P248 Next generation sequencing reveals a novel hla-a*24 allele associated with hla-dqb1*06:10 in asians

Abstract

Aim Next Generation Sequencing (NGS) for HLA typing improves not only the resolution of typing results but also aids in the discovery of novel alleles. Here we present a novel HLA-A∗24 allele associated with HLA-DQB1∗06:10, which was found in an Asian family and an unrelated donor. Methods Genomic DNA samples isolated from a patient, family members and potential unrelated donors were tested by reverse sequence specific oligonucleotide probes (rSSOP), sequence based typing (SBT) and NGS technology. In NGS typing, genomic DNA was amplified using the Illumina TruSight HLA v2 Sequencing Panel kit. After normalization, fragmentation/tagmentation and dual-indexing procedures, the amplicons were pooled onto the MiSeq sequencer to facilitate cluster generation and sequencing by synthesis on the flow cell. The generated data was then analyzed with Illumina TruSight HLA Assign software. Results A patient being evaluated for a potential hematopoietic stem cell transplant (HSCT) was typed by Sanger sequencing and found to be homozygous at HLA-A, B, C and DRB1. By Luminex LabType, the patient was heterozygous at DQA1, DQB1 and DPB1. However, subsequent NGS typing revealed heterozygousity for HLA-A, with an expected A∗24:02:01 and a novel A∗24 with a mutation in exon 6 which resulted in an amino acid change from lysine to glutamic acid. The same mutation was also found in one of his children and one of four unrelated donors. The patient, child and unrelated donor who had the HLA-A∗24:NEW allele all self-identified as Asian. In all three cases, the HLA-A∗24:NEW allele was associated with HLA-DQB1∗06:10; this association is found only in Asians at low frequency. Conclusions This HLA-A∗24:NEW allele may be associated with DQB1∗06:10 in Asians. Implementing NGS technology in clinical practice will not only allow us to choose the best donor from many potentially matched donors but also will fill in gaps in HLA databases to improve our typing.