Association Of Killer Cell Immunoglobin-like Receptor Gene With Bone Marrow Failure Syndromes And The Outcome Following HLA-identical Sibiling Hematopoietic Stem-cells Transplantation

Object: To investigate the correlation of killer immunoglobulin-like receptors gene polymorphism with bone marrow failure syndromes and study the effect of KIR-HLA receptor-ligand model on NK cell alloreactivity which predicted the outcomes in HLA-identical sibiling hematopoietic stem-cells transplantation(HSCT),For further study, investigate the effect of human NK cells from donor by blocking the inhibitory receptors KIR2DL1 and KIR2DL3 with monoclonal antibody CD158a and CD158b,to explore the possible mechanism about donor NK cells decrease the incidence of graft versus host disease (GVHD).Method: SSP-PCR was used to exam the genotypic makeup of KIR in patients with AA,MDS and 15 pairs of patients and their HLA-identical sibiling HSCT donors in our department. Peripheral blood mononuclear cells(PBMC) from 15 pairs of patients and their donors were extracted and preserved, donor NK cells were isolated by DYNABEADS UNTOUCHED HUMAN NK sorting kit. Methyl thiazolyl tetrazolium(MTT) reduction assay was used to detect donor NK cells killing activity against recipients DC. To observe the different NK killing activity before and after KIR receptors blockade, the anti-CD158a,CD158b monoclonal antibody was used to block KIR inhibitory receptors.Results: (1)All the 16 KIR genes which had been prescribed were identified. (2)The frequencies of KIR-2DS1,2DS2,2DS3,2DS5 and 3DS1 genes were showed increased in patients with AA,MDS than in healthy controls. The patients with AA had lower frequency of KIR-2DS5 than the patients with MDS. (3)Among all the 15 donor/recipient pairs,8 donors with KIR2DL2/L3 could find corresponding HLA-C1 ligands in their recipients;3 donors with KIR2DL1 could find corresponding HLA-C2 ligands in their recipients;9 donors with KIR3DL1 could find corresponding HLA-Bw4 ligands in their recipients;6 donors with KIR3DL2 could find corresponding HLA-A3,A11 ligands in their recipients. The incidence rate ofⅢ-Ⅳgrade acute graft versus host disease (aGVHD) was 0% and 35.7% in host versus graft(HVG) KIR ligand-mismatching pattern and KIR receptor-ligand matched pattern, HVG KIR ligand-mismatching pattern also had the lower incidence rate ofⅢ-Ⅳgrade aGVHD than GVH KIR ligand-mismatching pattern(0 vs 7.1%), but the total incidence rates of aGVHD or cGVHD showed no significant differences between HVG and GVH or KIR receptor-ligand matched pattern. Either of two donors who had activating KIRs(KIR2DS1,KIR2DS2) found corresponding ligands in their recipients occurred severe aGVHD. In the setting of our HLA-identical sibiling HSCT, KIR-ligand mismatch was associated with better rate of overall survival(OS) and lower rate of transplant related mortality(TRM). (4)The killing effect of donor NK cells significantly enhanced after inhibitory receptor KIRs blockades, The killing effect became stronger when the dose of antibody raised up.Conclusion: The increased frequencies of activated KIRs in patients with MDS and AA suggest that the abnormal immuno-Genetic might be related to the pathogenesis of bone marrow failure syndromes. KIR genetic background of Donors and recipients is correlate with the outcome of HLA-identical sibiling HSCT in incidence rates of aGVHD,OS and TRM. To avoid choosing the donors who had GVH receptor-ligand matched pattern could improve the prognosis of recipients. It is demonstrated that the killing effect of alloreactive NK cells could be enhanced by KIR functional modification in vitro, and it could be used as a new strategy of immunization therapy for GVHD.