Generation of Interleukin-2-Secreting Melanoma Cell Populations from Resected Metastatic Tumors

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Abstract
This study aimed to determine the feasibility of producing patient-specific, interleukin-2 (IL-2)-secreting tumor cell vaccines for the treatment of metastatic melanoma. Primary tumor cell cultures were established from 26/33 resected metastatic melanoma samples. Recombinant retroviral gene transfer and expression in these cultures was optimized using an amphotropic, defective retrovirus carrying the LacZ gene. All cell cultures were infectable; those that proliferated more rapidly were infected at a higher frequency. Addition of fibroblast growth factor to the culture medium increased the rate of cell proliferation and the efficiency of infection. A single infection with an identical retrovirus carrying a human IL-2 cDNA resulted in the generation of unselected cell populations secreting up to 300 units IL-2/106 cells · 48 hr. Multiple infections increased the level of IL-2 secretion to 5,000 units/106 cells · 48 hr. The recombinant viral genome could be detected at approximately single copy in the multiply infected cells; no helper virus was detected. IL-2 secretion from infected cultures was maintained following cryopreservation and x-irradiation. These data demonstrate that heterogeneous tumor cell populations secreting IL-2 can be generated from individual patients to be used as autologous, irradiated cell vaccines. Injection of IL-2-secreting tumor cells has been shown to stimulate the host antitumor immune response and prevent metastasis in animal tumor models. This approach is currently being applied to the treatment of metastatic human cancer. We describe the optimization of gene transfer in primary cultures of human metastatic melanoma cells. Nonselective recombinant retroviral infection was used to generate large numbers of tumor cells secreting high levels of IL-2. This protocol will maintain any antigenic heterogeneity of the original tumor and provides a cell vaccine suitable for clinical application.