Magnetic nanoparticle mediated delivery of neurotherapeutic genes to multipotent neural stem cell transplant populations

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The adult brain and spinal cord (together called the central nervous system or CNS) repair poorly after injury and disease. This leads to limited recovery for patients with severe brain/spinal cord injury and disease with profound consequences for their quality of life. Stem cells found in the nervous system called neural stem cells or NSCs have enormous potential for increasing repair at injury/disease sites in the CNS as they can give rise to new stem cells, replace lost cell types of the CNS and can also suppress injury responses. These cells can migrate long distances in the CNS and are particularly attracted into areas of damage in the nervous system. Therefore, an important use for these engineered stem cells is to function as as transport 'vehicles' to deliver therapeutic molecules to injury/disease areas in the CNS. In experimental neurology research, the most common way of delivering genes to stem cells is to use modified viruses as vehicles. Infection of cells with these viruses enables the transfer of genes of interest into the infected cells. However, experimental methods using viruses can be be technically difficult, time consuming and expensive and can alter the basic cell properties of neural stem cells such as their division capability and their genesis of new cell types. This method also has other drawbacks including major safety issues (such as reactions from the immune system and cancer causing effects) and difficulties in achieving the large scale production that is required for human and veterinary clinical therapies. State-of-the-art delivery systems using small particles with an iron core called magnetic nanoparticles (MNPs) have many benefits in this regard and can be used effectively for delivery of genes. Preliminary clinical trials have shown that injecting MNPs into the body appears to be safe. MNPs can be linked with genes and taken up into cells to mediate gene delivery. Although MNP based systems are thought to be less effective than viruses for delivering genes, we have shown that applying static or oscillating magnetic fields (a method called 'magnetofection') can dramatically increase gene delivery by MNPs. DMC's group proved recently that MNPs can be used to safely deliver genes to several major cell types (of both rodent and canine origin) that are used in neural cell transplantation therapies. The methods used were not associated with safety issues and did not alter basic stem cell properties like their division or migration capability or the types/numbers of daughter cells they give rise to. The methods we have developed for MNP based gene transfer are very safe, technically simple, quick and relatively inexpensive, therefore their use will provide an effective and convenient method for delivering genes and can result in significant cost savings to laboratories and funding agencies.This study will build on recent findings to develop methods to use MNPs to deliver genes coding for therapeutic molecules to stem cells (that will then be used for transplantation). We will assess different strategies to optimise uptake of MNPs coated with genes into stem cells and establish if this procedure has adverse effects on the survival and development of the cells. A further goal is to develop a new, technically easy and cheap method to examine the interactions of stem cells and MNPs at high magnification using a special 'electron' microscope. We will also investigate the underlying mechanisms by which oscillating magnetic fields increase gene delivery by MNPs. The repair enhancing potential of the engineered stem cells will be tested in a 'living' 3D slice model of the spinal cord, that can function well as injured 'host' tissue for transplant cells. This method provides a robust alternative to the use of surgical transplantation to study stem cell transplantation therapies in living animals, thereby significantly reducing animal usage and suffering in experimental research.