Selective tumor delivery of therapeutic genes is instrumental to the success of numerous cancer gene therapy protocols. The objective of this research proposal, which is a key step in our long-term goal to develop safe and efficient vectors for systemic human cancer gene therapy, is to prepare polyplexes capable of selective surface presentation of targeting ligands in response to small physiologically relevant decrease of pH. Well defined, end-functionalized copolymers of N-isopropylacrylamide and 1-vinylimidazole, which display pH-induced globule-to-coil phase transition at body temperature, will be synthesized by RAFT polymerization, conjugated with PEG, protamine, and targeting ligands, and used to prepare polyplexes with plasmid DNA and oligonucleotides. We hypothesize that pH-triggered phase transition will display targeting ligands selectively at slightly acidic pH and permit pH-selective cellular uptake. The phase state-controlled tropism of polyplexes will be investigated by analyzing pH-dependent binding of biotin-containing polyplexes to immobilized avidin and by measuring the levels of pH-dependent cellular uptake of non-specifically-binding and integrin- and asialoglycoprotein-binding polyplexes in cell culture. To our knowledge, this is the first use of stimuli-responsive polymers to control tropism of synthetic self-assembly vectors for gene delivery. While preliminary results demonstrating the feasibility of controlling surface properties of polyplexes by the phase state of the stimulus-responsive copolymers are presented, the modulation of ligand binding by pH needs to be clearly demonstrated to establish the feasibility of this approach. Funding through the NIBIB R21 mechanism will allow us to test the validity of our hypothesis and establish a design platform for further development of polyplexes with stimuli-responsive tropism. The polyplexes with pH-controlled tropism, proposed here, will be exploited in the development of gene delivery vectors that recognize and selectively display targeting ligands in slightly acidic extracellular pH of many solid tumors; thus minimizing the need for tumor-specific ligands.