Upregulation of TCRζ Chain Overcome T Cell Immunodeficiency in Patients with Chronic Myeloid Leukemia

Abstract 4719

TCRζ chain is the key molecular of TCR signaling, the defective TCRζ chain not only decreases the expression of TCR on the T cell surface and the quantity of circulation T cells, but also influences T cell activation and proliferation. Previous studies indicated that the obvious downregulation of TCRζ chain was one of main factors which caused T cell immunodeficiency in patients with chronic myeloid leukemia (CML), which lead to dysfunction of immune supervision to tumor. This study was undertaken to explore the possibility that forced expression of TCRζ chain may restored the T cell immune function. In present study, the freshly CD3 ⁺ T cells were isolated by MACS from de novo CML patients; freshly T cells were transfected with TCRζ chain recombinant vector (TCRζ-IRES-EGFP) and control vector (pIRES2-EGFP) by nucleoporation technique. The transfection efficiency was detected by FCM at 18 hours post-transfection, and TCRζ chain protein and its phosphorylation were detected by Western blotting after activation with OKT3 antibody for 1 minute. The supernatants and RNA were collected from transfected cells stimulated with OKT3 and anti-CD28 antibody, for analysis of IL-2 levels and the mRNA expression of ZAP70 and NF-κB. The results showed the transfection efficiency of TCRζ chain vector and control vector construct was 72.16±6.95% and 73.4±7.90% in CML T cells from different patients respectively. In T cells transfected with TCRζ chain, the expression of TCR chain was increased, the IL-2 production induced by OKT3 and anti-CD28 antibody in TCRζ chain transfected T cells (175.1±66.3pg/mL) were higher than that from control group (107.6±65.5pg/mL) (n=6, p=0.039). The bother expression levels of ZAP-70 and NF-κ B in the experimental group was higher than the control group (n = 4, p < 0.05), moreover the expression level between ZAP-70 and NF-κB showed linear correlation (n = 4, p = 0.013, r = 0.98). In conclusion, the results indicate that upregulation of deficient TCRζ chain may reverse the TCR/CD3-mediated signaling abnormalities, which may improve the T cell immune function in patients with CML.

This study was supported by grant from the Key project of Natural Science Foundation of Guangdong Province, China (No. 9251063201000001).