IMMUNOHEMATOLOGY HLA-DRB1*07:01 allele is primarily associated with the Diego a alloimmunization in a Brazilian population Wilson Baleotti Jr, 1 Marcelo Ortega Ruiz, 2 Antonio Fabron Jr, 2 Lilian Castilho, 3 Silvana Giuliatti, 4 and Eduardo Antonio Donadi 5 BACKGROUND: The Diego blood group presents a major polymorphic site at Residue 854, causing a proline (Di b antigen) to leucine (Di a antigen) substitution. Di a alloimmunization has been observed among Asian and Native South American populations. Considering that Brazilians represent a genetically diverse popula- tion, and considering that we have observed a high inci- dence of Di a alloimmunization, we typed HLA-DRB1 alleles in these patients and performed in silico studies to investigate the possible associated mechanisms. STUDY DESIGN AND METHODS: We studied 212 alloimmunized patients, of whom 24 presented immuno- globulin G anti-Di a , 15 received Di(a+) red blood cells and were not immunized, and 1008 were healthy donors. HLA typing was performed using commercial kits. In silico analyses were performed using the TEPITOPEpan software to identify Diego-derived anchor peptide binding to HLA-DRB1 molecules. Residue alignment was performed using the IMGT/HLA for amino acid identity and homology analyses. RESULTS: HLA-DRB1*07:01 allele was overrepre- sented in Di a -alloimmunized patients compared to nonimmunized patients and to healthy donors. Two motifs were predicted to be potential epitopes for Di a alloimmunization, the WVVKSTLAS motif was predicted to bind several HLA-DR molecules, and the FVLILTVPL motif exhibited highest affinity for the HLA-DRB1*07:01 molecule. Pocket 4 of the DRB1*07:01 molecule con- tained specific residues not found in other HLA-DRB1 molecules, particularly those at Positions 13(Y), 74(Q), and 78(V). CONCLUSION: Individuals carrying the HLA- DRB1*07:01 allele present an increased risk for Di a alloimmunization. The identification of susceptible indi- viduals and the knowledge of potential sensitization peptides are relevant approaches for transfusion care, diagnostic purposes, and desensitization therapies. T he solute carrier family 4, anion exchanger, member 1 (SLC4A1, AE1) gene presents 20 exons and codes the red blood cell (RBC) membrane protein band 3, which is one of the major pro- teins of the RBC membrane, exhibiting 911 amino acids and 1 million copies per membrane. The 40-kDa N-terminal domain binds to glycolytic enzymes and hemoglobin, whereas the 55-kDa glycosylated transmem- brane C-terminal domain is primarily responsible for anion transport. 1-4 The Diego blood group system com- prises 22 potentially immunogenic antigens of Band 3 variants: 1) two pairs of antithetical Di a /Di b and Wr a /Wr b antigens, 2) 17 antigens presenting low worldwide fre- quency, and 3) one antigen exhibiting high worldwide frequency. 5-11 The Di a antigen is considered to be the major antigen of the Diego blood group system due to its impor- tance in hemolytic transfusion reactions and hemolytic disease of the fetus and newborn. Serologic and molecular studies have shown that the Di a antigen is rare among European Caucasians and African Brazilians; 12,13 however, the antigen is commonly observed among Native South American (up to 5%-65%) and Asian populations (5%- 12%), and the Di a antigen is considered to be an important ABBREVIATION: EF = etiologic fraction. From the 1 Faculty of Medicine of Marília (FAMEMA), Marília, São Paulo, Brazil; 2 Laboratório Diagnósticos do Brasil (DB), Curitiba, Paraná, Brazil; 3 Hemocentro da Universidade Estadual de Campinas (UNICAMP), Campinas, São Paulo, Brazil; 4 Department of Genetics, Faculty of Medicine of Ribeirão Preto, University of São Paulo (USP), Ribeirão Preto, São Paulo, Brazil; 5 Division of Clinical Immunology, Department of Medicine, Faculty of Medicine of Ribeirão Preto, University of São Paulo (USP), Ribeirão Preto, São Paulo, Brazil. Address reprint requests to: Wilson Baleotti Jr, Hemocentro da Faculdade de Medicina de Marília, Rua Lourival Freire, n°240, CEP 17.519-050–Marília, São Paulo, Brasil; e-mail: baleotti@famema.br. Received for publication October 15, 2013; revision received February 12, 2014, and accepted February 14, 2014. doi: 10.1111/trf.12652 © 2014 AABB TRANSFUSION 2014;54:2468-2476. 2468 TRANSFUSION Volume 54, October 2014