Phase I/II study of treatment with matured dendritic cells with or without low dose IL-2 in patients with disseminated melanoma.

BACKGROUND: In the present study, we have examined whether treatment of patients with metastatic melanoma with matured dendritic cell (DC) vaccines with or without low dose IL-2 may improve treatment outcomes. METHODS: Sixteen patients received DC vaccines (DCs) sensitized with autologous melanoma lysates and 18 patients received DCs sensitized with peptides from gp100, MART-1, tyrosinase, MAGE-3.A2, MAGE-A10 and NA17. IL-2 was given subcutaneously (sc) at 1 MU/m2 on the second day after each injection for 5-14 days in half of each group. DCs were given by intranodal injection. RESULTS: There were 2 partial responses (PR) and 3 with stable disease (SD) in the nine patients receiving DCs + peptides + IL-2, and 1 PR and 1 SD in nine patients treated with DCs + peptides without IL-2. There were only two patients with SD in the group receiving DCs + autologous lysates and no IL-2. Median overall survival for all patients was very good at 18.5 months but this was most probably due to selection of a favourable group of patients for the study. There was no significant difference in survival between the groups by log rank analysis. Treatment was not associated with significant side effects. The quality and yield of the DCs in the preparations were generally good. CONCLUSIONS: We conclude that mature DC preparations may be superior to immature DC preparations for presentation of melanoma peptides and that IL-2 may increase clinical responses to the DCs plus peptides. However, in our view the low response rates do not justify the cost and complexity of this treatment approach.

Phenotype and morphology of murine NC-1.1+ natural cytotoxic cells.

The morphology and phenotype of cells identified by the anti-NC-1.1 mAb, 1C4, were studied in CBA and (CBA x C57BL/6) F1 mice. This mAb blocked splenic natural cytotoxic (NC) cell activity in vitro and in vivo. Single colour flow cytometric analysis showed that 6 +/- 1% of all CBA splenocytes were NC-1.1+, and that granular cells of varying sizes, as defined by their forward and side light scatter, contained the highest proportion of NC-1.1+ cells (29 +/- 7%). When analysed by two colour flow cytometry, the large and the granular NC-1.1+ spleen cells in CBA mice were shown to co-express Thy-1.2 and L3T4 (< or = 33%), Mac-1 (< or = 26%), IgM (< or = 60%) and FcR gamma II and J11d (< or = 100%). In contrast, T cell markers were not co-expressed on the small agranular NC-1.1+ spleen cells. This pattern of marker expression was evident in CBA spleens from 2 days of age. When (CBA x C57BL/6) F1 spleen cells were similarly analysed, 82% of granular NC-1.1+ cells also co-expressed NK-1.1. Single colour analysis of CBA bone marrow, thymus, lymph node and peripheral blood leucocytes revealed that < or = 10% of all cells examined were NC-1.1+ while the most granular cells in these organs were 16-43% NC-1.1+. These results support the 'horizontal lineage' theory that NC cells are cells of different lymphohaemopoietic cell lineages at particular stages of differentiation.

Correlation of growth of tumours in NC-cell-depleted mice with NC- and NK-cell-mediated lysis in vitro.

The anti-tumour surveillance activity of natural cytotoxic (NC) cells was studied in vivo using the transplantable tumours WEHI-164 fibrosarcoma, MPC-II plasmacytoma, WEHI-7 T-lymphoma, B16 melanoma and EL-4 thymoma in syngeneic and semi-allogeneic mice. Experimentally, mice were treated with the anti-NC-I.I monoclonal antibody (MAb) IC4 to abrogate splenic NC activity. This was followed by s.c. inoculation of MTD100 doses of the tumours. Comparison of the diameters of the tumours in the anti-NC-I.I-treated mice with control mice using non-parametric statistics showed significantly faster growth of WEHI-164 (p less than 0.01), MPC-II (p less than 0.05) and WEHI-7 (p less than 0.05) when the mean tumour diameters were less than 15 mm in the anti-NC-I.I-treated mice. Significantly faster growth was also observed in anti-NC-I.I-treated mice with the B16 tumour (p less than 0.05), but at a later stage of growth, when the tumour diameter was greater than 15 mm. In vitro, WEHI-164, MPC-II and WEHI-7 were shown to be predominantly sensitive to lysis by mouse splenic NC cells, while B16 was predominantly lysed by splenic natural-killer (NK) cells. Anti-NC-I.I treatment of mice did not affect the growth of EL-4 in vivo and in vitro experiments with anti-NK-I.I and anti-NC-I.I MAb indicated that this tumour was lysed by sub-sets of NK and NC cells distinct from those which lysed the other tumours. We conclude that, in mice at least, NC cells have an in vivo role in controlling the growth of some transplantable tumours, and this correlates with the in vitro NC cell lysis of these same tumours.

Production of a monoclonal allo-antibody to murine natural cytotoxic cells.

A mouse IgG1 monoclonal antibody (1C4), which recognizes a cell surface molecule on murine natural cytotoxic (NC) cells was produced. By flow cytometry, 1C4 preferentially reacted with less than 5% of fresh CBA spleen cells and 20-50% of CBA-interleukin-3 (IL-3) cells, an in vitro derived NC-like cell line. In vitro treatment of spleen cells from a number of inbred mouse strains either with 1C4 alone or 1C4 coupled to dynabeads markedly decreased or abolished NC activity of the cells against 51Cr-labelled WEH1-164 targets. Splenic NC activity of these same mouse strains was also reduced or abolished by in vivo administration of 1C4. The effect was evident within 2 h of treatment and persisted for at least 1 week. In contrast 1C4 had little or no effect on splenic NK activity against 51Cr-labelled YAC-1 targets over the same range of experiments in vitro and in vivo. Results of strain surveys for both in vitro and in vivo reduction of splenic NC activity by 1C4 treatment showed that CBA, C57BL/6, BALB/c and NZB mice were positive and CE and DBA/2 mice were negative, indicating that 1C4 recognizes an allo-antigen on mouse NC cells. This allo-antibody has been designated NC-1.1, and thus 1C4 is an anti-NC-1.1 monoclonal antibody.