Fetal/neonatal alloimmune thrombocytopenia due to anti‐CD36 antibodies: antibody evaluations by CD36‐transfected cell lines

Abstract

BACKGROUND: Isoantibodies against CD36 (platelet glycoprotein 4), developed in Type I CD36-deficient mothers are frequently reported as the cause of fetal/ neonatal alloimmune thrombocytopenia in the Asian population. Therefore, further detailed characterization of anti-CD36-mediated fetal/neonatal alloimmune thrombocytopenia is warranted. Here, we report the characterization of a patient with fetal/neonatal alloimmune thrombocytopenia in a Taiwanese family caused by anti-CD36 isoantibodies using a novel antigen-capture method. STUDY DESIGN AND METHODS:Platelets and monocytes were analyzed for CD36 expression by flow cytometry. Sequencing analysis of the CD36 gene was performed to identify the mutation underlying the CD36 deficiency. Stable transfected human embryonic kidney HEK293 cells expressing recombinant CD36 were established. These cells were used for the characterization of anti-CD36 isoantibodies by flow cytometry, immunoprecipitation, and antigen-capture assay. RESULTS:Flow cytometry analysis revealed a total absence of CD36 on both platelets and monocytes of the mother (Type I CD36-deficient) caused by heterozygous deletions of the CD36 gene (332_333delCA and c.1254 1 6_1254 1 11delTATTTG). Analysis of maternal serum with CD36-transfected HEK293 cells by flow cytometry, immunoprecipitation, and antigen-capture assay demonstrated the presence of anti-CD36 isoantibodies in maternal serum. Interestingly, this antibody could not be detected by the monoclonal antibody immobilization of platelet antigens assay when anti-CD36 monoclonal antibody (clone FA6-152) was used as the capture antibody.CONCLUSION: This case reemphasizes the role of anti-CD36 isoantibodies on the pathomechanism of fetal/ neonatal alloimmune thrombocytopenia. The fact that the monoclonal antibody immobilization of platelet antigens assay does not seem to be reliable for the identification of all anti-CD36 antibodies indicates that screening of anti-CD36 isoantibodies by a monoclonal antibody-independent method, as presented here, should be considered. Fetal/neonatal alloimmune thrombocytopenia (FNAIT), which occurs in from 1 of 800 to 1 of 1000 live births, is the most common cause of severe thrombocytopenia and intracranial hemorrhage in the fetus and in term newborns.1,2 FNAIT is caused by maternal antibodies, which recognize paternalderived alloantigens on fetal platelets, leading to platelet destruction. In Caucasians, more than 75% of FNAIT cases are induced by alloantibodies against human platelet antigen-1a (HPA-1a), 3,4 whereas the most common antibodies causing FNAIT in Japanese are anti-HPA-4b alloantibodies. The clinical importance of anti-CD36 isoantibodies in the development of FNAIT has been reported in different populations.9-13 Currently, the antigen-capture assay represents the gold standard for detecting platelet antibodies. 14 With this assay, platelet antigens captured by mouse monoclonal antibody are used as the target for platelet antibodies. Unfortunately, this approach does not seem to be reliable for the identi...

Frequency of CD36 deficiency in Thais analyzed by quantification of CD36 on cell surfaces and in plasma

“…In this study, we found that around 0.43% of Thai blood donors exhibit Type I CD36 deficiency and are therefore at risk of developing anti‐CD36 antibodies if exposed to the antigen through transfusion or pregnancy . These antibodies can cause PTR and serious FNAIT, as recently reported in different Asian populations . Indeed, a case of FNAIT has been reported in Thailand .…”

“…2 These antibodies can cause PTR and serious FNAIT, as recently reported in different Asian populations. 3,4 Indeed, a case of FNAIT has been reported in Thailand. 18 Based on this fact, routine diagnostics as well as treatment strategy for anti-CD36-mediated bleeding disorders with use of CD36-compatible platelets should be seriously taken into consideration.…”

“…Individuals with CD36 Type I deficiency can develop anti-CD36 antibodies during pregnancy or upon transfusion, a phenomenon that is responsible for a number of immune-mediated platelet disorders such as fetal/neonatal alloimmune thrombocytopenia (FNAIT), platelet transfusion refractoriness (PTR), and posttransfusion purpura. [2][3][4] In addition, anti-CD36s have been found in cases of transfusionrelated acute lung injury. 5 However, the exact mechanism by which anti-CD36s mediate these disorders is currently not fully understood.…”

“…Two types of CD36 deficiency are known; Type I–deficient individuals do not express CD36 on either monocytes or platelets, while those with Type II deficiency express the molecule on monocytes only. Individuals with CD36 Type I deficiency can develop anti‐CD36 antibodies during pregnancy or upon transfusion, a phenomenon that is responsible for a number of immune‐mediated platelet disorders such as fetal/neonatal alloimmune thrombocytopenia (FNAIT), platelet transfusion refractoriness (PTR), and posttransfusion purpura . In addition, anti‐CD36s have been found in cases of transfusion‐related acute lung injury .…”

New molecular basis associated with CD36‐negative phenotype in the sub‐Saharan African population

“…10 Besides, the lack of such a membrane GP can also have a negative impact, because it can lead to isoimmunization in Type I deficiency, in the contexts of pregnancy, PLT transfusion, and even RBC transfusion, as RBCs are contaminated by remaining PLTs and WBCs even after centrifugation and filtration and potentially by circulating CD36-positive microparticles, a source of the plasmaderived form of soluble CD36. 20,21 Anti-CD36 isoimmunization can engender serious complications such as PLT transfusion refractoriness, 13 PTP-like syndrome, 14 and fetal or neonatal immune thrombocytopenia, 15,[22][23][24] which explains the interest of our lab in searching for CD36neg patients and their risk of immunization as well as to study the underlying molecular defects.…”