Autoimmune Heparin-induced Thrombocytopenia Research Articles

Thrombopénies et thromboses induites par les anticorps anti-FP4

AbstractThrombocytopenia and thromboses induced by antibodies to platelet factor 4 (PF4) constitute a heterogeneous group of rare but severe clinical and biological syndromes, including classical heparin-induced thrombocytopenia (HIT), and more rarely autoimmune HIT, vaccine-induced immune thrombotic thrombocy-topenia (VITT), and, recently, VITT-like syndromes. These pathologies all induce major hypercoagulability, related to blood cell activation by anti-PF4 antibodies, leading to potentially severe thromboses, which require urgent treatment. The severity of thrombocytopenia and thromboses, and their location, vary with the disease; autoimmune HIT and VITT are associated with particularly aggressive forms, including cerebral venous thrombosis. The diagnosis of thrombocytope-nia and thromboses induced by anti-PF4 antibodies relies on a combination of clinical and biological factors, including the detection of anti-PF4 antibodies by immunological assays and, in most cases, confirmation of their pathogenicity by platelet activation tests. As the performance of these tools varies greatly depending on the pathology, the biologist plays a major role in the choice of the assays, and their interpretation, to ensure correct diagnosis and appropriate management of the patient.

Thrombotic anti-PF4 immune disorders: HIT, VITT, and beyond.

Antibodies against the chemokine platelet factor 4 (PF4) occur often, but only those that activate platelets induce severe prothrombotic disorders with associated thrombocytopenia. Heparin-induced thrombocytopenia (HIT) is the prototypic anti-PF4 disorder, mediated by strong activation of platelets through their FcγIIa (immunoglobulin G [IgG]) receptors (FcγRIIa). Concomitant pancellular activation (monocytes, neutrophils, endothelium) triggers thromboinflammation with a high risk for venous and arterial thrombosis. The classic concept of HIT is that anti-PF4/heparin IgG, recognizing antigen sites on (cationic) PF4 that form in the presence of (anionic) heparin, constitute the heparin-dependent antibodies that cause HIT. Accordingly, HIT is managed by anticoagulation with a nonheparin anticoagulant. In 2021, adenovirus vector COVID-19 vaccines triggered the rare adverse effect "vaccine-induced immune thrombotic thrombocytopenia" (VITT), also caused by anti-PF4 IgG. VITT is a predominantly heparin-independent platelet-activating disorder that requires both therapeutic-dose anticoagulation and inhibition of FcγRIIa-mediated platelet activation by high-dose intravenous immunoglobulin (IVIG). HIT and VITT antibodies bind to different epitopes on PF4; new immunoassays can differentiate between these distinct HIT-like and VITT-like antibodies. These studies indicate that (1) severe, atypical presentations of HIT ("autoimmune HIT") are associated with both HIT-like (heparin-dependent) and VITT-like (heparin-independent) anti-PF4 antibodies; (2) in some patients with severe acute (and sometimes chronic, recurrent) thrombosis, VITT-like antibodies can be identified independent of proximate heparin exposure or vaccination. We propose to classify anti-PF4 antibodies as type 1 (nonpathogenic, non- platelet activating), type 2 (heparin dependent, platelet activating), and type 3 (heparin independent, platelet activating). A key concept is that type 3 antibodies (autoimmune HIT, VITT) require anticoagulation plus an adjunct treatment, namely high-dose IVIG, to deescalate the severe anti-PF4 IgG-mediated hypercoagulability state.

Heparin-induced thrombocytopenia (and autoimmune heparin-induced thrombocytopenia): an illustrious review

Heparin-induced thrombocytopenia (and autoimmune heparin-induced thrombocytopenia): an illustrious review

Heparin-induced thrombocytopenia: An illustrated review

Heparin-induced thrombocytopenia (HIT) is an immune-mediated adverse drug effect from unfractionated or low-molecular-weight heparin that results in thrombocytopenia and potentially catastrophic thrombosis. HIT occurs due to the development of platelet-activating antibodies against multimolecular complexes of platelet factor 4 and heparin. Given the frequency of thrombocytopenia and heparin use among hospitalized patients, calculation of the 4Ts Score is recommended to identify patients at increased likelihood of HIT and direct further evaluation. In patients with an intermediate or high probability 4Ts Score, an immunoassay and functional assay are recommended to confirm or refute the diagnosis of HIT. Heparin avoidance and initiation of nonheparin anticoagulation are the mainstays of acute HIT management. In this illustrated review, we provide visual summaries of the diagnosis and management of HIT, highlighting connections between pathophysiology and clinical care as well as summarizing efforts in quality improvement in the field. We further emphasize common pitfalls and pearls in diagnosis and management to encourage evidence-based care. We include graphical representation of the unique challenges of HIT with cardiopulmonary bypass and also delineate autoimmune HIT and its subtypes.

