Single locus HLA sequencing with the nanopore technology for HLA disease association diagnosis.

ABSTRACT

Associations between HLA genotype and disease susceptibility encompass almost all the classic HLA loci. The level of typing resolution enabling a correct identification of an HLA disease susceptibility gene depends on the disease itself and/or on the accumulated knowledge about the molecular involvement of the HLA allele(s) engaged. Therefore, the application of Next Generation Sequencing technologies to HLA disease association, which would improve typing resolution, could prove useful to better understand disease severity. In the present study, we tested a nanopore sequencing approach developed by Omixon Biocomputing Ltd, dedicated to on-demand locus typing for HLA and disease, as an alternative to the conventional widely used sequence specific oligoprobe (SSO) approach. A total of 145 DNA samples used in routine diagnosis by SSO were retrospectively analyzed with nanopore technology, for HLA-A*02 immunotherapy decision for A*29, B*27, B*51, B*57 identification in class I, and DRB1, DQA1, and DQB1 for bullous dermatosis, rheumatoid arthritis, diabetes, and celiac disease requests in class II. Each locus was typed in a separate experiment, except for DQB1 and DQA1, which were analyzed together. Concordance between typings reached 100% for all the loci tested. Ambiguities by nanopore were only found for missing exon coverage. This approach was found to be very well adapted to the routine flow imposed by the SSO technique. This study illustrates the use of the new NanoTYPE MONO kit for single locus HLA sequencing for HLA and disease association diagnosis.