N-glycan–mediated shielding of ADAMTS13 prevents binding of pathogenic autoantibodies in immune-mediated TTP

Abstract

Immune-mediated thrombotic thrombocytopenic purpura (iTTP) is an autoimmune disorder caused by the development of autoantibodies targeting different domains of ADAMTS13. Profiling studies have shown that residues R568, F592, R660, Y661 and Y665 within exosite-3 of the spacer domain provide an immunodominant region of ADAMTS13 for pathogenic autoantibodies that develop in patients with iTTP. Modification of these 5 core residues with the goal of reducing auto-antibody binding revealed a significant trade-off between autoantibody resistance and proteolytic activity. Here, we employed structural bioinformatics to identify a larger epitope landscape on the ADAMTS13 spacer domain. Models of spacer-antibody complexes predicted that residues R568, L591, F592, K608, M609, R636, L637, R639, R660, Y661, Y665 and L668 contribute to an expanded epitope within the spacer domain. Based on bioinformatics-guided predictions we designed a panel of N-glycan insertions in this expanded epitope to reduce the binding of spacer domain autoantibodies. One N-glycan variant (NGLY3-ADAMTS13, containing a K608N substitution) showed strongly reduced reactivity with TTP patient sera (28%) as compared to WT-ADAMTS13 (100%). Insertion of an N-glycan at amino acid position 608 did not interfere with processing of VWF positioning the resulting NGLY3-ADAMTS13 variant as a potential novel therapeutic option for treatment of iTTP.

The standard of care for immune thrombotic thrombocytopenic purpura today

“…Several anecdotal case reports have demonstrated the safety and efficacy of using such a new strategy for managing acute iTTP. [82][83][84][85] Additionally, recombinant ADAMS-13 86,87 and its potentially antibody-resistant ADAMTS-13 variants 88,89 should be developed for clinical use. This will change how we treat iTTP tomorrow.…”

Syntaxin 5 determines Weibel-Palade body size and Von Willebrand factor secretion by controlling Golgi architecture

“…VWF string assays were essentially performed as described using 75,000 shCTRL or shSTX5 transduced cells per channel in gelatin-coated 6-channel IBIDI μ-slides VI 0.4. 28 Strings were visualized by supplementing perfusion mix with AF488-conjugated anti-VWF antibody (DAKO) at 2 μM concentration. Further experimental detail can be found in the supplemental material.…”

Autoantibody-resistant ADAMTS13 variant

“…In this issue of Blood, Ercig et al report that ADAMTS13 modified by N-glycosylation functions even in the presence of sera obtained from patients with immune-mediated thrombotic thrombocytopenic purpura (iTTP). 1 This variant may be promising for treatment of iTTP patients.…”

“…In this issue of Blood, Magid-Bernstein et al examine the question of the significance of ABO-incompatible platelet transfusion. 1 Like red blood cells, platelets express ABO antigens, and the general practice is to provide ABO-compatible platelets whenever possible. The rationale for this has primarily been concern over the infusion of ABO-incompatible plasma with the platelets and the associated risk of hemolysis in the recipient.…”