ABSTRACT
Tumor cells transduced with the IL-4 gene demonstrate reduction of growth associated with macrophage and eosinophilic infiltrates, generation of cytotoxic T-cells, and protective immunity. Using murine IL-4 retroviral vectors, murine fibroblasts and tumors that produce from 50 to 5000 U of IL-4/10(6) cells per 24 hours as determined by ELISA and bioassay were successfully transduced. In blinded studies using C57BL/6 and BALB/c mice, we have shown that tumor growth can be inhibited (mean delay of 10 days compared with controls; P < .05) and in some cases, completely suppressed by the coinjection of viable tumor with IL-4-producing fibroblasts (tumor free > 100 days; P < .001). Animals that are able to reject an initial tumor inoculate can also completely reject subsequent parental tumor challenge of 10(5) cells (P < .001) while challenge of 10(6) parental tumor cells results in a significant delay of tumor induction (P < .05). In addition, immunization with IL-4 transduced fibroblasts and irradiated tumor cells resulted in complete suppression of parental tumor challenge in animals that received the high-dose IL-4 delivery. Finally coadministration of systemic IL-2 led to enhancement of IL-4 gene therapy resulting in a 20-day delay of preestablished tumor growth compared with controls (P < .05).
Subject(s)
Fibroblasts/transplantation , Genetic Therapy , Immunologic Factors/therapeutic use , Interleukin-2/therapeutic use , Interleukin-4/administration & dosage , Neoplasms, Experimental/therapy , Recombinant Fusion Proteins/metabolism , Animals , Cells, Cultured/transplantation , Combined Modality Therapy , Fibroblasts/radiation effects , Genes, Reporter , Genetic Vectors/genetics , Graft Rejection , Immunization , Immunologic Factors/administration & dosage , Immunologic Factors/genetics , Interleukin-4/genetics , Interleukin-4/therapeutic use , Lung Neoplasms/prevention & control , Lung Neoplasms/secondary , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Neoplasm Transplantation , Neoplasms, Experimental/immunology , Neoplasms, Experimental/pathology , Retroviridae/genetics , Transfection , Tumor Cells, Cultured/immunology , Tumor Cells, Cultured/transplantationABSTRACT
Introduction of the interleukin-4 (IL-4) gene into cells derived from human tumor tissue provides a means for generating a specific tumor vaccine. Such a vaccine could be produced by either transducing tumor-derived stromal cells with the IL-4 vector and coinjecting tumor cells, or by transducing the tumor cells themselves. We have developed a protocol for culturing cells from non-small cell lung tumors and routinely produce tumor cultures from 25% of tumors, and stromal cultures from > 80% of specimens. Several of these cultures were transduced with the incompetent retroviral vector G1NaSvi4.25, which encodes the human IL-4 cDNA and the G418-resistance gene. Infection of cells by viral titers of 2-5 x 10(4) plaque-forming units/ml, and a multiplicity of infection of 0.1:1 to 1:1 yielded transfer efficiencies of 3.3-32.0 transfectants per 10(4) cells in six of eight attempts. Following selection with the neomycin analog G418, IL-4-producing cells were isolated. IL-4 titers ranged from 142 to 593 U/ml/10(6) in a 24-h collection. Successful transfer of the IL-4 gene was demonstrated by polymerase chain reaction amplification of cDNA derived from reverse-transcribed total RNA, by immunohistochemistry, and by enzyme-linked immunosorbent assay. The IL-4-producing cells were shown to stimulate the proliferation of autologous peripheral blood lymphocytes in one individual by 7.5-fold over control and by 4.1-fold over non-IL-4 producing tumor cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Subject(s)
Carcinoma, Non-Small-Cell Lung/therapy , Gene Transfer Techniques , Interleukin-4/genetics , Lung Neoplasms/therapy , Base Sequence , Carcinoma, Non-Small-Cell Lung/immunology , Cell Line , Genetic Therapy , Humans , Interleukin-4/biosynthesis , Lung Neoplasms/immunology , Lung Neoplasms/metabolism , Lymphocyte Activation , Molecular Sequence Data , Stromal Cells/metabolism , Transcription, GeneticABSTRACT
Melanoma represents the single best example of a human tumor that has been shown to elicit specific T-cell reactivity. The responsiveness of some patients with metastatic melanoma to treatment with the prototypic T-cell growth factor (TCGF), interleukin-2 (IL-2), indicates that T cells play a role in antitumor immunity. Interleukin-4 (IL-4), another TCGF that has been administered clinically to humans, was not associated with tumor response in our trials conducted at the Surgery Branch of the National Cancer Institute. Combination trials of IL-2 with IL-4 have shown no increase in responsiveness of melanoma or other tumors when compared to IL-2 alone. However, enhanced expansion of tumor-infiltrating lymphocytes (TILs) in vitro has been observed with combinations of low-dose IL-2 and IL-4. We have begun a study evaluating the trafficking of such expanded lymphocytes following their adoptive transfer in association with systemic administration of IL-2 and IL-4. We have established several TIL cultures from fresh tumor samples, maintained them in long-term culture, and marked them with the neomycin phosphotransferase gene using the LNL6 retroviral vector. Such TILs appear to demonstrate no notable alterations in phenotype or cytolytic activity when compared to their nontransduced counterparts. In addition to IL-2 and IL-4, there are a variety of other novel TCGFs that are now available for evaluation in preclinical and clinical trials. IL-7 induces proliferation and lymphokine-activated killer (LAK) cell activity from human peripheral blood mononuclear cells.(ABSTRACT TRUNCATED AT 250 WORDS)
