Ex vivo generation and expansion of anti-tumor cytotoxic T-cell lines derived from patients or their HLA-identical sibling.
Successful ex-vivo priming and long-term maintenance of anti-tumor cytotoxic T-cell (CTL) lines are preliminary conditions for their use in approaches of adoptive immunotherapy for patients with cancer. We describe the results of a novel procedure for generating in vitro anti-tumor CTL using CD8-enriched peripheral blood mononuclear cells (PBMC) and dendritic cells (DC), pulsed with irradiated tumor cells (TC) as source of tumor antigen. Eight patients were enrolled in our study: 4 sarcoma, 2 renal cell carcinoma, 1 ovarian carcinoma and 1 breast carcinoma. Ten anti-tumor CTL-lines cytotoxic towards patient TC were generated. Five CTL-lines were obtained using both DC and PBMC from the patients (autologous setting). For 5 CTL-lines, DC derived from an HLA-identical sibling were employed (allogeneic setting): patients or siblings PBMC were used to generate CTL-lines in 2 and 3 cases, respectively,. After tumor-specific rounds of stimulation, followed by antigen-independent cycle of expansion, CTL-lines obtained in both autologous and allogeneic setting showed an expansion of the absolute number of cultured cells. In 6 of 10 CTL-lines, the majority of effector cells (>70%) were CD3+/CD8+, while in the remaining 4, 40-70% of effector cells were CD3+/CD4+. Both CD8+ and CD4+ T cells displayed anti-tumor cytotoxic activity. Spectratyping analysis of the TCR-Vbeta subfamilies revealed a preferential expansion of oligoclonal populations in 18 of 24Vbeta subfamily. Altogether these results demonstrate that our experimental approach is suitable for efficiently generating and expanding anti-solid tumor CTL to be used for adoptive immunotherapy.
Montagna
Daniela
D
Immunology Laboratory and Pediatric Hematology/Oncology, IRCCS Policlinico San Matteo, University of Pavia, Piazzale Golgi, 27100 Pavia, Italy. d.montagna@smatteo.pv.it
Schiavo
Roberta
R
Gibelli
Nadia
N
Pedrazzoli
Paolo
P
Tonelli
Roberto
R
Pagani
Sara
S
Assirelli
Elisa
E
Locatelli
Franco
F
Pession
Andrea
A
Fregoni
Vittorio
V
Montini
Elisa
E
Da Prada
Gian Antonio
GA
Siena
Salvatore
S
Maccario
Rita
R
D000293:Adolescent; D000328:Adult; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D002460:Cell Line; D003602:Cytotoxicity, Immunologic; D005260:Female; D006650:Histocompatibility Testing; D006801:Humans; D008297:Male; D008875:Middle Aged; D009369:Neoplasms / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*