Modifying ADAMTS13 to modulate binding of pathogenic autoantibodies of patients with acquired thrombotic thrombocytopenic purpura
| Autor: | Jan Voorberg, Andres Männik, Paul H. P. Kaijen, Gerry A. F. Nicolaes, Karen Vanhoorelbeke, Paul Coppo, Nuno A. G. Graça, Agnès Veyradier, Bogac Ercig, Kadri Kangro, Leydi Carolina Velásquez Pereira |
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| Přispěvatelé: | Graduate School, ACS - Microcirculation, Experimental Vascular Medicine, Landsteiner Laboratory, AII - Inflammatory diseases, Biochemie, RS: Carim - B01 Blood proteins & engineering |
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
PROTEASE
VON-WILLEBRAND-FACTOR Mutant 030204 cardiovascular system & hematology VARIANTS Article Epitope ACTIVATION TSP1 03 medical and health sciences 0302 clinical medicine Asparagine Alanine Acquired Thrombotic Thrombocytopenic Purpura biology Chemistry Autoantibody SPACER DOMAIN ANTI-ADAMTS13 AUTOANTIBODIES Hematology CONTAINS Molecular biology ADAMTS13 ANTIBODIES biology.protein RESIDUES Antibody 030215 immunology |
| Zdroj: | Haematologica, 105(11), 2619-2630. Ferrata Storti Foundation Haematologica Haematologica-the Hematology Journal, 105(11), 2619-2630. Ferrata Storti Foundation |
| ISSN: | 0390-6078 |
| Popis: | Antibodies that develop in patients with immune thrombotic thrombocytopenic purpura commonly target the spacer epitope R568/F592/R660/Y661/Y665 (RFRYY) of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13). In this study we present a detailed contribution of each residue in this epitope for autoantibody binding. Different panels of mutations were introduced to create a large collection of full-length ADAMTS13 variants comprising conservative (Y F), semi-conservative (Y/F -> L), non-conservative (Y/F -> N) or alanine (Y/F/R -> A) substitutions. Previously reported gain-of-function (KYKFF) and truncated `MDTCS' variants were also included. Sera from 18 patients were screened against all variants. Conservative mutations of the aromatic residues did not reduce the binding of autoantibodies. Moderate resistance was achieved by replacing R568 and R660 by lysines or alanines. Semi-conservative mutations of aromatic residues showed a moderate effectiveness in autoantibody resistance. Non-conservative asparagine or alanine mutations of aromatic residues were the most effective. In the mixtures of autoantibodies from the majority (89%) of patients screened, autoantibodies targeting the spacer RFRYY epitope were preponderant compared to other epitopes. Reductions in ADAMTS13 proteolytic activity were observed for all full-length mutant variants, in varying degrees. The greatest reductions in activity were observed in the most autoanti-bodyresistant variants (15-35% residual activity in a FRETS-VWF73 assay). Among these, a triple-alanine mutant - RARAA - showed activity in a von Willebrand factor multimer assay. This study shows that non-conservative and alanine modifications of residues within the exosite-3 spacer RFRYY epitope in full-length ADAMTS13 resist the binding of autoantibodies from patients with immune thrombotic thrombocytopenic purpura, while retaining residual proteolytic activity. Our study provides a framework for the design of autoantibody-resistant ADAMTS13 variants for further therapeutic development. |
| Databáze: | OpenAIRE |
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