Polycations and cationic lipids enhance adenovirus transduction and transgene expression in tumor cells

Replication-deficient adenovirus vectors are efficient vehicles for deliver ing therapeutic genes into mammalian cells. However, the high doses require d to produce effective gene transfer in vivo can also cause unwanted cellul ar toxicity. To improve replication-deficient adenovirus transgene expressi on while minimizing adverse reactions, we have tested polycationic compound s for their ability to enhance adenovirus adsorption. We demonstrate increa sed transgene expression after mixing adenovirus preparations with polycati ons, cationic lipids, and CaCl2 prior to transduction in vitro. An E1-delet ed adenovirus vector was admired with various polycations, and beta-galacto sidase (beta-gal) activity was evaluated. The optimal polycation concentrat ions for augmenting adenovirus-mediated gene transfer were 5-10 mu g/mL pol ybrene, 400 mu g/mL protamine sulfate, 10 mu g/mL N-(1-[2,3-dioleoyloxylpro pyl)-N,N,N-trimethylammonium methylsulfate (DOTAP), 2.5 mu g/mL Lipofectami ne, and 62.5 mM CaCl2. Polycations enhanced beta-gal expression in three of six established cell lines. Similar results were obtained using primary tu mor cell cultures, where beta-gal expression was increased 1.5- to 10.7-fol d (mean = 3.6) by polybrene, 1.8- to 7.5-fold (mean = 3.4) by DOTAP, and 2. 3- to 10.4-fold (mean = 4.8) by protamine sulfate. Adenovirus transduction efficiency in two primary leukemia isolates was improved by 3- and 4.5-fold . We were unable to demonstrate any benefit when adenovirus was admired wit h protamine sulfate prior to intratumoral injection in a xenogeneic severe combined immunodeficient mouse melanoma tumor model. Further studies will d etermine whether polycations can improve intratumoral gene transfer.