Investigation of anti-PF4 versus anti-PF4/heparin reactivity using fluid-phase enzyme immunoassay for 4 anti-PF4 disorders: classic heparin-induced thrombocytopenia (HIT), autoimmune HIT, vaccine-induced immune thrombotic thrombocytopenia, and spontaneous HIT

BackgroundFour platelet-activating anti-platelet factor 4 (PF4) disorders have been recognized: classic heparin-induced thrombocytopenia (cHIT), autoimmune heparin-induced thrombocytopenia (aHIT), spontaneous heparin-induced thrombocytopenia (SpHIT), and vaccine-induced immune thrombotic thrombocytopenia (VITT). All test immunoglobulin G (IgG) positive using solid-phase enzyme immunoassay (solid-EIA) against PF4/heparin (PF4/H) and/or PF4 alone. Fluid-phase EIA (fluid-EIA) should better discriminate between anti-PF4 and anti-PF4/H antibodies since conformationally altered PF4 bound to solid phase is avoided. ObjectivesTo compare anti-PF4 vs anti-PF4/H antibody profiles for anti-PF4 disorders using solid- and fluid-EIA. MethodsWe developed a novel fluid-EIA to measure anti-PF4 vs anti-PF4/H antibodies. ResultsUsing fluid-EIA, 27 of 27 (100%) cHIT sera tested IgG positive with PF4/H, but only 4 of 27 (14.8%) tested positive against PF4 alone; all 27 exhibited heparin-enhanced binding. In contrast, 17 of 17 (100%) VITT sera tested IgG positive against PF4 alone, with markedly reduced binding against PF4/H; this distinct VITT antibody profile was not evident using solid-EIA. All 15 aHIT sera and all 11 SpHIT sera tested IgG positive against PF4 alone, with variable reactivity in PF4/H-EIA (heparin-enhanced binding in 14 of 15 and 10 of 11 aHIT and SpHIT sera, respectively). Remarkably, 1 SpHIT patient with a VITT-mimicking fluid-EIA profile (PF4 >> PF4/H) also clinically resembled patients with VITT (postviral cerebral vein/sinus thrombosis), with anti-PF4 reactivity correlating inversely with platelet count recovery; moreover, the single aHIT patient with a VITT-mimicking fluid-EIA profile also developed postviral cerebral vein/sinus thrombosis. ConclusioncHIT and VITT sera showed opposite fluid-EIA profiles (cHIT: PF4/H >> PF4, with most testing negative against PF4 alone; VITT: PF4 >> PF4/H, with most testing negative against PF4/H). In contrast, all aHIT and SpHIT sera reacted against PF4 alone but with variable (usually enhanced) reactivity against PF4/H. VITT-mimicking clinical/serologic profiles occurred in only a minority of patients with SpHIT and aHIT.

Platelet factor 4 triggers thrombo-inflammation by bridging innate and adaptive immunity.

Platelet factor 4 (PF4, synonym: CXCL4) is an evolutionary old chemokine with proposed roles in hemostasis and antimicrobial defense. In addition, PF4 has attracted considerable attention as a crucial mediator of one of the most prothrombotic adverse drug effects affecting blood cells, heparin-induced thrombocytopenia (HIT). Interest in PF4 substantially increased in 2021 when it was identified as the target antigen in the life-threatening adverse effect, vaccine-induced immune thrombotic thrombocytopenia (VITT). We address the concept that a major biological function of PF4-a strongly cationic chemokine-is to bind to negatively-charged prokaryotic microorganisms, resulting in structural changes in PF4 that trigger a danger signal recognized by the adaptive immune system. Application of biophysical tools has provided substantial insights into the molecular mechanisms by which PF4 becomes immunogenic, providing insights into a new mechanism of autoimmunity. Binding of autoantibodies with high affinity induces conformational change(s) in the endogenous protein, which are then recognized as foreign antigen, as exemplified by the prothrombotic disorders, autoimmune HIT and VITT. The final part of our review summarizes current assays for HIT and VITT, explaining how structural aspects of anti-PF4 pathobiology relate to assay design and performance characteristics. Currently, functional (platelet activation) assays using washed platelets detect HIT antibodies when heparin is added, and VITT antibodies when PF4 is added. Solid-phase PF4-dependent immunoassays using microtiter plates are sensitive for both HIT and VITT antibodies, while rapid immunoassays, in which the PF4/heparin antigen is coated on beads, are sensitive and specific for HIT, but not for VITT antibodies.

"Autoimmune HIT", a Potential Diagnostic Artifact Due to Remnant Heparin in HIT Samples, Is Not Associated with More Severe Outcomes

