensitive and PK-resistant PrP(Sc) species in rodents samples but also from patients with different CJD forms (sporadic and new variant); (iii) A correlation could be established between the amount of rSDS-PrP(Sc) oligomers revealed by MR100 and the duration of the symptomatic phase of the disease in CJD patients; and (iv) Bioassay experiments showed that MR100 can trap prion infectivity more efficiently than P30 drug.
  
  
  CONCLUSIONS
  MR100 is a powerful tool not only for studying the prion aggregation pathways regarding oligomeric and sPrP(Sc) species, but also for developing alternative methods for the detection of prion-infected samples. Considering our bioassay results, MR100 is a promising molecule for the development of prion decontamination approaches.
authors.affiliation: Université Montpellier, Montpellier, F-34095, France.|Inserm, U1198, Montpellier, F-34095, France.|EPHE, Paris, F-75007, France.
authors.forename: Thibaut
authors.initials: T
authors.lastname: Imberdis
authors.affiliation: Université Montpellier, Montpellier, F-34095, France.|Inserm, U1198, Montpellier, F-34095, France.|EPHE, Paris, F-75007, France.
authors.forename: Adeline
authors.initials: A
authors.lastname: Ayrolles-Torro
authors.affiliation: Institut Charles Gerhardt Montpellier, UMR 5253 CNRS-UM2-ENSCM-UM, Matériaux Avancés pour la Catalyse et la Santé, ENSCM, 8 rue de l'Ecole Normale, 34296, Montpellier cedex 5, France.
authors.forename: Alysson
authors.initials: A
authors.lastname: Duarte Rodrigues
authors.affiliation: Université Montpellier, Montpellier, F-34095, France.|Inserm, U1198, Montpellier, F-34095, France.|EPHE, Paris, F-75007, France.
authors.forename: Joan
authors.initials: J
authors.lastname: Torrent
authors.affiliation: Etablissement Confiné d'Expérimentation ECE, CECEMA, US009 Biocampus, UMS 3426, Université Montpellier, Montpellier, F-34095, France.
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Alvarez-Martinez
authors.affiliation: Institute of Neurology, Medical University Vienna, A-1097, Vienna, Austria.
authors.forename: Gabor G
authors.initials: GG
authors.lastname: Kovacs
authors.affiliation: Université Montpellier, Montpellier, F-34095, France.|Inserm, U1198, Montpellier, F-34095, France.|EPHE, Paris, F-75007, France.
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Verdier
authors.affiliation: Institut Charles Gerhardt Montpellier, UMR 5253 CNRS-UM2-ENSCM-UM, Matériaux Avancés pour la Catalyse et la Santé, ENSCM, 8 rue de l'Ecole Normale, 34296, Montpellier cedex 5, France.
authors.forename: Mike
authors.initials: M
authors.lastname: Robitzer
authors.affiliation: Université Montpellier, Montpellier, F-34095, France. veronique.perrier@univ-montp2.fr.|Inserm, U1198, Montpellier, F-34095, France. veronique.perrier@univ-montp2.fr.|EPHE, Paris, F-75007, France. veronique.perrier@univ-montp2.fr.
authors.forename: Véronique
authors.initials: V
authors.lastname: Perrier
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000379:methods; D001921:Brain / Q000378:metabolism*; D004195:Disease Models, Animal; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D051379:Mice; D018621:PrPC Proteins / Q000378:metabolism*; D017096:Prion Diseases / Q000175:diagnosis* / Q000378:metabolism*; D011328:Prions; D011743:Pyrimidines / Q000378:metabolism*
chemical_list: D005456:Fluorescent Dyes; D018621:PrPC Proteins; D011328:Prions; D011743:Pyrimidines; C030986:pyrimidine
references.citation: Sci Transl Med. 2015 Aug 5;7(299):299ra123
references.pmid: 26246168
references.citation: Chemistry. 2014 Sep 22;20(39):12537-43
references.pmid: 25111601
references.citation: Eur J Biochem. 2000 May;267(10):2833-9
references.pmid: 10806380
references.citation: Lancet. 2001 Jul 21;358(9277):171-80
references.pmid: 11476832
references.citation: Biochemistry. 2002 Oct 22;41(42):12868-75
references.pmid: 12379130
references.citation: Annu Rev Neurosci. 2003;26:267-98
references.pmid: 12704221
references.citation: J Neurochem. 2004 Apr;89(2):454-63
references.pmid: 15056288
references.citation: Science. 1982 Apr 9;216(4542):136-44
references.pmid: 6801762
references.citation: Cell. 1986 Aug 15;46(4):503-11
references.pmid: 3015416
references.citation: Cell. 1989 Dec 1;59(5):847-57
references.pmid: 2574076
references.citation: Biochemistry. 1991 Aug 6;30(31):7672-80
references.pmid: 1678278
references.citation: Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):10962-6
references.pmid: 7902575
references.citation: Nat Med. 1998 Oct;4(10):1157-65
references.pmid: 9771749
references.citation: J Mol Biol. 2005 Feb 18;346(2):645-59
references.pmid: 15670611
references.citation: Proc Natl Acad Sci U S A. 2005 Mar 1;102(9):3501-6
references.pmid: 15741275
references.citation: Nature. 2005 Sep 8;437(7056):257-61
references.pmid: 16148934
references.citation: Biochemistry. 2006 Dec 26;45(51):15710-7
references.pmid: 17176093
references.citation: Expert Rev Anti Infect Ther. 2007 Aug;5(4):613-30
references.pmid: 17678425
references.citation: PLoS Pathog. 2007 Aug 31;3(8):e125
references.pmid: 17784787
references.citation: FEBS J. 2008 May;275(9):2021-31
references.pmid: 18355314
references.citation: J Biol Chem. 2008 Jun 6;283(23):15988-96
references.pmid: 18400757
references.citation: Ann Neurol. 2008 Jun;63(6):697-708
references.pmid: 18571782
references.citation: PLoS One. 2008;3(7):e2773
references.pmid: 18648643
references.citation: Int J Mol Sci. 2009 Mar;10(3):976-99
references.pmid: 19399233
references.citation: Neuropathol Appl Neurobiol. 2009 Dec;35(6):628-32
references.pmid: 19671081
references.citation: J Neurol Neurosurg Psychiatry. 2010 Sep;81(9):1052-5
references.pmid: 20547632
references.citation: Ann Neurol. 2010 Aug;68(2):162-72
references.pmid: 20695009
references.citation: J Mol Biol. 2010 Jul 23;400(4):908-21
references.pmid: 20553730
references.citation: J Neurosci. 2011 Oct 19;31(42):14882-92
references.pmid: 22016521
references.citation: Nat Chem Biol. 2012 Jan;8(1):93-101
references.pmid: 22101602
references.citation: Acta Neuropathol. 2013 Jun;125(6):795-813
references.pmid: 23604588
references.citation: Chemistry. 2013 Jul 29;19(31):10179-92
references.pmid: 23780508
references.citation: J Alzheimers Dis. 2013;37(1):19-28
references.pmid: 23727899
references.citation: Curr Top Med Chem. 2013;13(19):2477-83
references.pmid: 24059332
references.citation: J Virol. 2000 Jan;74(1):320-5
references.pmid: 10590120