The absorption enhancer sodium deoxycholate promotes high gene transfer in skeletal muscles.

Gene delivery to skeletal muscle is a promising strategy for the treatment of muscle disorders and for the systemic secretion of therapeutic proteins. In addition, muscle is an attractive target tissue because it is easily accessible. However, very few synthetic vectors proved capable of surpassing naked DNA mediated muscle gene transfer. In fact, only neutral copolymers, in particular poloxamers, demonstrated capacities to increase transgene expression in skeletal muscles. Here, we studied in vitro and in vivo behaviour of different bile salts. We report that sodium deoxycholate (DOC) and derivatives thereof increase after intramuscular injection by more than 100-fold the levels of the reporter gene luciferase compared to naked DNA. Using a LacZ expression cassette, we found that more than 20% of the muscle fibers expressed the reporter gene. Prolonged expression of a secreted reporter gene derived from a natural murine alkaline phosphatase enzyme could be documented. Altogether, our results demonstrate that bile salts belong to the most efficient chemicals identified so far for skeletal muscle gene transfer. Importantly, since these compounds are naturally found in the body, there is no risk of immune response against them and in addition several bile salts are already used in human medicine. Bile salt mediated muscle gene transfer may thus have broad applications in gene therapy.

The reverse block copolymer Pluronic 25R2 promotes DNA transfection of skeletal muscle.

Muscle is an important and attractive target for gene therapy. Recent findings have shown that neutral amphiphilic triblock copolymers with a PEO-PPO-PEO arrangement significantly increase muscle transfection as compared to naked DNA. We were interested in evaluating whether reverse Pluronics (PPO-PEO-PPO) also possess transfection properties. Therefore, we measured the in vitro and in vivo transfection activity of 25R2 and 25R4, two copolymers that differ by their hydrophilic/hydrophobic balance. The results show that 25R2 significantly increases the transfection level in muscle compared to naked DNA. Taken together, this work demonstrates that the reverse Pluronic 25R2 possesses interesting properties for in vivo transfection.

Evaluation of the muscle gene transfer activity of a series of amphiphilic triblock copolymers.

BACKGROUND: Amphiphilic triblock copolymers such as the polyethylene oxide-polypropylene oxide-polyethylene oxide L64 (PEO(13)-PPO(30)-PEO(13)) significantly increase transgene expression after injection of DNA/polymer mixtures into skeletal muscles. To better understand the way such copolymers act, we studied the behaviour of different poloxamers, including L64, both in vitro and in vivo. METHODS: The in vitro and in vivo transfection activity of five copolymers that differ either by their molecular weight or by their hydrophilic/hydrophobic balance was evaluated. Furthermore, we also studied the membrane permeabilizing properties of the poloxamers. RESULTS: The results obtained indicate that, after intramuscular administration of DNA/poloxamer formulations, all five compounds were able to significantly increase the expression levels of luciferase compared to an injection of naked DNA. Using a LacZ expression cassette, up to 30% of the muscle fibers expressed the reporter gene. Furthermore, we show that the effect can be obtained using different promoters. Finally, we document that, to some extent, all five poloxamers possess membrane permeabilizing properties. CONCLUSIONS: Taken together, the results obtained in the present study show that there is a large flexibility in terms of molecular weight and EO/PO ratio for obtaining increased levels of transgene expression in vivo.