In recent years, there has been increasing use of the term "autoimmune" heparin-induced thrombocytopenia (HIT) to describe patients with HIT antibodies that activate platelets in the absence of exogenous heparin (i.e., the "zero heparin" serotonin release assay, SRA), with the proposed implication that such patients present with more severe phenotypes. Platelet activation in the SRA in the absence of exogenous heparin is likely a real phenomenon in rare/infrequent patients who did not receive proximate heparin therapy (Spontaneous HIT) and in patients who develop HIT after heparin treatment has ceased (delayed-onset HIT). However, in other patients deemed to have "autoimmune" HIT based on reactivity in the "zero heparin" SRA, we hypothesized that platelet reactivity in this SRA test system might result from remnant heparin in the patient's diagnostic blood sample. To further complicate the interpretation of autoimmune HIT testing, there is a lack of rigorous data demonstrating a correlation between this diagnosis and disease severity. This study was undertaken to evaluate the technical validity and clinical relevance of an autoimmune HIT diagnosis. First, thirty "real-world" diagnostic HIT patient samples were subjected to anti-Xa testing. Remarkably, fourteen of the 30 samples (47%) had detectable heparin, with a range of 0.11-0.55 IU/mL for thirteen samples, while one sample had heparin <0.1 IU/mL (data not shown due to Abstract 2-figure panel limit). Next, to determine the extent to which low concentrations of unfractionated or low molecular weight heparin spiked into patient samples impact reactivity in the SRA, spiked samples were tested in the "conventional" low dose heparin (0.1 IU/ml) SRA and the "zero heparin" SRA (autoimmune HIT test). Fig 1A demonstrates that even ultra-low concentrations of unfractionated heparin (0.06 IU/mL) spiked into the sample facilitate serotonin release in the "zero heparin" SRA. On the other hand, the conventional SRA was not impacted by heparin spiked into samples unless concentrations were 5 IU/mL or higher, several-fold greater than what would be seen in patient samples. Similarly, LMWH induced platelet reactivity in the zero heparin SRA at concentrations between 0.25-1.0 IU/mL, while there was no effect on the conventional SRA at these concentrations (data not shown). As proof-of-concept that low levels of heparin mediate reactivity in a zero-heparin functional test, a HIT patient sample devoid of heparin was spiked with 0.05 IU/ml of heparin and evaluated with and without digestion with heparinase I. Heparin-spiked samples induced platelet activation in a zero-heparin functional assay which was abrogated upon heparinase I digestion (data not shown). Together, these results are consistent with the hypothesis that at least some cases of "autoimmune HIT" are labeled as such due to residual heparin present in patient samples rather than due to the presence of a qualitatively different ("autoimmune") HIT antibody. To evaluate the clinical relevance of an "autoimmune" HIT diagnosis, platelet nadir and the presence of thrombosis were interrogated as markers of disease severity in a cohort of 48 HIT patient samples with unknown heparin levels. Thirty-six patients were positive in the zero heparin SRA ("Autoimmune HIT" cohort; >50% serotonin release), while 12 were negative ("Classical HIT" cohort; positive in the conventional SRA, but <50% serotonin release in the zero heparin SRA). Fig 1B demonstrates that there was no difference in platelet nadir between the two groups (p=0.562); further, there were no differences between these groups in rates of confirmed thrombosis/skin necrosis and acute system reactions (p=0.324; data not shown). In summary, "autoimmune HIT" diagnostic testing (zero-heparin SRA) should not be performed until samples are confirmed to be devoid of heparin, and rigorous large-scale studies are needed to evaluate whether results of validly-conducted autoimmune HIT tests correlate with disease severity. Figure 1. (A) Low concentrations of spiked heparin in a HIT patient sample facilitates platelet activation in the "autoimmune" HIT test (zero-heparin SRA; red line) but does not affect the conventional SRA (black line). (B) Platelet nadir was similar between "autoimmune" (n=36) and conventional HIT (n=12) cohorts. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Vaccine-Induced Immune Thrombotic Thrombocytopenia: Where does The PF4 Fit?

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a severe and usually fatal syndrome with an aggressive thrombotic process in unusual sites (notably, sinus veins of the brain and/or splanchnic veins) that accompanies profound thrombocytopenia. If not well managed, VITT may progress to a more severe systemic disease such as disseminated intravascular coagulation. VITT was associated with two adenoviral vector vaccines, mainly ChAdOx1 nCoV-19 (AstraZeneca) and Ad26.COV2.S (Janssen/ Johnson &amp; Johnson) was first reported in late February 2021 and mid-April 2021, respectively. This thrombotic phenomenon closely resembled that of autoimmune heparin-induced thrombocytopenia. Yet, in 2021, the MHRA (Medicines &amp; Healthcare products Regulatory Agency) and the European Medicines Agency (EMA) claim no association between thrombosis and vaccines. Not long after, three scientific societies from Norway, Germany, and the UK reported in the press and social media the detection of thrombocytopenia with cerebral venous sinus thrombosis (CVST) and anti-platelet factor 4 (anti-PF4) antibodies in patients succeeding adenovirus-based vaccination. The immune response against PF4; (also known as CXCL4) in VITT is probably triggered by the proinflammatory milieu; compounds such as human cell-line proteins, non-assembled adenoviral proteins, and potentially EDTA (edetic acid) could be contributing to the prothrombotic state. This review will address functional aspects of PF4 in the thromboinflammatory phenomena, its role in the current anti-SARS-CoV-2 adenoviral-based vaccines, and evidence for its role in triggering VITT. Physicians as well as the public need to be aware of this new disease to quickly provide accurate diagnosis and timely treatment.

PB2302: HEPARIN-INDUCED THROMBOCYTOPENIA ANTIBODIES CAN MAINTAIN PLATELET-ACTIVATING PROPERTIES AFTER TWO SESSIONS OF THERAPEUTIC PLASMA EXCHANGE

