Naturally processed peptides spanning the HPA-1a polymorphism are efficiently generated and displayed from platelet glycoprotein by HLA-DRB3*0101–positive antigen-presenting cells

Abstract

IntroductionHuman platelet antigen (HPA)-1a and HPA-1b alloantigens are determined, respectively, by the polymorphism Leu or Pro at position 33 of the ␤ 3 chain (glycoprotein IIIa [GPIIIa]) of the platelet integrin ␣ IIb ␤ 3 . 1 Mismatch between fetal and maternal platelet antigens can lead to the production of maternal immunoglobulin G anti-HPA-1a alloantibodies that cross the placenta and cause thrombocytopenia in an HPA-1a-positive fetus. 2,3 Approximately 2% of white women are HPA-1b homozygous, and 10% of these develop anti-HPA-1a alloantibodies during pregnancy. 2,3 The estimated incidence of severe neonatal alloimmune thrombocytopenia (NAIT) is approximately 1 in 1100 neonates, and the clinical manifestations range from mild purpura to intracranial hemorrhage. 2,3 Maternal responsiveness to HPA-1a shows strong associations with major histocompatibility complex class II, with greater than 95% of responders carrying the DRB3*0101 allele encoding human histocompatibility leukocyte antigen (HLA)-DR52a, 4,5 and up to 94% may carry the DQB1*02 allele. 6 Classical immunoglobulin G antibody responses are dependent on T-helper (Th) cells that recognize antigen as short, processed, linear peptides bound to major histocompatibility complex molecules on antigen-presenting cells (APCs). The identification of the immunodominant helper epitopes on the integrin sequence containing the HPA-1a polymorphic site would be an important step toward understanding and controlling the immune response to HPA-1a. Earlier studies have shown that short, synthetic HPA-1a peptides containing the Leu 33 polymorphism bind to recombinant DR52a molecules, whereas the Pro 33 version does not, with Trp 25 and Leu 33 predicted to be important anchor residues. 7 The dominant synthetic HPA-1a peptides recognized by peripheral blood Th cells from women alloimmunized with anti-HPA-1a contain the Trp 25 -Leu 33 core sequence, with Leu 33 preferentially at the C-terminus. 8 Recent characterization of the fine specificity of Th cell clones derived from women with NAIT also mapped a putative core epitope spanning Trp 25 -Leu 33 . 9 However, antigen processing within APCs is highly selective, with very few peptides typically generated from any given protein for efficient display to Th cells, 10-13 and there is no direct evidence that this core sequence is indeed presented from platelet glycoprotein. The aim of this study was therefore to determine whether DRB3*0101 homozygous APCs, pulsed with HPA-1a, naturally process and present peptides spanning the predicted Trp 25 -Leu 33 core epitope. Methods Antigen preparationDNA encoding HPA-1a amino acids 1-62 was cloned into the expression vector pGEX-6P-1, and Escherichia coli BL21(DE3) cells were transformed. This sequence corresponds to the complete plexin-semaphorinintegrin (PSI) domain of platelet GPIIIa, which spans the Leu 33 HPA-1a polymorphism and expresses the HPA-1a antigen. 14 Because the PSI domain is predicted to be on the exterior of the GPIIIa protein 14 and accessible to processing enzymes, ...

Modeling and Molecular Dynamics of HPA-1a and -1b Polymorphisms: Effects on the Structure of the β3 Subunit of the αIIbβ3 Integrin

“…Such structural differences may modify β3 antigenicity and possibly induce differences in the immune response to the HPA-1a and HPA-1b alleles. As far as we know, the high specific HPA-1a immunogenicity is attributed to the ability of the processed L33 β3 peptides to bind, with high affinity, the HLA-DRB3*0101 allele carried by the antigen-presenting cells [40]–[42]. However, although our study shows clear structural differences between the two β3 variants that may affect the antigenicity, we cannot conclude as to immunogenicity.…”

Current perspectives on fetal and neonatal alloimmune thrombocytopenia – increasing clinical concerns and new treatment opportunities

“…This strong association suggests that CD4 T-cell activation by the DRA/DRB3*01:01 molecule is a determining event in the immune response against HPA-1a. In support of this notion, it has been shown that integrin β3-derived peptides with Leu33 (HPA-1a peptide), but not with Pro33, bind well to the DRA/DRB3*01:01 molecule and thus can be presented by antigen-presenting cells 30,31. Also, HPA-1a-specific DRA/DRB3*01:01-restricted CD4 T cells have been isolated from women who have had a child affected by FNAIT 32,33.…”

Molecular insight into human platelet antigens: structural and evolutionary conservation analyses offer new perspective to immunogenic disorders

“…The second frequent antigen causing NAIT is HPA‐5b (10%‐15% of NAIT cases), while all the rest cause NAIT very rarely 11 . Only 10% of HPA‐1b–homozygous women with an HPA‐1a–positive fetus develop anti‐HPA‐1a alloantibodies during pregnancy 12 . Maternal responsiveness to HPA‐1a shows strong association with DRB*0101 allele encoding human histocompatibility leukocyte antigen DR52a.…”