Open ADAMTS13 Conformation in Immune-Mediated Thrombotic Thrombocytopenic Purpura Is Induced By Anti-ADAMTS13 Autoantibodies and Corresponds with an Ongoing ADAMTS13 Pathology

Abstract Background. Deficient ADAMTS13 activity (TS13:act <10%) induced by anti-ADAMTS13 autoantibodies (autoAbs) causes immune-mediated thrombotic thrombocytopenic purpura (iTTP). Recently we showed that an open ADAMTS13 conformation is characteristic for acute iTTP patients, while folded ADAMTS13 was found in 78% of iTTP patients in remission with an TS13:act >50%. However, also iTTP patients in remission with a persistent (<10%) or moderately restored (10-50%) TS13:act have been described, but their ADAMTS13 conformation is unknown. Intriguingly, the factor responsible for inducing open ADAMTS13 in iTTP patients remains elusive. Identifying the cause of open ADAMTS13 in iTTP will help better understand the pathophysiology of iTTP and could help appreciate the prognosis and better manage the prevention of subsequent relapses. Aim. Determine ADAMTS13 conformation in plasma of iTTP patients during acute TTP and remission when TS13:act is <10%, moderately restored (10-50%) or >50% and investigate if anti-ADAMTS13 autoAbs induce conformational changes in ADAMTS13. Methods. TS13:act was determined in 120 iTTP plasma samples from 4 different centers (Marseille, Milan, Budapest, Utrecht). Samples were categorized according to the presence of clinical symptoms (acute versusremission) and their TS13:act in remission (>50%, 10-50%,<10%). Next, ADAMTS13 conformation was determined in all samples using our ADAMTS13 conformation ELISA. Additionally, presence of anti-ADAMTS13 autoAbs was also determined via ELISA. Finally, IgG's from 18 acute iTTP plasma samples were purified and added to folded ADAMTS13 from healthy donor (HD) plasma to test whether iTTP IgG's are able to induce the open HD ADAMTS13 conformation. Results. Of the 120 iTTP plasma samples, 46 were obtained during the acute (clinical signs present) and 74 during the remission phase (clinical signs absent). Further subdividing remission samples showed that TS13:act was >50% in 41, 10-50% in 14 and <10% in 19 samples. ADAMTS13 was open in 98% (45/46) of the acute samples and folded in 71% (29/41) of the remission samples with TS13:act >50%, confirming our previous results. Interestingly, ADAMTS13 was open in 93% and 89% of remission samples with TS13:act 10-50% and <10%, respectively (chi square, P<0.0001). Since anti-ADAMTS13 autoAbs influence TS13:act in iTTP patients, we next could demonstrate that open ADAMTS13 conformation was linked with presence of anti-ADAMTS13 autoAbs (chi square, P<0.0001) suggesting that anti-ADAMTS13 autoAbs could be a factor able to induce an open ADAMTS13 conformation in iTTP patients. To further test this hypothesis, we purified IgG's from 18 acute iTTP plasma's with open ADAMTS13 and added them to HD plasma containing closed ADAMTS13, where 14 of the 18 patient IgG pools (78%) did induce the open conformation in HD ADAMTS13, indicating that patient anti-ADAMTS13 autoAbs indeed can induce conformational changes in ADAMTS13. Conclusion. We show that ADAMTS13 is not only in the open conformation in iTTP patient plasma during the acute phase but also in remission when TS13:act is <10% or 10-50%. Hence, the presence of open ADAMTS13 in those remission patients indicates that the underlying pathophysiology is still ongoing, emphasizing the need for a close monitoring of those patients. In addition, anti-ADAMTS13 autoAbs were identified as a factor responsible for the change in conformation in ADAMTS13 in iTTP. Disclosures Peyvandi: Octapharma US: Honoraria; Kedrion: Consultancy; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Grifols: Speakers Bureau; Sobi: Speakers Bureau; Shire: Speakers Bureau; Novo Nordisk: Speakers Bureau; Octapharma US: Honoraria; Octapharma US: Honoraria; Novo Nordisk: Speakers Bureau; Kedrion: Consultancy; Roche: Speakers Bureau; Novo Nordisk: Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Shire: Speakers Bureau; Sobi: Speakers Bureau; Roche: Speakers Bureau; Roche: Speakers Bureau; Shire: Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Octapharma US: Honoraria; Roche: Speakers Bureau; Roche: Speakers Bureau; Grifols: Speakers Bureau; Sobi: Speakers Bureau; Ablynx: Other: Member of Advisory Board, Speakers Bureau; Sobi: Speakers Bureau; Shire: Speakers Bureau; Sobi: Speakers Bureau; Novo Nordisk: Speakers Bureau; Shire: Speakers Bureau; Kedrion: Consultancy; Novo Nordisk: Speakers Bureau; Grifols: Speakers Bureau; Grifols: Speakers Bureau; Kedrion: Consultancy; Kedrion: Consultancy; Grifols: Speakers Bureau; Octapharma US: Honoraria. Coppo:Ablynx: Consultancy. Veyradier:LFB: Other: Investigator. Vanhoorelbeke:Shire: Consultancy; Ablynx: Consultancy.

