Targeted adenoviral delivery to the heart by magnetic microbubbles and highly focused ultrasound and imaging of gene transfer related processes
Final Report Abstract
Adenoviral gene transfer is still one oft he most effective and most intensively used methods for gene delivery. One of the major limitations in using adenoviral vectors is their immunogenicity and in in vivo gene transfer the predominantly targeting of liver cells (hepatocytes and Kupffer cells). To address both points we were able to device a methods of allowing for locally enhanced gene transfer to endothelial cells combined with shielding of vectors from interaction with neutralizing antibodies. This was achieved by using magnetic nanoparticels and additional electrostatic coating with the polymer P6YE5C. These nanoparticles were able to arrest the complexes under flow conditions as measured in arterioles and enhanced local gene transfer under guidance of a magnetic field and expression of reporter genes and vascular endothelial growth factor. Whereas the association with the magnetic nanoparticles already resulted in protection from neutralizing antibodies the addition of the polymer coat slightly enhanced further protection and still allowed gene transfer in the presence of magnetic nanoparticles and magnetic field. Most AdV-MNP complexes are large and rigid and in analogy to LV-MNP complexes might not allow for passage of the capillary system, but might lead to micro-vessel occlusion. Magnetic microbubbles (MMB) are assumed to be better suited for capillary passage. Therefore, during the second funding period we focused on establishing the magnetic microbubble-based technique combined with ultrasound for localized enhancement of gene transfer to the cardio-vascular system. The overall aim was to reduce scar formation in the infarcted heart by localized viral IL-10 production as IL-10 is known to reduce the release of pro-inflammatory cytokines and chemokines and thus to reduce macrophage infiltration. We successfully produced AdV MMB that lead to enhanced gene transfer in presence of magnetic field, which was further increased by disruption of the magnetically arrested MMB by ultrasound. After elimination of toxicity by means of adopting formulation and physical parameters this was finally successful under static as well as flow conditions for reporter gene expressing AdV as well as the antiinflammatory gene vIL expressing Ad. The physical and biological parameters of the complexes were determined and additionally it was demonstrated that the locally released vIL10 was finctional: The release of pro-inflammatory cytokines (TNFa, IL6 and IL1b) was reduced as the vIL10 expression was increased by magnetic field and application of ultrasound. It was also investigated, if the macrophages themselves were activated by the treatment, which was not observed. Furthermore the Tet-on system for regulation of IL-10 gene expression as well as for tissue-specific gene expression was successfully developed.
Publications
- (2010). Magnetic and Acoustically Active Lipospheres for Magnetically Targeted Nucleic Acid Delivery. Adv. Funct. Mater. 20, 3881-3894
Vlaskou, D., Mykhaylyk, O., Krotz, F., Hellwig, N., Renner, R., Schillinger, U., Gleich, B., Heidsieck, A., Schmitz, G., Hensel, K., and Plank, C.
(See online at https://doi.org/10.1002/adfm.200902388) - (2011). Magselectofection: an integrated method of nanomagnetic separation and genetic modification of target cells. Blood 117, e171-181
Sanchez-Antequera, Y., Mykhaylyk, O., van Til, N.P., Cengizeroglu, A., de Jong, J.H., Huston, M.W., Anton, M., Johnston, I.C., Pojda, Z., Wagemaker, G., and Plank, C.
(See online at https://doi.org/10.1182/blood-2010-08-302646) - 2012. Optimization of magnetic nanoparticles assisted lentiviral gene transfer. Pharm. Res. Pharm. Res. 29(5): 1255-1269
Christina Trueck, Katrin Zimmermann, Olga Mykhaylyk, Martina Anton, Sarah Vosen, Daniela Wenzel, Bernd K. Fleischmann, Alexander Pfeifer
(See online at https://doi.org/10.1007/s11095-011-0660-x) - 2012. Optimizing Adenoviral transduction of endothelial cells under flow conditions. Pharm. Res. 29(5): 1219-31
Martina Anton, Anja Wolf, Olga Mykhaylyk, Christian Koch, Bernd Gansbacher, Christian Plank
(See online at https://doi.org/10.1007/s11095-011-0631-2) - 2012. Silica-Iron Oxide Magnetic Nanoparticles Modified for Gene. Delivery; A Search for Optimum and Quantitative Criteria. Pharm. Res. 29(5): 1344-65
Olga Mykhaylyk, Titus Sobisch, Isabella Almstätter, Yolanda Sanchez-Antequera, Sabine Brandt, Martina Anton, Markus Döblinger, Dietmar Eberbeck, Marcus Settles, Rickmer Braren, Dietmar Lerche and Christian Plank
(See online at https://doi.org/10.1007/s11095-011-0661-9)