Background: Therapeutic plasma exchange (TPE) has been used as a salvage therapy in a small subset of heparin-induced thrombocytopenia (HIT) patients (severe or refractory HIT, before heparin re-exposure during cardiac surgery, in bleeding patients and in autoimmune HIT). Existing data show that HIT antibodies can lose platelet-activating properties after one or two TPE sessions, while more sessions may be needed to decrease the antibody titer. However, results may be assay dependent. The utility of Heparin-Induced Multiple aggregometry (HIMEA) in this setting has not been evaluated. Aims: To assess the effect of TPE on pathogenic HIT antibodies using HIMEA before an emergency surgery for gynecologic malignancy. Methods: TPE was performed in a 72-year-old patient necessitating emergency surgery for a voluminous uterine sarcoma in the acute phase of HIT (10 days after diagnosis). TPE aimed to reduce post-operative thrombotic risk by decreasing pathogenic HIT antibodies in a patient presenting a high risk of post-operative thrombotic complications. Two consecutive sessions of TPE using Spectra Optia ® Apheresis System were performed immediately before surgery. During each plasma exchange 2 L of plasma were removed and replaced by 5% human albumin. Acid-Citrate-Dextrose (ACD) was used as anticoagulant. Blood samples were taken before and after each TPE session (TPE1 pre, TPE1 after, TPE2 pre, TPE2 after). Removed patient’s plasma collected to the plasmapheresis bag was also tested. IgG anti-PF4/heparin antibodies tested with Enzyme Immunosorbent Assay (EIA) (Hyphen ZYMUTEST™ HIA IgG, Hyphen Biomed) and Heparin-Induced Multiple aggregometry (HIMEA) (Multiplate, Roche) in the absence of heparin (HIMEA saline), in the presence of 1UI/ml unfractionated heparin (UFH) (HIMEA low) and in the presence of 100IU/ml UFH (HIMEA high) were assessed. Results: Patient’s 4T’s score was 7 (2 for platelet count fall, 2 for timing, 2 for catheter associated venous thrombosis and 1 for other causes), strongly positive IgG anti-PF4 antibodies (OD: 2.43, cut-off<0.3 units) and a HIMEA profile favoring HIT (typical sigmoid curve at HIMEA low, >50% decrease of AUC at HIMEA high, absence of platelet aggregation at HIMEA saline). She was treated with fondaparinux at 1.5 mg/ day due to sepsis-associated acute kidney injury requiring hemodialysis. Platelet count (PLT) were 168 G/l before surgery. After two TPE sessions antibody titers remained unchanged (TPE1 pre: 2.63 units, TPE1 after:2.19 units, TPE2 pre:2.51 units, TPE2 after:5.52 units). Moreover, platelet reactivity remained also strong as demonstrated by high area under the curve (AUC) values in HIMEA low (figure 1). Plasma from the collection bag showed strong EIA and platelet activation test reactivity. No thrombosis nor thrombocytopenia were observed in one month follow-up. Image:Summary/Conclusion: Two TPE sessions failed to reduce platelet-activating properties of HIT antibodies as assessed by HIMEA in a cancer patient. TPE could be considered as a therapeutic intervention that can reduce thrombotic risk in patients necessitating an urgent non-cardiac surgery in the acute phase of HIT. However, more data is needed on the clinical benefit and the optimal number of TPE sessions in this clinical setting. Functional assays assessing Fcγ receptor mediated platelet activation may help guide the timing of the surgery.

Novel disease entities due to antiplatelet factor 4 antibodies: HIT, aHIT, and VITT

Antiplatelet factor 4 (PF4) antibodies, also known as anti-PF4/heparin complex antibodies, are measured to diagnose heparin-induced thrombocytopenia (HIT). In HIT, anti-PF4 antibodies induced by heparin exposure cause thrombocytopenia and thrombosis. However, in recent years, autoimmune HIT (aHIT) that develops without heparin exposure has been getting attention. In 2021, anti-PF4 antibodies were reported to cause the fatal vaccine-induced immune thrombotic thrombocytopenia (VITT) that developed after adenoviral vector vaccination for COVID-19. HIT, aHIT, and VITT are considered to be caused by anti-PF4 antibodies, and their pathological conditions are similar. However, they have different levels of severity, and the detection sensitivity of their antibodies varies depending on the assay. Herein, we review three pathologies, namely, HIT, aHIT, and VITT, associated with anti-PF4 antibodies.

Solid organ procurement and transplantation from deceased donors with vaccine-induced thrombosis and thrombocytopenia

In late February 2021, the first cases of vaccine-induced thrombosis and thrombocytopenia (VITT) were observed in patients after immunization with the ChAdOx1 nCoV-19 adenoviral vector vaccine (Oxford, AstraZeneca) against SARS-CoV-2.1 This prothrombotic response is presumed to be mediated by anti-platelet factor 4 (PF4) antibodies that activate platelets and induce activation and extracellular traps formation of neutrophils.2 These antibodies can also be seen in autoimmune heparin-induced thrombocytopenia, but all the described cases had no previous exposure to heparin.

No Diagnostic Utility of Zero Heparin Control Buffer in Serotonin Release Assay: Findings from a Validation Study

