P059 Using NGS in solid organ transplantation to resolve anti-HLA antibodies to self antigens

Abstract

Next generation sequencing (NGS) is an innovative technology that is redefining the landscape of human molecular genetic testing. NGS technology has entered the clinical lab space with validated machines and reagents to streamline the process of HLA typing. This case highlights the utility of HLA typing by NGS for a solid organ laboratory. A 45 year old African American male presented to our center for new patient evaluation. The patient had received a previous kidney transplant in June 2013 at another institution. We performed HLA typing by SSO method and screened the patient for anti-HLA antibodies by Flow PRA (FPRA) and reflexed to single antigen bead identification using One Lambda reagents. The patient was HLA typed as DRB1∗10:XX, DRB1∗14:04; DQA1∗01:MV (01/04/05), DQA1∗01:SXYS (01/04/05/12) with DQB1∗05:XX, DQB1∗05:XX by SSO. We confirmed the patient’s HLA typing with the previous transplant center. Due to previous sensitizing events, the patient’s FPRA was 99% class I and 100% class II. Examination of the class II single antigen bead profile revealed strong DR14 and DQ5 antibodies. (DRB1∗14:54, MFI:10,640; DRB1∗14:01, MFI:10299; DRB1∗14:02, MFI: 9379) (DQB1∗01:02-DQB1∗05:02, MFI: 15707) (DQB1∗01:01-DQB1∗05:01, MFI: 7141). For further resolution of the patient’s DR and DQ typings we performed NGS high resolution typing using Illumina TruSight v2 sequencing panel. The patient was typed (DRB1∗10:01:01, DRB1∗14:04:01) (DQA1∗01:04:02, DQA1∗01:05:01) with (DQB1∗05:01:01, DQB1∗05:03:01). DQA1∗01:04:02 was not common and well defined, differing from DQA1∗01:04:01 at position 5433 A→C in exon 3 leading to a synonymous base substitution. The DR and DQA-DQB pairs of the patient found through sequencing differ from the DR14 and DQ5 single antigen beads used for calling unacceptables. The patient was not generating anti-HLA antibodies to self, but rather common DR DQA-DQB allele variants. We confirmed there was no self-reactivity by auto crossmatch which was negative by Flow Cytometry and Cytotoxicity. Given that deceased donor typing is performed at lower resolution and single antigen beads are unable to cover all HLA alleles, a combination of virtual XM and real time crossmatches will need to be performed to safely get this patient re-transplanted.