P074 : IDENTIFICATION OF NOVEL HLA ALLELES IN HEMATOPOIETIC STEM CELL TRANSPLANT PATIENTS

Abstract

Identification of novel HLA alleles is a common finding in the HLA lab. This finding is particularly challenging in hematopoietic cell transplant (HCT) patients since may prevent them from finding HLA matched donors. In addition, somatic mutations can occur in blast cells of patients with hematological malignancies and are attributed to a mechanism of immune evasion. Herein, we describe the approach used at the University of Chicago Transplant Immunology Laboratory for patients with hematological malignancies who were evaluated for HCT work-up. DNA from recipients who carried novel HLA alleles detected in peripheral blood (PBL) were confirmed in recipient’s germline DNA obtained from buccal cells. DNA was isolated from peripheral blood cells and buccal cells of 5 patients by automated and manual methods. The novel HLA-A, B, C, and DQB1 genes were amplified by polymerase chain reaction (PCR) using locus specific primers. Exons 2 through 4 for HLA-A, B, C, and exons 2 and 3 for HLA-DQB1 were sequenced by Atria sequencing kits, and analyzed by Assign 3.6 SBT Software (Abbot Molecular, Des Plaines, IL). Novel alleles were confirmed in buccal cells of patients 1, 2, 3 and 4. Patient 5 only carried the new allele in the blast cells. Patient 1 had a matched related donor (MRD) that carried the same polymorphism at HLA-DQB1 locus. Patients 2 and 3 did not have MRD, and due to presence of novel allele in the HLA-B locus, an 8/8 matched unrelated donor (MUD) could not be found. For patient 4, the polymorphism was located outside of the hypervariable region and was classified as a group of alleles which have the same antigen binding domains (P group). Patient 4 was most likely to find an 8/8 MUD due to this classification. For patient 5 the new allele was only present on DNA from somatic cells, did not affect donor searches, and multiple MUD were available through the National Marrow Donor Registry (NMDP). Identification of novel alleles is a common finding that can affect HCT donor searches. Confirmation of genomic versus somatic nature of the polymorphisms through typing related relatives and/or DNA from an alternative source such as buccal cells might be of help for donor selection.