Skewed TCR Repertoire of Vδ1+ γδT Cells in Recipients of Allogeneic Hematopoietic Stem Cell Grafts and Possible Involvement of Epstein-Barr Virus in Clonal Expansion of Vδ1+ T Cells.

Background: We previously demonstrated that stable clonal expansion of Vδ1+ γδT lymphocytes persisted for several years after human allogeneic hematopoietic stem cell transplantation (allo-HSCT). These Vδ1+ T cells are derived from mature T cells in the graft. In the present study, we have extended our observation to learn whether oligoclonal expansions of Vδ1+ γδT lymphocytes might be associated with clinical outcome and GVHD, and whether consensus sequences of the CDR3 region of clonally expanded Vδ1+ T cells would be observed among different individuals. We also examined the possible role for Epstein-Barr virus (EBV) in clonal expansion of Vδ1+ T cells. Methods: Forty-two patients receiving allo-HSCT for hematological malignancies were included in this study. Grafts included bone marrow (n=33), G-CSF mobilized peripheral blood stem cells (n=7) and cord blood (n=2). Clonality of the Vδ1+ T cell subset was determined by CDR3 size spectratyping analysis. Junctional sequences were determined by DNA sequencing. In some experiments, PBMCs from healthy volunteer donors were stimulated with autologous EBV-LCL and were analyzed for clonality of TCRs. Results: CDR3 size spectratyping analysis revealed that twenty-three out of forty-two patients had highly skewed TCR repertoires of the Vδ1+ T cells. There was no apparent association between the oligoclonality of Vδ1+ TCRs and clinical outcome such as GVHD and leukemia relapse. In eight out of seventeen patients examined, the -WGI- amino acid sequence was observed in the CDR3 region of TCRs of clonally expanded Vδ1+ T cells. The -YWG- sequence was observed in four patients. All recipients examined were serologically positive for EBV-VCA IgG and EBNA. Bacterial or fungal components failed to stimulate Vδ1+ T cells to proliferate in vitro, but autologous EBV transformed B cells could induce the expansion of Vδ1+ T cells. The CDR3 size distribution patterns of Vδ1+ TCRs became skewed after stimulation with autologous EBV-LCL, and the T cell clone with the -LEEYWGLPH- CDR3 sequence predominated in the culture with autologous EBV-LCL, whereas this clone was not detectable before culture. Moreover, allogeneic EBV-positive Raji cells also induced the oligoclonal expansion of Vδ1+ T cells carrying the -WGI- or -YWG- junctional sequence. These results suggest the CDR3 structure may contribute to recognition of EBV-associated antigens by Vδ1+ T cells Conclusion: Skewing of the Vδ1+ TCR after allo-HSCT may be the result of the response to infectious antigens widely existing in humans such as EBV. Table 1. Junctional diversity of Vδ1 TCR of γδ T cells expanded in response to autologous EBV-LCL and allogeneic Burkitt lymphoma cells Stimulation δV1 N-D-N Jδ Colony frequency T cell clones appearing more than once are presented. Nil CALGE GLPHALIMWGDLAY TDKLIFGKG 3/20 EBV-LCL CALGE LEEYWGLPH TDKLIFGKG 10/27 CALGE GLPHALIMWGDLAY TDKLIFGKG 7/27 CALG GVLYWGIRR TDKLIFGKG 2/27 CALGE SLWGIRY TDKLIFGKG 2/27 CALGE LGETTPLLGGYSFA LTAQLFFGKG 2/27 Raji CALG VSGLARGGSL KLIFGKG 6/25 CALGE ADWGIRARILY TDKLIFGKG 4/25 CALGE PRAILGDTRIKRMY TDKLIFGKG 4/25 CALGE LEEYWGLPH TDKLIFGKG 3/25 CALGE DPGLPFLWY TDKLIFGKG 2/25 CALG DLNLLWGIRSILPG TDKLIFGKG 2/25