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Receptor-mediated gene transfer in vivo

by Perales, Jose Carlos

Abstract (Summary)
Receptor-mediated gene transfer is an attractive method for therapeutically correcting human genetic diseases since it permits the targeting of DNA to cellular receptors in specific tissues of adult animals. The gene delivery vehicle is composed of the DNA of interest, a DNA binding moiety (i.e., poly(L-lysine)) and a ligand for the target receptor. Genes introduced by this technique have been shown to be expressed in the target tissue for varying periods. However, to be useful for gene therapy, it is critical that both the chemical properties and physical interactions of the reagents involved in the design of the DNA delivery vehicle (DNA/ligand complex) be rigorously characterized. To obtain a DNA/ligand complex suitable for in vivo gene transfer, I have developed a multi-step protocol for the interaction of poly(L-lysine) with DNA in solution. I demonstrate here that the slow increase in the concentration of poly(L-lysine) in a vortexing solution of DNA results in the formation of DNA/ligand-poly(L-lysine) particles of defined size and shape. I hypothesize that binding of poly(L-lysine) to DNA initiates condensation by forming a nucleus of condensation along the DNA length. Further addition of poly(L-lysine) results in the formation of aggregated DNA/ligand-poly(L-lysine) complexes. The second step in the formation of stable condensed DNA particles consists in equilibrating the ionic strength of the solvent to facilitate the solubilization of the condensed DNA particles. Several biochemical parameters have been studied and correlated with the functional activity of DNA/ligand-poly(L-lysine) particles in transferring genes to the liver of adult animals by receptor-mediated endocytosis. Based on these findings, a model for the interaction between the DNA and the ligand/poly(L-lysine) is proposed and the implications for the effective transfer of genes into animals in vivo discussed. The model proposed indicates a role for the polycation in both the formation of suitable particles for gene transfer, and the efficient transfer of the DNA to the nucleus of cells
Bibliographical Information:

Advisor:

School:Case Western Reserve University

School Location:USA - Ohio

Source Type:Master's Thesis

Keywords:receptor mediated gene transfer

ISBN:

Date of Publication:01/01/1995

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