Cytokine-modified tumour cells have been used in clinical trials for immunotherapy of neuroblastoma, but primary tumour cells from surgical biopsies are difficult to culture. Autologous fibroblasts, however, are straightforward to manipulate in culture and easy to transfect using nonviral or viral vectors. Here we have compared the antitumour effect of fibroblasts and tumour cells transfected ex vivo to coexpress interleukin-2 (IL-2) and IL-12 in a syngeneic mouse model of neuroblastoma. Coinjection of cytokine-modified fibroblasts with Neuro-2A tumour cells abolished their in vivo tumorigenicity. Treatment of established tumours with three intratumoral doses of transfected fibroblasts showed a significant therapeutic effect with reduced growth or complete eradication of tumours in 90% of mice, associated with extensive leukocyte infiltration. Splenocytes recovered from vaccinated mice showed enhanced IL-2 production following Neuro-2A coculture, and increased cytotoxicity against Neuro-2A targets compared with controls. Furthermore, 100% of the tumour-free mice exhibited immune memory against tumour cells when rechallenged three months later. The potency of transfected fibroblasts was equivalent to that of tumour cells in all experiments. We conclude that syngeneic fibroblasts cotransfected with IL-2 and IL-12 mediate therapeutic effects against established disease, and are capable of generating immunological memory. Furthermore, as they are easier to recover and manipulate than autologous tumour cells, fibroblasts provide an attractive alternative immunotherapeutic strategy for the treatment of neuroblastoma.