Background: Heparin induced thrombocytopenia (HIT) is a rare immune mediated complication that is triggered in a subset of patients temporal to therapeutic heparin exposure. Laboratory testing is based on screening for the presence of serum anti-PF4 antibodies using sensitive solid-phase immunoassays. If antibodies are detected, functional testing to demonstrate the platelet activating properties and heparin dependence of these immune complexes is then performed. Previous studies have reported possible clinical utility in identifying non heparin dependent platelet activating antibodies using a buffer control with zero heparin in the serotonin release functional assay (SRA). These reports suggested a correlation between reactivity in the zero heparin buffer control and pathogenicity of HIT antibodies which may define a subtype of HIT, referred to as autoimmune HIT. We aimed to investigate the utility of zero heparin buffer control as a part of an inhouse validation study of a mass spectrometry-coupled SRA (Mayo-SRA).Methods: Three hundred archived serum samples were tested using anti-PF4 IgG antibody enzyme-linked immunosorbent assay (ELISA; Immucor Diagnostics, GA, USA). SRA was preformed on all samples using Mayo-SRA and reference 14C SRA methods. Zero heparin control buffer was included in the Mayo-SRA assay. Serotonin release >20% in the low dose heparin (0.1U/mL, LDH) and ELISA optical density (OD) >0.4 were considered positive. Drug interference studies were performed by spiking known SRA-positive samples with increasing concentrations of unfractionated heparin (UFH), low molecular weight heparin (LMWH) or fondaparinux in the LDH and zero heparin SRA. The clinical 4T score was calculated retrospectively calculated for all patients.Results: Of the 300 tested samples, 57 were anti-PF4 ELISA positive. 33 of the 57 samples were positive using the reference 14C SRA method. Whereas 43 samples were positive by Mayo-SRA assay. Three additional samples were positive by Mayo-SRA, but negative by both screening anti-PF4 ELISA and the reference SRA method (Fig- 1A). Lastly, 13 samples were anti-PF4 ELISA positive, but SRA negative by both methods in comparison. Interestingly, 44 of 46 (95%) samples interpreted positive by LDH were also interpreted positive (serotonin release >20%) under zero heparin conditions. These included the 3 samples that were positive by Mayo-SRA but negative by both screening anti-PF4 ELISA and the reference SRA. The overall % serotonin release using zero heparin control was significantly lower (P= 0.003, paired Student T-test) compared to LDH (Fig-1B). In addition, zero heparin followed a similar pattern as LDH, with highest levels at ELISA OD units >2 (Fig-1C).Strikingly, drug interference studies showed artifactual serotonin release in the zero heparin reaction, which was not detected in the absence of spiked drugs. For UFH, serotonin release in zero heparin control occurred at very low spiked concentrations, ≥0.063 U/mL. LMWH and fondaparinux spiking experiments also displayed similar zero heparin serotonin release patterns (Fig-1D). Of note, none of these interferences were detected in UFH spiked SRA-negative samples (data not shown).Conclusion: Contrary to prior results suggesting that less than 50% of LDH SRA positive samples are also positive in the zero heparin SRA, our results show high zero heparin SRA positivity rate of >95%. Zero heparin SRA showed a pattern with highest levels at ELISA OD units >2 suggesting that reactivity in this condition is a function of antibody strength rather than a qualitative difference (i.e. “autoimmune” HIT antibodies vs “non-autoimmune” HIT antibodies). In addition, contamination of patient sera with small amounts of remnant heparin can significantly impact platelet activation in the zero heparin SRA test. Thus, zero heparin SRA positive results may be artifactual and represent residual heparin contained in the patient sample.Figure legend:Fig-1A. Scatter plots of SRA positive samples grouped by 4T scores. Red dots are Mayo-SRA only positive samples. Fig-1B and Fig-1C. Scatter plots of % serotonin release of Mayo-SRA positive samples at LDH (circles) vs. zero heparin buffers (squares), and grouped by ELISA OD values, respectively. Fig-1D. % Serotonin release of known SRA-positive samples spiked with increasing concentrations of UFH, LMWH, or fondaparinux at LDH (white) or zero heparin buffers (red). [Display omitted] DisclosuresPruthi: CSL Behring: Honoraria; Genentech: Honoraria; Bayer Healthcare AG: Honoraria; HEMA Biologics: Honoraria; Instrumentation Laboratory: Honoraria; Merck: Honoraria. Padmanabhan: Veralox Therapeutics: Membership on an entity's Board of Directors or advisory committees.

Relapsed Refractory Acquired Thrombotic Thrombocytopenic Purpura (aTTP) Following COVID-19 Vaccination

