Ex vivo generation and expansion of autologous anti-tumor CTL lines for adoptive cell therapy in patients with solid tumors

Abstract

12516 Background: Ex-vivo generation of cytotoxic T cell (CTL) lines able to selectively kill autologous tumor cells (TC) is crucial for adoptive cell therapy of cancer. We have described the ex vivo generation of anti-tumor HLA-restricted CTL using CD8-enriched blood mononuclear cells (PBMC) and dendritic cells, pulsed with apoptotic TC as source of tumor antigens (Montagna, Int J Cancer 2004). These cells possess a marked cytotoxicity against TC but very low levels of cytotoxicity against autologous non-malignant controls, suggesting their safe in vivo use. Aim of this study was to evaluate the possibility of generating a large number of anti-tumor CTL lines, in compliance with good manufacture procedure (GMP), to be utilized for adoptive cell therapy. Methods: Four patients with advanced, pretreated solid tumors have been evaluated: 2 sarcoma, 1 renal carcinoma and 1 ovarian carcinoma. Tumor sample was obtained during surgical procedures planned for therapeutic purposes, while PBMC were obtained by a single apheresis. All reagents employed for cultures were produced under GMP. Based on our reported method, the current protocol included two or three rounds of tumor-specific stimulation, followed by two rounds of antigen-independent expansion. Results: We have obtained, after the first round of expansion and in total, a range of 0.5–2×109 and 5–20×109 CTL, respectively. Such large amounts of anti-tumor CTL were generated even if a low number of viable TC was available. In 3 out of 4 anti-tumor CTL lines, >85% of effector cells were CD3+/CD8+. Conclusions: We demonstrated the feasibility of obtaining, in GMP conditions, large quantities of anti-tumor specific CTL suitable for in vivo use. A phase I protocol of lymphoablative therapy followed by CTL transfer in vivo has been designed and submitted for approval to the competent regulatory agency. No significant financial relationships to disclose.