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The HLA Variant rs6903608 Is Associated with Disease Onset and Relapse of Immune-Mediated Thrombotic Thrombocytopenic Purpura in Caucasians

Journal of Clinical Medicine ◽

10.3390/jcm9103379 ◽

2020 ◽

Vol 9 (10) ◽

pp. 3379

Author(s):

Ilaria Mancini ◽

Elisa Giacomini ◽

Silvia Pontiggia ◽

Andrea Artoni ◽

Barbara Ferrari ◽

...

Keyword(s):

Thrombotic Thrombocytopenic Purpura ◽

Risk Allele ◽

Absolute Risk ◽

Disease Onset ◽

Close Monitoring ◽

Reference Allele ◽

Thrombocytopenic Purpura ◽

Immune Mediated ◽

The Impact ◽

Risk Of Relapse

Immune-mediated thrombotic thrombocytopenic purpura (iTTP) is a rare, life-threatening thrombotic microangiopathy caused by severe ADAMTS13 (a disintegrin and metalloproteinase with thrombospondin motifs 13) deficiency, recurring in 30–50% of patients. The common human leukocyte antigen (HLA) variant rs6903608 was found to be associated with prevalent iTTP, but whether this variant is associated with disease relapse is unknown. To estimate the impact of rs6903608 on iTTP onset and relapse, we performed a case-control and cohort study in 161 Italian patients with a first iTTP episode between 2002 and 2018, and in 456 Italian controls. Variation in rs6903608 was strongly associated with iTTP onset (homozygotes odds ratio (OR) 4.68 (95% confidence interval (CI) 2.67 to 8.23); heterozygotes OR 1.64 (95%CI 0.95 to 2.83)), which occurred over three years earlier for each extra risk allele (β −3.34, 95%CI −6.69 to 0.02). Of 153 survivors (median follow-up 4.9 years (95%CI 3.7 to 6.1)), 44 (29%) relapsed. The risk allele homozygotes had a 46% (95%CI 36 to 57%) absolute risk of relapse by year 6, which was significantly higher than both heterozygotes (22% (95%CI 16 to 29%)) and reference allele homozygotes (30% (95%CI 23 to 39%)). In conclusion, HLA variant rs6903608 is a risk factor for both iTTP onset and relapse. This newly identified biomarker may help with recognizing patients at high risk of relapse, who would benefit from close monitoring or intensified immunosuppressive therapy.

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Open ADAMTS13, induced by antibodies, is a biomarker for subclinical immune-mediated thrombotic thrombocytopenic purpura

Blood ◽

10.1182/blood.2019004221 ◽

2020 ◽

Cited By ~ 4

Author(s):

Elien Roose ◽

An-Sofie Schelpe ◽

Edwige Tellier ◽

György Sinkovits ◽

Bérangère S Joly ◽

...

Keyword(s):

Thrombotic Thrombocytopenic Purpura ◽

Adamts13 Activity ◽

Thrombocytopenic Purpura ◽

Follow Up Study ◽

Open Conformation ◽

Immune Mediated ◽

Long Term Follow Up ◽

Donor Plasma

Recently, we showed that during the acute phase of immune-mediated thrombotic thrombocytopenic purpura (iTTP), ADAMTS13 circulates in an open conformation. Although the cause of this conformational change in acute iTTP remains elusive, ADAMTS13 is mainly closed in iTTP patients (i) in remission with an ADAMTS13 activity >50% and undetectable anti-ADAMTS13 autoantibodies, and (ii) after rituximab treatment, suggesting a role for anti-ADAMTS13 autoantibodies. Therefore, IgGs from 18 acute iTTP patients were purified and added to closed ADAMTS13 in healthy donor plasma. This resulted in open ADAMTS13 in 14/18 (78%) samples, proving that indeed anti-ADAMTS13 autoantibodies can induce an open ADAMTS13 conformation. To further elucidate the conformation of ADAMTS13 in iTTP patients, we studied a novel iTTP patient cohort (n=197) that also included plasma samples of iTTP patients in remission where ADAMTS13 activity was <50%. The open ADAMTS13 conformation was not only found during acute iTTP but also in patients in remission with an ADAMTS13 activity <50% and in half of the patients with an ADAMTS13 activity >50%, although free anti-ADAMTS13 autoantibodies were not always detected. Thus open ADAMTS13 is not only a hallmark of acute iTTP, but also a novel biomarker to detect subclinical iTTP in patients in remission. Finally, a long term follow-up study in one iTTP patient showed that the open conformation precedes a severe drop in ADAMTS13 activity. In conclusion, we have shown that anti-ADAMTS13 autoantibodies from iTTP patients induce an open ADAMTS13 conformation. Most importantly, an open ADAMTS13 conformation is a biomarker for subclinical iTTP and could become an important tool in TTP management.

