Development of a new micro-injection-system for adenoviral gene delivery (MAGD)

Abstract

e22167 Background: Safe and effective delivery of viral vectors is important for gene therapy. Local application of such vectors is more efficient than systemic application. However, currently available injection systems are not optimal because placement of the injection needle and the vector application itself are generally not reproducibile. We developed a new injection system providing controlled injection of adenoviral vectors in tumors (xeno-transplants) in nude mice (NMRI nu/nu). Methods: The new MAGD consists of two fixation devices mounted on a 40x40cm plexi-glass plate, four injection units, and one pump unit with 4 infusion-pumps (Bee hive, Bioanalytical, Chesire, UK). Each injection unit is linked to a pump with commercially available plastic tube. Because the injection arms can be moved in x-,y- and z-direction, virus injection can easily performed for irregular tumor volumes. The entire injection volume is equally distributed to the 4 injection pumps allowing precise and steady injection rates between 0.1μl/min and 100μl/min. Success was confirmed with MRI-scans (Magnetom symphony, Siemens, Erlangen, Germany) 10 min after the injection. The efficacy of the MAGD was tested in 4 human squamous cell carcinoma cell lines from oro-/hypopharynx (FaDu, UD- SCC2, UD-SCC6, and UD-SCC7a). Virus transfection was performed with an adenovirus (serotype 5) expressing enhanced green fluorescent protein (eGFP). Transfection rates were quantified with flow cytometry. Successfully transfected cells expressed eGFP resulting in green fluorescence. In-vitro cell lines (concentration: 10 particles per cell, MOI 10) of FaDu, UD-SCC2, UD-SCC6 and UD-SCC7a served as controls. Results: The MAGD was easy to handle. Injection of 100μl (25μl per infusion pump) at an injection rate of 5μl/min took only 5 min. The in-vivo transfection rates achieved with the MAGD were 9±1% for FaDu, 39±2% for UD- SCC, 54±1% for UD-SCC6 and 54±9% for UD-SCC7a, respectively. The in-vitro transfection rates (controls) for these 4 cell lines were 2±1%, 23±4%, 29±4% and 3±2%, respectively. Conclusions: The new MAGD provided controlled injection of adenoviral vectors in tumors in nude mice. It proved to be effective, as it resulted in comparably high transfection rates. No significant financial relationships to disclose.