The splenic autoimmune response to ADAMTS13 in thrombotic thrombocytopenic purpura contains recurrent antigen-binding CDR3 motifs

Abstract

Key Points• The spleen harbors ADAMTS13-specific memory B cells following acute acquired TTP.• The splenic anti-ADAMTS13 antibody repertoire is characterized by a set of unique and novel CDR3 motifs, 4 shared by 2 patients.Acquired thrombotic thrombocytopenic purpura (TTP) is the consequence of a severe ADAMTS13 deficiency resulting from autoantibodies inhibiting ADAMTS13 or accelerating its clearance. Despite the success of plasma exchange the risk of relapse is high. From 2 patients (A and B), splenectomized for recurrent episodes of acquired TTP, the splenic B-cell response against ADAMTS13 was characterized through generation of human monoclonal anti-ADAMTS13 autoantibodies (mAbs) by cloning an immunoglobulin G (IgG) 4 k-and IgG 4 l-Fab library using phage display technology and by Epstein-Barr virus transformation of switched memory B cells (CD19 1 /CD27 1 /IgG 1 ). Sequence analysis of the anti-ADAMTS13 IgGs of both patients revealed that the V H gene use was limited in our patients to V H 1-3 (55%), V H 1-69 (17%), V H 3-30 (7%), and V H 4-28 (21%) and contained 8 unique and thus far not reported heavy-chain complementarity determining region 3 motifs, of which 4 were shared by the 2 patients. The discovery of several highly similar anti-ADAMTS13 autoantibodies in 2 unrelated TTP patients suggests that the autoimmune response is antigen driven, because the probability that such similar immunoglobulin rearrangements happen by chance is very low (<10 29). (Blood. 2014;124(23):3469-3479)

High-resolution epitope mapping by HX MS reveals the pathogenic mechanism and a possible therapy for autoimmune TTP syndrome

“…4). Among the identified epitopic loops, exosite 3 (motif A) is known to play an important role in antibody binding (8,14,35,36). The role of the other loop motifs has not previously been defined nor the fact that all five loops are required for high-affinity ADAMTS13 binding to its inhibiting autoantibodies (Fig.…”

“…NMR spectroscopy offers a similar level of structural detail but is limited to relatively small proteins that can be expressed with 15 Nand 13 C-labeled residues and prepared in large amounts at high concentration. Peptide arrays can provide information on linear epitopes but with limited structural resolution and will often miss the more common discontinuous epitopes (14). A different approach, the measurement of protein hydrogen exchange together with a peptide separation approach incorporating mass spectrometry analysis [hydrogen-deuterium exchange mass spectrometry (HX MS)] can specify protein-protein interaction regions even for large protein complexes using only a tiny amount of Significance Acquired thrombotic thrombocytopenic purpura (TTP) is primarily caused by autoantibodies that inhibit the ability of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) to proteolyze von Willebrand factor (VWF).…”

Novel therapies in thrombotic thrombocytopenic purpura

“…They found that splenic B cells can produce anti‐ADAMTS13 autoantibodies that can inhibit the protease in the same way as those present in plasma. Interestingly, these two unrelated patients shared four heavy‐chain complementarity determining region 3 motifs 53. At the ISTH Congress in 2017, the same group reported additional findings from two patients that had a splenectomy for iTTP.…”

ADAMTS13 and von Willebrand factor interactions

“…Binding epitopes, including residues Glu376, Gln159–Asp166, Tyr 305–Glu327, and Asn308 in the metalloprotease and disintegrin domains have been identified [91]. However, the most frequent antibody epitope region is found within the cysteine-rich and spacer domain, particularly the spacer domain [75,90–95,96 ▪▪ ,97]. In the mapping of the antigenic regions on ADAMTS13, the most common characteristic is the involvement of five loops located on the spacer domain (Fig.…”