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The D173G mutation in ADAMTS-13 causes a severe form of congenital thrombotic thrombocytopenic purpura

Thrombosis and Haemostasis ◽

10.1160/th15-02-0119 ◽

2016 ◽

Vol 115 (01) ◽

pp. 51-62 ◽

Cited By ~ 5

Author(s):

Maria Teresa Pagliari ◽

Andrea Cairo ◽

Safwat Abdel-Azeim ◽

Edrisse Chermak ◽

Ilaria Lazzareschi ◽

...

Keyword(s):

Thrombotic Thrombocytopenic Purpura ◽

Conformational Changes ◽

Clinical Signs ◽

Severe Form ◽

Hek293 Cells ◽

Homozygous Mutation ◽

Thrombocytopenic Purpura ◽

Expression Studies ◽

Congenital Thrombotic Thrombocytopenic Purpura

SummaryCongenital thrombotic thrombocytopenic purpura (TTP) is a rare form of thrombotic microangiopathy, inherited with autosomal recessive mode as a dysfunction or severe deficiency of ADAMTS-13 (A Disinte-grin And Metalloprotease with ThromboSpondin 1 repeats Nr. 13), caused by mutations in the ADAMTS-13 gene. About 100 mutations of the ADAMTS-13 gene were identified so far, although only a few char-acterised by in vitro expression studies. A new Asp to Gly homozygous mutation at position 173 of ADAMTS-13 sequence was identified in a family of Romanian origin, with some members affected by clinical signs of TTP. In two male sons, this mutation caused a severe (< 3 %) deficiency of ADAMTS-13 activity and antigen level, associated with periodic thrombocytopenia, haemolytic anaemia and mild mental confusion. Both parents, who are cousins, showed the same mutation in heterozygous form. Expression studies of the mutant ADAMTS-13, performed in HEK293 cells, showed a severe decrease of the enzyme’s activity and secretion, although the protease was detected inside the cells. Molecular dynamics found that in the D173G mutant the interface area between the metalloprotease domain and the disintegrin-like domain significantly decreases during the simulations, while the proline-rich 20 residues linker region (LR, 285–304) between them undergoes extensive conformational changes. Inter-domain contacts are also significantly less conserved in the mutant compared to the wild-type. Both a decrease of the inter-domain contacts along with a substantial conformational rearrangement of LR interfere with the proper maturation and folding of the mutant ADAMTS-13, thus impairing its secretion.Supplementary Material to this article is available online at www.thrombosis-online.com.

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Alternate-Day Dosing of Caplacizumab for Immune-Mediated Thrombotic Thrombocytopenic Purpura

Blood ◽

10.1182/blood-2021-151283 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2087-2087

Author(s):

Lucas Kühne ◽

Dennis A. Eichenauer ◽

Paul Knöbl ◽

Linus A. Völker ◽

Jessica Kaufeld ◽

...

Keyword(s):

Thrombotic Thrombocytopenic Purpura ◽

Autoimmune Disorder ◽

Daily Treatment ◽

Treatment Modalities ◽

Close Monitoring ◽

Dosing Regimen ◽

Thrombocytopenic Purpura ◽

Normal Platelet ◽

Immune Mediated ◽

Risk Patients

Abstract Background: The anti-VWF nanobody caplacizumab directly prevents the fatal microthrombi formation in immune-mediated thrombotic thrombocytopenic purpura, thereby adding a new therapeutic principle to the treatment of this autoimmune disorder. However, real-world treatment modalities beyond clinical trials remain heterogeneous. Methods: Here, we describe the risks and benefits of an alternate-day dosing regimen for caplacizumab, by thoroughly analyzing the timing and outcome of this treatment approach in a retrospective cohort of 25 iTTP patients treated with caplacizumab in Austria and Germany between 2018 and 2021. Results: Alternate-day dosing of caplacizumab appeared feasible and led to persisting normal platelet counts in the majority of patients, who were converted after a median time of 17 days daily treatment. Four patients experienced iTTP exacerbations or relapses that led to the resumption of daily caplacizumab application and/or other therapies. VWF activity was repeatedly measured in 17 out of 25 patients and documented sufficient suppression by caplacizumab after 24 and 48 hours, in line with published pharmacokinetics. Conclusion: Extension of caplacizumab injection intervals from daily to alternate-day dosing may be safely considered in selected patients after 30 days of daily treatment. Earlier modifications may be considered in low-risk patients, but require close monitoring for clinical and laboratory features of thrombotic microangiopathy. Disclosures Völker: Sanofi-Genzyme: Honoraria, Other: counselling fees.

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