Peak ADAMTS13 activity to assess ADAMTS13 conformation and risk of relapse in immune-mediated thrombotic thrombocytopenic purpura.

ABSTRACT: Previous studies have demonstrated that >38% of patients with immune-mediated thrombotic thrombocytopenic purpura in remission with activity >50% had an open ADAMTS13 (a disintegrin and metalloproteinase with thrombospondin type 1 motif, member 13) conformation. We assessed ADAMTS13 conformation in remission (ADAMTS13 activity >60%), focusing on peak ADAMTS13 activity levels and longitudinal assessment in 420 samples across 157 patients. Fewer cases had an open conformation at peak ADAMTS13 activity than unselected remission samples with ADAMTS13 activity >60% (23% vs 43%). Patients with a closed ADAMTS13 conformation at peak ADAMTS13 activity had an eightfold lower relapse rate in the subsequent year (9% vs 46%) and a fivefold lower relapse rate within 2 years (23% vs 62%) compared with cases with an open conformation. Patients with an open conformation at peak ADAMTS13 activity required preemptive anti-CD20 treatment earlier than those with a closed conformation (median, 10 vs 25 months). Longitudinally, an open conformation was evident at, and often preceded relapse. When the conformation was already open before relapse, an increase in the conformation index at relapse was seen despite the undetectable anti-ADAMTS13 immunoglobulin G (IgG) antibody. In cases with detectable anti-ADAMTS13 IgG antibody, these became undetectable before achieving a closed conformation, highlighting the relapse risk even with undetectable anti-ADAMTS13 IgG antibody and the clinical utility of open/closed during monitoring. To our knowledge, this is the first study to show an association between relapse risk and ADAMTS13 conformation when activity levels are at a peak. The open conformation identifies antibody-mediated subclinical disease that is not detectable by the current ADAMTS13 testing.

ADAMTS13 activity testing: evaluation of commercial platforms for diagnosis and monitoring of thrombotic thrombocytopenic purpura.

Background: ADAMTS13 activity is one of the key investigations needed to diagnose thrombotic thrombocytopenic purpura, and there are a number of different assays available to measure it. HemosIL AcuStar, a chemiluminescent immunoassay, was developed and used as a quicker, automated test. In clinical practice, discrepancies between AcuStar and the gold standard FRETS-VWF73 have been documented in a manner that would affect diagnosis and treatment. Objectives: We aimed to identify and highlight clinical situations where this discrepancy occurs and to attempt to determine the cause. Method: Therefore, we undertook a study to compare the FRETS-VWF73 assay with AcuStar, the Technozym Activity ELISA, and Ceveron FRET assays using a mixture of 94 retrospective and prospective patient samples. Results: We found that although the concordance between FRETS-VWF73 and the other methods was generally very good, discrepancies were found in a small number tested on AcuStar affecting diagnosis (5 of 32) and follow-up (7 of 51). A Wilcoxon test comparing FRETS-VWF73 to the AcuStar results suggested that the AcuStar results were significantly lower in 42 samples tested on all 4 platforms. We investigated potential causes for this difference by testing the impact of high vWF levels and addition of a monoclonal ADAMTS13 autoantibody (3H9) to samples. We found no impact of high vWF levels on interassay variability but found that 3H9 reduced ADAMTS13 activity levels much more in AcuStar and ELISA assays than in FRETS assays. Conclusion: Based on our findings, we would suggest that when AcuStar is used upfront to guide management, a second testing method should be used in patients with an atypical thrombotic thrombocytopenic purpura presentation or unexpectedly slow ADAMTS13 recovery.

Alterations in B- and circulating T-follicular helper cell subsets in immune thrombotic thrombocytopenic purpura.

T follicular helper (Tfh) cells regulate development of antigen-specific B-cell immunity. We prospectively investigated B-cell and circulating Tfh (cTfh) cell subsets in 45 patients with immune thrombotic thrombocytopenic purpura (iTTP) at presentation and longitudinally after rituximab (RTX). B-cell phenotype was altered at acute iTTP presentation with decreased transitional cells and post-germinal center (post-GC) memory B cells and increased plasmablasts compared with healthy controls. A higher percentage of plasmablasts was associated with higher anti-ADAMTS13 IgG and lower ADAMTS13 antigen levels. In asymptomatic patients with ADAMTS13 relapse, there were increased naïve B cells and a global decrease in memory subsets, with a trend to increased plasmablasts. Total circulating Tfh (CD4+CXCR5+) and PD1+ Tfh cells were decreased at iTTP presentation. CD80 expression was decreased on IgD+ memory cells and double-negative memory cells in acute iTTP. At repopulation after B-cell depletion in de novo iTTP, post-GC and double-negative memory B cells were reduced compared with pre-RTX. RTX did not cause alteration in cTfh cell frequency. The subsequent kinetics of naïve, transitional, memory B cells and plasmablasts did not differ significantly between patients who went on to relapse vs those who remained in remission. In summary, acute iTTP is characterized by dysregulation of B- and cTfh cell homeostasis with depletion of post-GC memory cells and cTfh cells and increased plasmablasts. Changes in CD80 expression on B cells further suggest altered interactions with T cells.