Introduction:Acquired thrombotic thrombocytopenic purpura (aTTP) due to an acquired deficiency in the enzyme ADAMTS13 leads to ultra-large von Willebrand multimers, thrombocytopenia and microangiopathic hemolytic anemia. Complications include microvascular and macrovascular thrombosis. We present an unusual case of a patient with a history of refractory aTTP who experienced relapsed aTTP following COVID-19 vaccine.Case Description:A 57-year-old African-American male with a history of refractory aTTP experienced a relapse following 3 years of remission after receiving COVID-19 vaccination. The patient was initially diagnosed with aTTP in 2016, after presenting with symptoms of dark urine, mild headaches and transient episodes of aphasia and paresthesia. Due to symptoms and persistently low ADAMTS13 levels, he required prolonged and extensive treatment including over 5 weeks of daily therapeutic plasma exchange (TPE), followed by gradual reduction in frequency of TPE sessions, as well as trials of rituximab, eculizumab, steroids, mycophenolate mofetil and bortezomib. Ultimately, he achieved remission after 9 months of intermittent TPE, 3 months of weekly bortezomib 1 mg/m 2, mycophenolate mofetil up-titrated to 1,750 mg twice daily, and then slowly tapered off over a 2-year period. The patient was doing well for 3 years without manifestations of aTTP (2 years off all therapeutics), until he developed a petechial rash 7 weeks after receiving the second dose of the Moderna COVID-19 vaccine. He was found to have acute thrombocytopenia with platelets of 38 x 10 9/L (normal range 135-317 x 109/L), from a baseline of 200-300 x 10 9/L. He was referred to the emergency department, where additional labs were notable for mildly elevated LDH of 508 U/L (normal range 122-222 U/L), hemoglobin of 12.4 g/dL (normal range 13.2-16.6 g/dL), creatinine at baseline, and peripheral blood smear showing 1-3 schistocytes per high-powered field. ADAMTS13 activity level was t <5% (normal >/= 70%), with positive ADAMTS13 inhibitor screen and titer of 1.5 (normal <0.4), consistent with relapsed aTTP.The patient was admitted to the hospital, and initiated on daily TPE, with steroids and diphenhydramine prior to each TPE session. He quickly improved with TPE alone , but given his history of refractory aTTP, he was discharged on weekly rituximab for 4 weeks and caplacizumab 11 mg daily for 30 days. His platelets remained stable within the upper limit of normal during his 30 day course of caplacizumab. However, 3 weeks after completion of caplacizumab, he had an acute drop in his platelets to 23 x 10 9/L. His ADAMTS13 level was again found to be <5%, and inhibitor level was the highest that it had ever been at 11.4. He was again hospitalized and underwent 8 sessions of daily TPE, as well as re-initiation of caplacizumab, mycophenolate mofetil 500 mg bid (with increasing taper), and a prednisone taper. Intravenous Cyclophosphamide 750 mg/m 2 was also added every 3 weeks. With this regimen, patient's platelet count normalized and remain stable, and his ADAMTS13 activity level has reached 52-59%.Discussion:Cases of vaccine-induced immune thrombotic thrombocytopenia (VITT) have been described as a complication following vaccination with formulations containing replication-defective adenoviral vectors (AstraZeneca-Oxford and Johnson&Johnson COVID-19 vaccines)(Arepally and Ortel 2021, Simpson, Shi et al. 2021). VITT and aTTP are both immune-mediated, however, VITT is distinct and pathogenically linked to autoimmune heparin-induced thrombocytopenia (HIT), given the presence of anti-platelet factor 4 antibodies in these patients, whereas aTTP is due to reduction in ADAMTS13 level, secondary to an antibody inhibitor of ADAMTS13 (Arepally and Ortel 2021). Recently, cases have been reported of de novo aTTP developing shortly after COVID-19 vaccination with all available vaccines, except the Moderna (mRNA-1273) vaccine (Al-Ahmad, Al-Rasheed et al. 2021, de Bruijn, Maes et al. 2021, Maayan, Kirgner et al. 2021, Ruhe, Schnetzke et al. 2021, Waqar, Khan et al. 2021, Yocum and Simon 2021). Additionally, cases of relapsed aTTP have been described following only the BNT162B2 (Pfizer-BioNTech) vaccine (Maayan, Kirgner et al. 2021, Sissa, Al-Khaffaf et al. 2021). This is the first case, to our knowledge, reported in the literature of aTTP following vaccination with Moderna's mRNA-1273 vaccine. DisclosuresNo relevant conflicts of interest to declare.

