Nanomagnetic formulations for integrated nucleic acid delivery, cell labelling and imaging with a focus on mRNA transfection for cell reprogramming

• Antragsteller: Professor Dr. Christian Plank
• Fachliche Zuordnung: Pharmakologie
• Förderung: Förderung von 2009 bis 2016
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 40403621

This project will continue to provide the Research Unit Partners with comprehensively characterised magnetic nanoparticles (MNPs) for viral and nonviral nucleic acid delivery, magnetic cell labelling and manipulation, magnetic resonance imaging, relaxometry and the generation of magnetic microbubbles for magnetically targeted and ultrasound-activated nucleic acid delivery. In this context, our focus will be on understanding the nano-interactions involved in these processes to optimise the design of nanomagnetic composites including MNPs with immobilised affinity molecules and shielding of MNPs and magnetic delivery complexes. The specific focus of research project P7 is to establish an efficient and high yield procedure for mRNA magnetofection/magselectofection and to demonstrate the utility of the procedure for cell reprogramming with a focus on the generation of induced pluripotent stem cells (iPSs) in a DNA- and virus-free manner. This will be accomplished by introducing modified messenger RNAs encoding for the reprogramming factors into the cells using magnetofection and magselectofection, which are nanomagnetic nucleic acid delivery methods that we developed previously. First, we will develop optimised and well-characterised nanomagnetic formulations and protocols for delivering mRNA into the primary cells of interest, which will be established with the use of reporter constructs. Then, these methods will be translated for use with pluripotency factor mRNAs, which will involve a detailed analysis of timing and repeated dosing to obtain the optimal yield, viability and pluripotency of the derived cell populations. The latter part of the work will be carried out in the laboratory of Research Unit Partner ►Sasse and will result in a robust protocol for research and possible therapeutic applications. Partner ►Sasse will advance the optimized mRNA magnetofection protocols for the induction of cell differentiation towards cardiomyocytes. Collaborative accomplishments over the first funding period provide an excellent basis for achieving the new project goals. During the first funding period, a collection of optimised magnetic nanoparticles was tailor-made for highly efficient nonviral, lentiviral and adenoviral nucleic acid delivery. Optimised magnetic adenoviral formulations were used to boost the oncolytic potential of replicating adenovirus up to 50-fold. We also achieved high-dose magnetic cell labelling with low toxicity and with particles of high transverse r2* relaxivity for magnetic resonance imaging, allowing the detection of as few as 1,500 cells in a volume of 50 μl. Scalable magnetic nanoparticle synthesis was established in a system with automated process control. Furthermore, magnetic microbubbles with high magnetic and acoustic responsiveness were developed and successfully used for magnetically targeted and ultrasound-triggered nucleic acid delivery in vitro and in vivo, and this approach shows promising therapeutic results in a skin-flap model. Integrated magnetic cell separation and magnetofection, termed “magselectofection”, a method for genetically modifying cell lines and primary cells, including hematopoietic and mesenchymal stem cells, will serve as the technological basis for cell reprogramming within this Research Unit. Our work during the first funding period has resulted in 21 published project-related papers. Another six manuscripts are published online.

• DFG-Verfahren: Forschungsgruppen
• Teilprojekt zu: FOR 917: Targeting von Gen- und Zellbasierten Therapien mittels Magnetischer Nanopartikel
• Beteiligte Person: Privatdozent Dr. Carsten Rudolph