B LOOD G ROUP R EVIEW CD59: A long-known complement inhibitor has advanced to a blood group system C. Weinstock, M. Anliker, and I. von Zabern The blood group system number 35 is based on CD59, a 20-kDa by Zalman et al.1 from the group of H.J. Muller-Eberhard in La membrane glycoprotein present on a large number of different Jolla, California, and in 1988 by Schönermark et al. from the cells, including erythrocytes. The major function of CD59 is to group of G.M. Hänsch in Heidelberg (Germany).2 This inhibitor protect cells from complement attack. CD59 binds to complement components C8 and C9 and prevents the polymerization of C9, was published under the designations “HRF (homologous which is required for the formation of the membrane attack restriction factor)” and “C8bp (C8 binding protein),” complex (MAC). Other functions of CD59 in cellular immunity respectively, to describe its properties.1,2 “C8bp” indicates the are less well defined. CD59 is inserted into the membrane by a glycosylphosphatidylinositol (GPI) anchor. A defect of this anchor binding capacity for C8. The name “HRF” points to a species causes lack of this protein from the cell membrane, which leads to incompatibility of this “factor” that provides protection from an enhanced sensitivity towards complement attack. Patients with complement attack more effectively in a homologous (e.g., paroxysmal nocturnal hemoglobinuria (PNH) harbor a varying human erythrocytes as a target of human complement) than in percentage of red blood cell clones with a defect in GPI-anchored proteins, including CD59. The most characteristic symptoms of a heterologous system (e.g., human erythrocytes as a target of this disease are episodes of hemolysis and thromboses. Although guinea pig complement). In 1987, both groups independently CD59 has been classified as a membrane protein for more than described a deficiency of this factor on red blood cells (RBCs) of 25 years, an alloantibody directed against CD59 was found patients with paroxysmal nocturnal hemoglobinuria (PNH).3,4 only recently. So far, the first and sole alloantibody described was detected in a CD59-deficient child. In 2014, CD59 received A similar protein was reported in 1990 by Watts et al. from the status of a blood group system by the International Society B.P. Morgan’s group in Cardiff (UK), which they called “MIP for Blood Transfusion Red Cell Immunogenetics and Blood (membrane attack complex inhibiting protein).”5 All three of Group Terminology Working Party. Among a variety of almost 20 synonyms, the designation CD59 was chosen for the blood these groups ascribed to this inhibitor a molecular weight of group system and CD59.1 for the wild-type protein. The only approximately 65 kDa, which is significantly higher than that three alleles published to date are null alleles. All CD59-deficient known for CD59 today. This contradiction has never been individuals recognized so far were severely ill, two of whom have resolved; impurities or the formation of protein aggregates are died. Most of the reported cases present with a typical clinical picture within the first year of life that includes neuropathy, possible explanations. strokes, and mild Coombs-negative hemolysis. In one published The first description of an inhibitor with functional case, the application of the complement inhibitor eculizumab characteristics identical to HRF, C8bp, and MIP, but with a caused a pronounced improvement of the clinical situation. molecular weight of approximately 20 kDa, appeared in 1988 Immunohematology 2015;31:145–151. under the designation “P18.”6 Within 1 year, this protein was Key Words: CD59, HRF20, MIRL, complement regulatory rediscovered by several other groups,7–10 who each named protein, blood group, hemolysis it differently: “MIRL,” for which absence from PNH RBCs was confirmed, “HRF20,” “H19,” and “MEM43.” In 1990, History and Nomenclature Lachmann’s group in Cambridge added another name, “protectin,” to indicate its function in the protection of cells In 2014, the 35th blood group system was allocated from complement attack;11 the CD number 59 was assigned by to CD59, a small membrane glycoprotein of approximately the International Leucocyte Workshop. 