Multicenter phase II study of matured dendritic cells pulsed with melanoma cell line lysates in patients with advanced melanoma.

BACKGROUND: Several single center studies have provided evidence of immune activation and antitumor activity of therapeutic vaccination with dendritic cells (DC) in patients with metastatic melanoma. The efficacy of this approach in patients with favorable prognosis metastatic melanoma limited to the skin, subcutaneous tissues and lung (stages IIIc, M1a, M1b) was tested in a multicenter two stage phase 2 study with centralized DC manufacturing. METHODS: The vaccine (IDD-3) consisted 8 doses of autologous monocyte-derived matured DC generated in serum-free medium with granulocyte macrophage colony stimulating factor (GM-CSF) and interleukin-13 (IL-13), pulsed with lysates of three allogeneic melanoma cell lines, and matured with interferon gamma. The primary endpoint was antitumor activity. RESULTS: Among 33 patients who received IDD-3 there was one complete response (CR), two partial responses (PR), and six patients had stable disease (SD) lasting more than eight weeks. The overall prospectively defined tumor growth control rate was 27% (90% confidence interval of 13-46%). IDD-3 administration had minimal toxicity and it resulted in a high frequency of immune activation to immunizing melanoma antigens as assessed by in vitro immune monitoring assays. CONCLUSIONS: The administration of matured DC loaded with tumor lysates has significant immunogenicity and antitumor activity in patients with limited metastatic melanoma. CLINICAL TRIAL REGISTRATION: NCT00107159.

Analysis and characterization of antitumor T-cell response after administration of dendritic cells loaded with allogeneic tumor lysate to metastatic melanoma patients.

The primary goal of cancer vaccines is to induce CD8+ T cells specific for tumor-associated antigens (TAA) but the characterization of these cells has been difficult because of the low sensitivity of ex vivo assays. Here, we focused on TAA-specific CD8+ T-cell responses in melanoma patients after vaccination with autologous dendritic cells loaded with lysates derived from allogeneic tumor-cell lines (Lysate-DC). Out of 40 patients treated, 16 patients developed immune response to tumor-cell lysate and/or CD8+ T cells specific for differentiation and cancer-testis antigens. TAA-specific CD8+ T-cell responses were detected by interferon (IFN)-gamma enzyme-linked immunospot after in vitro sensitization and were, either transient during the treatment period or delayed, that is, observed after completion of all vaccinations. We could not correlate these immune responses to clinical data as none of the patients achieved an overall objective response according to Response Evaluation Criteria in Solid Tumors criteria. Three patients were reported as stable disease and 10 patients presented evidence of antitumor activity. We found that TAA-specific T cells characterized in 4 patients produced perforin ex vivo, but no IFN-gamma in enzyme-linked immunospot. Differential expression of IFN-gamma and perforin was also observed for viral-specific T cells. Altogether, our results show that Lysate-DC therapy elicited tumor-specific CD8+ T cells nonlimited to human leukocyte antigen-A2+ patients, with some T cells secreting perforin ex vivo and IFN-gamma only after restimulation. The differential expression of perforin and IFN-gamma by antitumor and antiviral CD8+ T cells supports that the sole use of IFN-gamma production to monitor T cells overlooks functional T-cell subpopulations triggered by vaccines.

Generation of dendritic cell-tumor cell hybrids by electrofusion for clinical vaccine application.

Vaccination with hybrids comprising fused dendritic cells (DCs) and tumor cells is a novel cancer immunotherapy approach designed to combine tumor antigenicity with the antigen-presenting and immune-stimulatory capacities of DCs. For clinical purposes, we have incorporated a large-scale process for the generation of clinical-grade DCs together with novel electrofusion technology. The electrofusion system provides for ease and standardization of method, efficient DC-tumor cell hybrid formation, and large-quantity production of hybrids in a high-volume (6-ml) electrofusion chamber. In addition, we have evaluated DC electrofusion with a variety of allogeneic human tumor cell lines with the rationale that these tumor cell partners would prove a ready, suitable source for the generation of DC-tumor cell hybrid vaccines. The DC production process can generate 6x10(8) to 2x10(9) DCs from a single leukapheresis product (approximately 180 ml). As determined by FACS analysis, electrofusion of 6x10(7) total cells (1:1 ratio of DC and tumor cells) resulted in a consistent average of 8-10% DC-tumor cell hybrids, irrespective of the tumor type used. Hybrids were retained in the population for 48 h postfusion and following freezing and thawing. Upon pre-irradiation of the tumor cell partner for vaccine purposes, the overall fusion efficiency was not altered at doses up to 200 Gy. Evaluation of DC-tumor cell hybrid populations for their ability to stimulate T-cell responses demonstrated that electrofused populations are superior to mixed populations of DCs and tumor cells in generating a primary T-cell response, as indicated by IFN-gamma release. Moreover, hybrids comprising HLA-A*0201 DCs and allogeneic melanoma tumor cells (Colo 829 cell line) stimulated IFN-gamma secretion by antigen-specific CD8+ T cells, which are restricted for recognition of a melanoma gp100 peptide antigen (gp100(209-217)) within the context of the DC HLA haplotype. Maturation of the DC-Colo 829 cell hybrid population served to further improve this T-cell gp100-specific response. Overall, our results are promising for the large-scale generation of electrofused hybrids comprising DCs and allogeneic tumor cells, that may prove useful in human vaccine trials.