Killer Immunoglobulin-Like Receptor Allele Determination Using Next-Generation Sequencing Technology

Area of Science:

• Immunogenetics
• Transplantation Immunology
• Genomic Technologies

Background:

• The impact of natural killer (NK) cell alloreactivity on hematopoietic stem cell transplantation (HSCT) outcomes remains debated due to complex graft, HLA, and killer cell immunoglobulin-like receptor (KIR)/KIR ligand genetic factors.
• KIR genes exhibit significant polymorphism in gene content, copy number, and alleles, potentially influencing KIR+ NK cell phenotype, function, and alloreactivity post-HSCT.

Purpose of the Study:

• To develop and validate a next-generation sequencing (NGS) method for investigating broad KIR allele polymorphisms.
• To assess the impact of KIR allelic variations on donor KIR+ NK cell phenotype and function in the context of HSCT alloreactivity.

Main Methods:

• Development of an NGS-based technology on a MiSeq platform for high-resolution KIR typing.
• Utilized genomic DNA from well-characterized cell lines for method validation.
• Employed two distinct bioinformatic pipelines (BiRD and Profiler) for sequence read attribution and KIR allele assignment.

Main Results:

• Demonstrated successful long-range KIR gene amplifications and complete sequencing of all KIR genes with high read depth (mean 317×) and mapping percentage (mean 93%).
• Achieved a 95% concordance rate for KIR typing compared to previous exome capture data for centromeric and telomeric KIR genes.
• Confirmed the reliability and specificity of the NGS method for comprehensive KIR allelic polymorphism investigation.

Conclusions:

• NGS technology is a robust tool for investigating extensive KIR allelic polymorphism.
• This advancement enhances understanding of KIR+ NK cell alloreactivity in HSCT.
• Findings contribute to knowledge regarding the role of KIR+ NK cells in controlling viral infections and diseases.