A highly systematic approach for the development of both orally bioavailable and bioactive cyclic N-methylated hexapeptides as high affinity ligands for the integrin αvβ3 is based on two concepts: a) screening of systematically designed libraries with spatial diversity and b) masking of the peptide charge with a lipophilic protecting group. The key steps of the method are 1) initial design of a combinatorial library of N-methylated analogues of the stem peptide cyclo(d-Ala-Ala ); 2) selection of cyclic peptides with the highest intestinal permeability; 3) design of sublibraries with the bioactive RGD sequence in all possible positions; 4) selection of the best ligands for RGD-recognizing integrin subtypes; 5) fine-tuning of the affinity and selectivity by additional Ala to Xaa substitutions; 6) protection of the charged functional groups according to the prodrug concept to regain intestinal and oral permeability; 7) proof of biological effects in mice after oral administration.
MyD88 is a cytoplasmic adaptor protein that plays a central role in signaling downstream of the TLRs and the IL1R superfamily. We previously demonstrated that MyD88 plays a critical role in EAE, the murine model of multiple sclerosis, and showed that the MyD88 BB-loop decoy peptide RDVLPGT ameliorates EAE. We now designed and screened a library of backbone cyclized peptides based on the linear BB loop peptide, to identify a metabolically stable inhibitor of MyD88 that retains the binding properties of the linear peptide. We identified a novel cyclic peptide protein mimetic that inhibits inflammatory responses to TLR ligands, and NFκB activation in response to IL-1 activation. The inhibitor, c(MyD 4-4), is metabolically stable in comparison to the linear peptide, blocks MyD88 in a specific manner, and inhibits MyD88 function by preventing MyD88 dimerization. Finally, treatment of mice with c(MyD 4-4) reduced the severity of clinical disease in the murine EAE model of multiple sclerosis. Thus, modulation of MyD88-dependent signaling using c(MyD 4-4) is a potential therapeutic strategy to lower innate immune inflammation in autoimmune CNS disease.
Ein systematischer Ansatz fürdie Entwicklung oral verfügbarer wie auchb iologischa ktiver,c yclischer,N -methylierter Hexapeptide als hoch affine Liganden fürd as Integrin avb3b eruht auf zwei Konzepten:a )Screening systematisch entwickelter Peptidbibliotheken mit räumlicher Diversitätund b) Maskierung der Peptidladung mithilfe lipophiler Schutzgruppen. Die Schlüsselschritte dieser Methode sind 1) Erstellung einer kombinatorischen Bibliothek N-methylierter Analoga des Stammpeptids cyclo(d-Ala-Ala 5 ); 2) Auswahl der cyclischen Peptide mit der hçchsten intestinalen Permeabilität; 3) Design von Unterbibliotheken mit der biologischa ktiven RGD-Sequenz in allen mçglichen Positionen;4)Auswahl der besten Liganden fürR GD-erkennende Integrinsubtypen; 5) Optimierung der Affinitätund Selektivitätdurch zusätzliche Ala-zu-Xaa-Substitutionen;6 )Schützen der geladenen funktionalen Seitengruppen nach dem Prodrug-Konzept zur Wiederherstellung der intestinalen und oralen Permeabilität; 7) Nachweis der biologischen AktivitätinMäusen nachoraler Gabe.
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