Mechanisms of gene transfer mediated by lipoplexes associated with targeting ligands or pH-sensitive peptides

Association of a targeting ligand such as transferrin, or an endosome disru pting peptide such as GALA, with cationic liposome-DNA complexes ('lipoplex es') results in a significant enhancement of transfection of several cell t ypes (Simoes S et al, Gene Therapy 1998; 5: 955-964). Although these strate gies can overcome some of the barriers to gene delivery by lipoplexes, the mechanisms by which they actually enhance transfection is not known. In stu dies designed to establish the targeting specificity of transferrin, we fou nd that apo-transferrin enhances transferrin enhances transfection to the s ame extent as transferrin, indicating that internalization of the lipoplexe s is mostly independent of transferrin receptors. These observations were r einforced by results obtained from competitive inhibition studies either by preincubating the cells with an excess of free ligand or with various 'rec eptor-blocking' lipoplexes. Transfection of cells in the presence of drugs that interfere with the endocytotic pathway provided additional insights in to the mechanisms of gene delivery by transferrin- or GALA-lipoplexes. Our results indicate that transferrin-lipoplexes deliver transgenes by endocyto sis primarily via a non-receptor-mediated mechanism, and that acidification of the endosomes is partially involved in this process.