Pre-clinical evaluation of Magnetically labeled Cells for Cellular MRI

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Previously, we demonstrated that by combining ferumoxytol (F) with protamine (P) and heparin (H) resulted in a self assembling nanocomplex that could be used to label cells. Transmission electron microscopy of HPF revealed that these nanocomplexes were spheroid shaped with the HP in center surrounded by F. Incubating stem cells and T-cells in HFP nanocomplexes in serum free media for 2 hours followed by complete media resulted in cell labeling. HPF labeling did not impair the cells viability, proliferative capacity, apoptotic rate, activation, phenotypic surface marker expression, or capacity to differentiate. MRI at 3T of HPF labeled BMSC implanted in the rat brain demonstrated the ability to detect 1000 cells with a 50% decrease in T2* in the rat brain compared to the surrounding parenchyma. Pre-clinical safety/toxicity studies of intracerebrally administrated HPF-labeled NSCs in mice were also performed, and demonstrated no significant clinical or behavioral changes, no neuronal or systemic toxicities, and no abnormal accumulation of iron in the liver or spleen. The HPF labeling technique has been scaled up, and the NIH Cell Processing Section cGMP facility was able to label BMSCs in biofactories with no changes in BMSC function or viability of the labeled cryopreserved cell product. The HPF method was also used to label genetically engineered neural stem cells (NSC) that express cytosine deaminase as part of an ongoing clinical trial to treat patients with recurrent glioblastoma. The HPF labeled NSC are to be directly implanted deep into brain around the periphery of the surgical resection sight in order that they can migrate to areas of residual tumors or satellite metastasis. MRI monitored the migration of the HPF labeled NSC in the brain over 30 days. Initial studies performed at City of Hope report that there was essential no adverse events following the implantation of labeled cells. We have recently optimized the HPF labeling approach by changing the order that the three drugs are added to cells to FHP resulted in significant increase in the intracellular iron content in BMSC and NSC as compared to HPF labeled cells. We also report on the physicochemical characteristics for optimizing the H, P, and F components in different ratios, and mixing sequences, producing NCs that varied in hydrodynamic size. NC size depended on the order in which drugs were mixed in media. Electron microscopy of HPF or FHP showed that F was located on the surface of spheroidal shaped HP complexes. Human stem cells incubated with FHP NCs resulted in a significantly greater iron concentration per cell compared to that found in HPF NCs with the same concentration of F. These results indicate that FHP could be useful for labeling stem cells in translational studies in the clinic. Long-term neurological deficits due to immature cortical development are emerging as a major challenge in congenital heart disease (CHD). By injection superparamagnetic iron oxide nanoparticles into the lateral ventricles of hypoxic exposed of the gyrencephalic piglet brain a model of CHD we were able to label neural stem cells in vivo and track their pattern of distribution into the cortex and olfactory bulbs by ex vivo MRI. The results showed that hypoxia reduces proliferation and neurogenesis in the subventricular zone (SVZ), which is accompanied by reduced cortical growth. The findings demonstrated that SVZ neuronal stem cells (NSC) contribute to perinatal corticogenesis and suggest that restoration of SVZ NSCs' neurogenic potential is a candidate therapeutic target for improving cortical growth in CHD.