TARGETED DELIVERY OF DNA ENCODING CYTOTOXIC PROTEINS THROUGH HIGH-AFFINITY FIBROBLAST-GROWTH-FACTOR RECEPTORS

Nonviral DNA delivery strategies for gene therapy have generally been limited by a lack of specificity and efficacy. However, ligand-mediate d endocytosis can specifically deliver DNA in vitro to cells bearing t he appropriate cognate receptors. Similarly, in order to circumvent pr oblems related to efficacy, DNA must encode proteins with high intrins ic activities. We show here that the ligand basic fibroblast growth fa ctor (FGF2) can target FGF receptor-bearing cells with DNA encoding th erapeutic proteins. Delivery of genes encoding saporin, a highly paten t ribosomal inactivating protein, or the conditionally cytotoxic herpe s simplex virus thymidine kinase, a protein that can kill cells by act ivating the prodrug ganciclovir, is demonstrated. The saporin gene was codon optimized for mammalian expression and demonstrated to express functional protein in a cell-free assay. FGF2-mediated delivery of sap orin DNA or thymidine kinase DNA followed by ganciclovir treatment res ulted in a 60 and 75% decrease in cell number, respectively. Specifici ty of gene delivery was demonstrated in competition assays with free F GF2 or with recombinant soluble FGF receptor. Alternatively, when hist one H1, a ligand that binds to cell surface heparan sulfate proteoglyc ans (''low-affinity'' FGF receptors), was used to deliver DNA encoding thymidine kinase, no ganciclovir sensitivity was observed. These find ings establish the feasibility of using ligands such as FGF2 to specif ically deliver genes encoding molecular chemotherapeutic agents to cel ls.