Immune response to green fluorescent protein: implications for gene therapy

Green fluorescent protein (GFP) is a widely used intracellular reporter mol ecule to assess gene transfer and expression. A potential use for GFP is as a co-expressed marker, to select and enrich gene-modified cells by flow cy tometry. Processed peptides derived from GFP and presented by the major his tocompatibility complex on the cell surface could potentially induce T cell immune responses against GFP+ cells. This clinical application of GFP is p remature, since in vivo studies on its immunogenicity are lacking. Therefor e, we investigated immune responses against EGFP (enhanced-GFP) in two tran splantable murine models: the BALB/c (H-2(d)) BM185 pre-B leukemia and the C57BL/6 (H-2(b)) EL-4 T cell lymphoma. BM185 and EL-4 cell lines modified t o express high levels of EGFP showed drastic reduction of disease developme nt when transplanted into immunocompetent mice. BM185/EGFP did lead to rapi d development of disease in immunodeficient Nu/Nu mice. Mice surviving BM18 5/EGFP leukemia challenge developed high cytotoxic T lymphocyte (CTL) respo nses against EGFP-expressing cells. Furthermore, immune stimulation against BM185/EGFP cells could also be induced by immunization with EGFP+ transduc ed dendritic cells. The effects of the co-expression of EGFP and immunomodu lators (CD80 plus GM-CSF) were also investigated as an irradiated leukemia vaccine. EGFP co-expression by the vaccine did not interfere with the devel opment of CTLs against the parental leukemia or with the anti-leukemia resp onse in vivo. These results indicate that the immune response against EGFP may interfere with its applicability in gene insertion/replacement strategi es but could potentially be employed for leukemia cell vaccines.