Successful in vitro priming of EBV-specific CD8+ T cells endowed with strong cytotoxic function from T cells of EBV-seronegative children.
Epstein-Barr virus (EBV)-seronegative transplant recipients are at high risk of developing EBV-associated post-transplant lymphoproliferative disorder (PTLD), and would maximally benefit from an EBV-directed T-cell therapy for prevention or treatment of PTLD. So far, efforts to activate CD8+ EBV-specific cytotoxic T lymphocytes (CTL) endowed with high specific cytotoxicity from EBV-seronegative children have failed. We compared the CD8+ CTL priming efficiency of three different modified activation protocols, based on lymphoblastoid cell lines (LCL) stimulation potentially enhanced by either LCL presentation through dendritic cells, or selection of IFN-gamma+ cultured cells, or culture in the presence of rhIL-12 and rhIL-7, according to the standard protocol for reactivation of EBV-specific CTL. We found that only specific LCL stimulation in the presence of rhIL-12 and rhIL-7 was able to reproducibly expand EBV-specific CD8+ CTL endowed with strong cytotoxic activity from truly EBV-seronegative children. The lines thus activated, which included specificities toward EBV latent and lytic proteins, showed high percentage CD8+ T cells, with <10% naïve CD8+/CCR7+/CD45RA+ cells. Overall, the total number of CD8+ central memory cells, and of CCR7 T-cell effectors was comparable to that observed in healthy EBV-seropositive controls. In conclusion, it is feasible to activate EBV-specific CD8+ CTL with suitable characteristics for in vivo employment from EBV-seronegative children.
Comoli
P
P
Laboratory of Transplant Immunology and Pediatric Hematology/Oncology, IRCCS Policlinico S. Matteo, 27100 Pavia, Italy. pcomoli@smatteo.pv.it
Ginevri
F
F
Maccario
R
R
Frasson
C
C
Valente
U
U
Basso
S
S
Labirio
M
M
Huang
G-C
GC
Verrina
E
E
Baldanti
F
F
Perfumo
F
F
Locatelli
F
F
D000328:Adult; D000914:Antibodies, Viral / Q000097:blood*; D016026:Bone Marrow Transplantation / Q000009:adverse effects*; D018414:CD8-Positive T-Lymphocytes / Q000821:virology*; D002648:Child; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D005260:Female; D004854:Herpesvirus 4, Human / Q000276:immunology*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D018664:Interleukin-12 / Q000494:pharmacology; D015851:Interleukin-7 / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000821:virology; D008232:Lymphoproliferative Disorders / Q000209:etiology; D008297:Male; D010641:Phenotype; D011994:Recombinant Proteins / Q000494:pharmacology; D013602:T-Lymphocytes, Cytotoxic / Q000821:virology*
D000914:Antibodies, Viral; D015851:Interleukin-7; D011994:Recombinant Proteins; D018664:Interleukin-12; D007371:Interferon-gamma