Monoclonal and Oligoclonal Anti-PF4 Antibodies Mediate VITT

Background: ChAdOx1 nCoV-19 (AstraZeneca) and Ad26.COV2.S (Janssen Johnson & Johnson) vaccines against COVID-19 have been associated with thrombotic thrombocytopenic reactions referred to as vaccine-induced immune thrombotic thrombocytopenia (VITT) characterized by the presence of platelet-activating, anti-PF4 antibodies. While VITT shares key clinical features with a similar but separate entity, Heparin-induced thrombocytopenia (HIT), there appear to be important differences: 1) VITT patients have extremely high thrombosis rates and are very strongly positive in PF4-polyanion ELISAs, and 2) Many patients with VITT frequently present with refractoriness to therapy or have disease recurrence that suggests distinct antibody characteristics due to a strong autoimmune anti-PF4 response.Aims: The goal of this study was to characterize anti-PF4 antibodies in VITT.Methods: Five VITT patients were studied, one after ChAdOx1 nCoV-19 vaccination and four after Ad26.COV2. Reactivity of VITT anti-PF4 antibodies to uncomplexed PF4, PF4-Polyvinyl sulfonate (PVS), and PF4-heparin targets was evaluated, and the platelet-activating ability of these antibodies was examined in the PF4-dependent P-selectin Expression assay (PEA). Anti-PF4 antibodies were isolated from patient blood samples using PF4-treated heparin sepharose beads, and isolated antibodies were subject to mass spectrometric evaluation (Liquid Chromatography Electrospray Ionization Quadrupole time-of-flight mass spectrometry [LC-ESI-QTOF MS]).Results: Antibodies from all VITT patients recognized both uncomplexed and complexed PF4 (Fig. 1A). Interestingly, recognition of PF4 by VITT antibodies was lower if PF4 targets were complexed with polyanions, PVS, or heparin (Fig. 1A). These results contrasted with those obtained in a “classical” HIT patient which showed reactivity to PF4/polyanion complexes, but not to uncomplexed PF4 (Fig 1A). All samples activated platelets in the PEA (data not shown). Mass spectrometric evaluation of anti-PF4 antibodies isolated from VITT patients demonstrated monoclonal anti-PF4 antibodies in three patients, and bi- and tri-clonal antibodies in one patient each (a representative monoclonal antibody anti-PF4 antibody is shown in Fig 1B). Consistent with current dogma, polyclonal anti-PF4/polyanion antibodies were seen in “classical” HIT (Fig 1C). Evaluation of anti-PF4 antibodies in spontaneous HIT, a type of autoimmune HIT seen in pro-inflammatory milieus such as orthopedic surgery and infectious prodromes also demonstrated monoclonal anti-PF4 antibodies (Fig 1D). Eluates from control heparin-sepharose beads did not reveal any immunoglobulins (data not shown).Conclusion: Although development of platelet-activating anti-PF4 antibodies and the thrombotic thrombocytopenia syndrome seen after ChAdOx1 nCoV-19 and Ad26.COV2.S vaccination resembles HIT, these findings demonstrate that clonally restricted anti-PF4 antibodies mediate VITT while polyclonal anti-PF4 antibodies mediate HIT. In addition, we noted clonally-restricted anti-PF4 antibodies in another condition that does not require proximate heparin exposure, spontaneous (“autoimmune”) HIT. In VITT, the strong immune response after vaccine administration may result in the activation of a single or few pre-existing anti-PF4 reactive clones, and development of clonally restricted anti-PF4 antibodies with a similar pathophysiology to Spontaneous HIT. It is also likely that high levels of monoclonal/oligoclonal anti-PF4 antibodies cause the severe thrombotic phenotypes seen in VITT and Spontaneous HIT. The high mortality rate and reports of disease refractoriness to therapy in VITT may warrant consideration of additional therapeutic modalities like rituximab and therapeutic plasma exchange in select cases.Figure Legends: (A): VITT (Patient 1-ChAdOx1 nCoV-19; Patients 2-5, Ad26.COV2.S) patient samples were tested in ELISA against uncomplexed PF4 (white), and PF4 in complex with polyvinyl sulfonate (light grey), or unfractionated heparin (dark gray). (B-D) Mass spectrometric evaluation of anti-PF4 antibodies isolated from VITT (B), HIT (C) and spontaneous HIT patient sera (D). “Relative Intensity” refers to abundance of the Ig light chain relative to the polyclonal background. Numbers above Ig light chain peaks depict mass/charge ratios. NC- Normal control. [Display omitted] DisclosuresMurray: Mayo Clinic: Other: Has received patents for the Mass-Fix technology which has been licensed to the Binding Site with potential royalties.. Padmanabhan: Veralox Therapeutics: Membership on an entity's Board of Directors or advisory committees.

Recognizing Vaccine-Induced Immune Thrombotic Thrombocytopenia.

OBJECTIVES:Vaccine-induced immune thrombotic thrombocytopenia is an unexpected consequence of the coronavirus disease 2019 pandemic era. We reviewed the pathogenesis, clinical presentation, diagnosis, and treatment of this rare side effect.DATA SOURCES:Online search of published medical literature through PubMed, Scopus, Web of Science, and Google Scholar using the terms “COVID-19,” “vaccine,” “thrombosis” was performed.STUDY SELECTION:Articles were chosen for inclusion based on their relevance to coronavirus disease 2019, vaccine, and thrombosis.DATA SYNTHESIS:Vaccine-induced immune thrombotic thrombocytopenia manifests most often as unusual thromboses (cerebral venous sinus thrombosis, splanchnic vein thrombosis) but sometimes also “usual” thromboses (arterial stroke, pulmonary embolism, deep-vein thrombosis), with oftentimes severe thrombocytopenia, that becomes clinically evident 5–30 days after adenovirus-vectored coronavirus disease 2019 vaccine administration. Most patients have disseminated intravascular coagulation. These features are the result of vaccine-triggered formation of anti-platelet factor 4 immunoglobulin G that activate platelets, clinically mimicking autoimmune heparin-induced thrombocytopenia. Early recognition based on thrombosis (sometimes, hemorrhage), thrombocytopenia, and d-dimer elevation within the day 5–30 postvaccine “window” is important given treatment with high-dose IV immunoglobulin plus nonheparin anticoagulation.CONCLUSIONS:Vaccine-induced immune thrombotic thrombocytopenia is a serious complication of vaccination that is not feasible to anticipate or prevent. When the patient presents with sustained headache, neurologic symptoms/signs, abdominal pain, dyspnea, or limb pain/swelling beginning 5–30 days post vaccination, platelet count and d-dimer must be measured, and imaging for thrombosis performed. Confirmation of vaccine-induced immune thrombotic thrombocytopenia diagnosis should be ordered (platelet factor 4/polyanion enzyme-linked immunosorbent assay; platelet factor 4–enhanced platelet activation testing) as treatment is initiated (nonheparin anticoagulation, IV immunoglobulin).

Vaccine‐induced Immune Thrombocytopenia and Thrombosis (VITT)

Vaccine‐induced Immune Thrombocytopenia and Thrombosis (VITT)

Malignant middle cerebral artery syndrome with thrombotic thrombocytopenia following vaccination against SARS-CoV-2.

