OR26 BW4+ HLA class I and KIR3DL1 allotypic pairing influences licensing and effector function of KIR3DL1+ NK cells

Abstract

Aim The interaction of HLA class I proteins with inhibitory receptors expressed by NK cells impacts their capacity to mediate effector responses via a process called licensing. The strength of these interactions is influenced by polymorphisms in genes encoding both receptor and ligand. Using the interaction between KIR3DL1 and HLA-Bw4 allotypes as a model, we aimed to further understand how the strength of this interaction correlates with licensing of KIR3DL1+ NK cells, assessing both HLA-A and -B Bw4+ molecules and determining the impact of having multiple Bw4+ allotypes of differing ligand strengths. Methods NK cells were purified from HLA-typed, healthy donor PBMC and incubated with HLA class I deficient 221 cells and the expression of CD107a and IFNγ by KIR3DL1+ NK cells assessed by flow cytometry. The expressed KIR3DL1 allotypes were further categorized into five subtypes, spanning ∼95% of the population, using a multiplex PCR. The data was then stratified by donor HLA and KIR3DL1 type and correlated against KIR3DL1/HLA-Bw4 affinity measurements based on KIR3DL1 tetramer binding to a panel of 100 different HLA class I allotypes. Results KIR3DL1+ NK cells from Bw4+ individuals had elevated degranulation and IFNγ responses following coculture with 221 cells compared to those from individuals who lacked Bw4 allotypes. However, there was no strict relationship between Bw4 copy number and the magnitude of the response or cell surface expression of Bw4 allotypes. When the NK response to 221 cells was correlated with the binding strength of the KIR3DL1/HLA-Bw4 interactions, as assessed by KIR3DL1 tetramer binding to bead-bound HLA class I, a spectrum of licencing efficiency was revealed. NK cells from donors with weak KIR3LD1/HLA-Bw4 interactions had more modest responses than those from individuals with combinations that interacted strongly. Some KIR3DL1 allotypes including *002 and *015 showed greater variability in effector function compared to other KIR3DL1 subgroups. When multiple Bw4 allotypes were present, the strongest correlation to NK activation was drawn when the contribution of both Bw4 allotypes was averaged. Conclusions NK cell effector function correlates with KIR3DL1/HLA-Bw4 binding strength, suggesting that licensing is dependent on the strength of the receptor/ligand interaction.