20 kDa molecular weight, which is inserted into the Within that same year, the first case of a congenital membrane of all circulating cells and most tissues by a homozygous CD59 deficiency was detected in Japan.12,13 It glycosylphosphatidylinositol (GPI) anchor. An important was not until 2013 that several further publications on CD59- function of this protein is to protect cells from damage by deficient children of North African Jewish or Turkish origin unwanted complement attack. appeared.14–17 Among these cases was a child with a history An inhibitor of the terminal stages of complement with the of transfusions who had formed an antibody directed against functional characteristics of CD59 was first described in 1986 CD59. The identification of this alloantibody allowed CD59 IMMUNOHEMATOLOGY, Volume 31, Number 4, 2015 145 C. Weinstock et al. to be defined as a blood group antigen.16 Finally, in 2014, (Table 3). These introduce premature stop codons, which likely CD59 was accepted by the International Society for Blood prevent the synthesis of functionally active protein. Transfusion (ISBT) Red Cell Immunogenetics and Blood All identified CD59-deficient children were found Group Terminology Working Party as the 35th blood group to carry homozygous alleles, and most were offspring of system (Table 1).18 Although there was a choice among almost consanguineous parents. The expression of CD59 on RBCs of 20 different pre-existing synonyms, the CD classification was heterozygous parents was reported to be lower than normal.14,16 adopted for the designation not only of the antigen but also of the Although the heterozygous patients were apparently without blood group system. The rationale for the uncommon choice of clinical signs, all children with homozygous congenital total a CD number was merely practical: It should be easy to identify deficiency of CD59 suffered from severe disease.12–17 the protein molecule underlying the new blood group system. Furthermore, this decision was made in agreement with the Molecular Basis and Biochemistry Nomenclature Committee of the Complement Society, which suggested the use of the designation CD59 for this complement The CD59 gene (HGNC: 1689; Entrez Gene: 966) was inhibitor. mapped to chromosome 11p13, where it spans over 33 kbp.19,20 Alternative splicing of four to seven exons leads Table 1. Blood group system CD59 to multiple transcript variants; eight of them have been Antigen number deposited as reference sequences in the National Center for System 001 Biotechnology Information database. They all differ only in the 035 CD59 CD59.1 5´ untranslated region but encode the same 128–amino acid Captured from the Table of Blood Group Antigens, version 4.18 preproprotein. The amino-terminal 25–amino acid residues function as signal peptides and are removed during processing The only three alleles published at present are null alleles of the polypeptide. The carboxyl-terminal 26 residues are (Table 2). removed during the attachment of the GPI anchor, leaving a mature CD59 protein, which consists of 77 amino acids.21 Table 2. Alleles encoding the CD59 phenotype null The three-dimensional structure of CD59 has DNA Protein Year of first description been investigated using nuclear magnetic resonance c.123delC p.Val42Serfs*38 199012,13 spectroscopy21,22 and X ray crystallography.23 The core of c.361delG p.Ala121Glnfs c.266G>A p.Cys89Tyr 201214 c.146delA p.Asp49Valfs*31 201315 Genetics and Inheritance The CD59-deficient individuals encountered to date were of Japanese, North African Jewish, or Turkish origin (Table 3). The amino acid substitution Cys89Tyr is a founder mutation in North African Jews that occurs with a carrier rate of 1 in 66.14 The underlying amino acid substitution involves the loss of a cysteine residue. In the normal protein, Cys89 contributes to a disulfide bond. Lack of this bond may destabilize the tertiary structure of the protein and cause abnormal processing. The authors conclude from their experiments that this missense mutation does not prevent biosynthesis of a CD59-like Fig. 1 Human CD59 consists of (a) a central three-stranded β-sheet molecule. Rather, a failure of the transport to the membrane and (b) an α-helix on one side of the sheet. On the other side is (c) an anti-parallel β-sheet finger carrying the (d) N-glycan. CD59 is or of membrane integration seems to cause the lack of CD59 anchored to the cell membrane via (e) glycosylphosphatidylinositol. expression.14 The presently known CD59-deficient individuals The star indicates the putative binding site for C8/C9 and intermedilysin. of Japanese and Turkish origin carry frameshift mutations 146 IMMUNOHEMATOLOGY, Volume 31, Number 4, 2015 CD59 blood group system Table 3. Clinical presentation of patients with CD59 deficiency Number of Parents Age of disease Protein Ethnic origin patients Sex consanguineous? onset Clinical presentation References p.Val42Serfs*38, Japanese 1 Male Yes 13 years Symptoms from age 13 to 22 years Yamashina et al.,12 p.Ala121Glnfs • 9 Episodes of PNH-like hemolysis Motoyama et al.13 • No RBC transfusions • 2 Cerebral infarctions • No peripheral nervous system involvement reported p.Cys89Tyr North 5 3 Males, 1 Patient 3–7 months • Chronic and paroxysmal Coombs-negative Nevo et al.14 African 2 females hemolysis Jewish • Occasional RBC transfusions