Cases of thromboses at unusual sites with thrombocytopenia have been reported following vaccination against Sars-CoV-2. This new syndrome, christened vaccine-induced thrombotic thrombocytopenia (VITT), mainly results in venous thromboses. We report the case of a young woman with a right middle cerebral artery stroke following vaccination with ChAdOx1 nCoV-19. A diagnosis of VITT was made and platelet counts began to recover shortly after commencing treatment with argatroban, intravenous immunoglobulins and corticosteroids. On day 6 following admission, the patient deteriorated neurologically and decision made to proceed with decompressive hemicraniectomy. There were no perioperative complications and anticoagulation with argatroban was reinitiated on the first postoperative day. VITT is a rare condition resembling auto-immune heparin-induced thrombocytopenia. All critical care staff should be aware of the rare link between vaccination against SARS-CoV-2 and VITT and the need to rapidly commence both anticoagulation, using heparin alternatives, and immunomodulation.

The first known case of vaccine‐induced thrombotic thrombocytopenia in Australia

The first known case of vaccine‐induced thrombotic thrombocytopenia in Australia

HIT in the head: a systematic review of cerebral venous sinus thrombosis in classical and autoimmune heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) causes thrombosis and thrombocytopenia, usually due to prior heparin exposure, so-called classical HIT. However, in the autoimmune form, the signs and symptoms of HIT occur without prior heparin exposure. Development of cerebral venous sinus thrombosis (CVST) secondary to HIT is a rare occurrence, with relatively few reports in the literature. There is a need to better understand the clinical presentation and treatment paradigms in these rare cases. Therefore, we present the first systematic review of CVST occurring in classical and autoimmune HIT. Cases of HIT-induced CVST were identified through a systematic search of Pubmed from the date of inception to March 2021. Literature search revealed 21 cases of HIT and associated CVST with six cases (28.6%) of autoimmune HIT. Patients presented with signs and symptoms consistent with increased intracranial pressure, intracerebral hemorrhage (ICH), and/or focal neurologic deficits. Headache was the most common symptom with 12 patients (60.0%) presenting as such. 10 patients (47.6%) included in the study developed ICH. Non-heparin anticoagulants, especially direct thrombin inhibitors, were the first-line treatment for the majority of patients (55.6%). Intravenous immunoglobulin (IVIG) was used as treatment for select patients (16.7%) with autoimmune HIT. Few patients received surgical intervention for CVST (14.3%) or ICH (30.0%). Four patients had a full recovery, four patients had residual deficits, and seven patients ultimately expired. Symptoms of HIT-induced CVST are often related to CNS dysfunction. Non-heparin anticoagulants are important to treat CVST, even when patients have concomitant ICH, and may be supplemented with IVIG if treating autoimmune HIT. Rapid identification and treatment of HIT-induced CVST is imperative in order to prevent morbidity and mortality.

US Case Reports of Cerebral Venous Sinus Thrombosis With Thrombocytopenia After Ad26.COV2.S Vaccination, March 2 to April 21, 2021

Cerebral venous sinus thrombosis (CVST) with thrombocytopenia, a rare and serious condition, has been described in Europe following receipt of the ChAdOx1 nCoV-19 vaccine (Oxford/AstraZeneca), which uses a chimpanzee adenoviral vector. A mechanism similar to autoimmune heparin-induced thrombocytopenia (HIT) has been proposed. In the US, the Ad26.COV2.S COVID-19 vaccine (Janssen/Johnson & Johnson), which uses a human adenoviral vector, received Emergency Use Authorization (EUA) on February 27, 2021. By April 12, 2021, approximately 7 million Ad26.COV2.S vaccine doses had been given in the US, and 6 cases of CVST with thrombocytopenia had been identified among the recipients, resulting in a temporary national pause in vaccination with this product on April 13, 2021. To describe reports of CVST with thrombocytopenia following Ad26.COV2.S vaccine receipt. Case series of 12 US patients with CVST and thrombocytopenia following use of Ad26.COV2.S vaccine under EUA reported to the Vaccine Adverse Event Reporting System (VAERS) from March 2 to April 21, 2021 (with follow-up reported through April 21, 2021). Receipt of Ad26.COV2.S vaccine. Clinical course, imaging, laboratory tests, and outcomes after CVST diagnosis obtained from VAERS reports, medical record review, and discussion with clinicians. Patients' ages ranged from 18 to younger than 60 years; all were White women, reported from 11 states. Seven patients had at least 1 CVST risk factor, including obesity (n = 6), hypothyroidism (n = 1), and oral contraceptive use (n = 1); none had documented prior heparin exposure. Time from Ad26.COV2.S vaccination to symptom onset ranged from 6 to 15 days. Eleven patients initially presented with headache; 1 patient initially presented with back pain and later developed headache. Of the 12 patients with CVST, 7 also had intracerebral hemorrhage; 8 had non-CVST thromboses. After diagnosis of CVST, 6 patients initially received heparin treatment. Platelet nadir ranged from 9 ×103/µL to 127 ×103/µL. All 11 patients tested for the heparin-platelet factor 4 HIT antibody by enzyme-linked immunosorbent assay (ELISA) screening had positive results. All patients were hospitalized (10 in an intensive care unit [ICU]). As of April 21, 2021, outcomes were death (n = 3), continued ICU care (n = 3), continued non-ICU hospitalization (n = 2), and discharged home (n = 4). The initial 12 US cases of CVST with thrombocytopenia after Ad26.COV2.S vaccination represent serious events. This case series may inform clinical guidance as Ad26.COV2.S vaccination resumes in the US as well as investigations into the potential relationship between Ad26.COV2.S vaccine and CVST with thrombocytopenia.