Aim: Killer cell immunoglobulin-like receptors (KIRs) expressed on Natural killer (NK) cells play a vital role in cancer immune surveillance by screening of human leukocyte antigen (HLA) class I on cancer cells. Inhibitory KIR3DL1 exhibited hierarchical recognition for Bw4+HLA-I which is influenced by polymorphisms as well repertoire of presented peptides which may lead to various clinical outcomes. This study set out to determine the varying potency of peptides in differential KIR3DL1 recognition of HLA-A*24:02.
Methods: 3DL1*001/*005/*015 were expressed in HEK293 F/S cells while HLA-A*24:02 heavy chain and B2-microglobulin were produced as inclusion bodies and refolded in the presence of peptide. Purified proteins were utilized to define the molecular/structural basis underpinning these interactions using surface plasmon resonance (SPR) assay and X-ray crystallography.
Results: Measuring the binding affinity of 3DL1 to HLA-A*24:02 exhibited a varying potency of endogenous and cancer peptides in inhibition of 3DL1*001/*005/*015+ NK cells. The interlineage *001 allotype was less sensitive to peptide variation in HLA-A*24:02 recognition compared with *005/*015. Furthermore, self-peptides presented by HLA-A*24:02 mediated more robust inhibition (tolerance) of 3DL1+NK cells than cancer peptides. Among tumor specific peptides, EYLQLVFGI as part of melanoma-associated antigen expressed in gastric/colorectal carcinomas could efficiently bind to three allotypes of 3DL1 offering a mechanism of evasion from NK cell attack. The most striking observation revealed that the peptide discrimination toward the carboxyl terminus at position (p)7 and 8 can remarkably influence the inhibition of KIR3DL1*001/*005/*015+NK cells. Solving ternary structure of the KIR3DL1*001-HLA-A*24:02-NFLSRSFYW displayed no direct contact between peptide and KIR3DL1 domains (D0, D1, D2), yet a network of water mediated bridges connected tyrosine at PΩ-1 of peptide to metionne165, leucine166, alanaine167 of D1 of 3DL1*001 that may in fact facilitate binding of 3DL1*001 to HLA-A*24:02.
Conclusion: These results provide additional evidence for peptide specificity of KIR3DL1 by highlighting the role of aromatic amino acids at p7/p8 in enhancing its binding affinity to ligand. Further structural analysis would be worthwhile for designing peptide based immunotherapeutic agents as a personalized therapy for cancer patints.
