Enabling Technologies for Cell-Based Clinical Translation Ferumoxytol Labeling of Human Neural Progenitor Cells for Diagnostic Cellular Tracking in the Porcine Spinal Cord With Magnetic Resonance Imaging JASON J. LAMANNA, a,b JUANMARCO GUTIERREZ, a LINDSEY N. URQUIA, a C. VICTOR HURTIG, a ELMAN AMADOR, b NATALIA GRIN, a CLIVE N. SVENDSEN, c THAIS FEDERICI, a JOHN N. OSHINSKI, b,d NICHOLAS M. BOULIS a,b Key Words. Cell transplantation x Clinical translation x In vivo tracking x Neural stem cell x Pig model x Stem cell transplantation x Magnetic resonance imaging x Spinal cord ABSTRACT We report on the diagnostic capability of magnetic resonance imaging (MRI)-based tracking of ferumoxytol-labeled human neural progenitor cells (hNPCs) transplanted into the porcine spinal cord. hNPCs prelabeled with two doses of ferumoxytol nanoparticles (hNPC-F Low and hNPC-F High ) were in- jected into the ventral horn of the spinal cord in healthy minipigs. Ferumoxytol-labeled grafts were tracked in vivo up to 105 days after transplantation with MRI. Injection accuracy was assessed in vivo at day 14 and was predictive of “on” or “off” target cell graft location assessed by histology. No dif- ference in long-term cell survival, assessed by quantitative stereology, was observed among hNPC- F Low , hNPC-F High , or control grafts. Histological iron colocalized with MRI signal and engrafted human nuclei. Furthermore, the ferumoxytol-labeled cells retained nanoparticles and function in vivo. This approach represents an important leap forward toward facilitating translation of cell-tracking tech- nologies to clinical trials by providing a method of assessing transplantation accuracy, delivered dose, and potentially cell survival. STEM CELLS TRANSLATIONAL MEDICINE 2016;5:1–12 SIGNIFICANCE This is the first report to document the diagnostic capabilities of iron oxide nanoparticle-labeled cells in the central nervous system of a large animal model. The major findings of this study were (a) the diagnostic capability of in vivo magnetic resonance imaging for evaluating ferumoxytol-labeled cell graft injection accuracy and initial volume; (b) the long-term survival of ferumoxytol-labeled cells in a large animal model; and (c) postmortem confirmation that ferumoxytol-labeled cells retain nanopar- ticles and function in vivo. The use of a U.S. Food and Drug Administration (FDA)-approved iron oxide nanoparticle, ferumoxytol, in combination with research-grade human neural progenitor cells, a large animal transplantation model, and a clinical magnetic resonance scanner make this study immedi- ately applicable to clinical investigation and informative to FDA Investigational New Drug enabling applications. INTRODUCTION Stem cell transplantation into the spinal cord presents a promising therapeutic strategy to overcome the regenerative limitations of the central nervous system (CNS) in degenerative and traumatic pathologies. Cell transplantation has been investigated clinically for amyotrophic lateral sclerosis (ALS), multiple sclerosis, and traumatic spinal cord injury [1]. Emerging evi- dence from these reports indicates that direct transplantation into the cord is safe, feasible, and well tolerated and may have therapeutic benefits [2, 3]. However, assessing therapeutic efficacy has been complicated by the inability of clinical investigators to measure transplanta- tion targeting accuracy, delivered dose, or cell survival because of ineffective postmortem his- tology and the lack of a diagnostic marker for identifying cell grafts. Calculating targeting accuracy for individual patients is important because only cell grafts de- livered “on” target will generate a therapeutic benefit (e.g., the ventral horn for ALS). Reliable targeting of the injection cannula to the ventral horn is based on preoperative imaging and visual observation of spinal cord anatomy but is compli- cated by spinal cord surface vasculature, its small size, and its relative depth in the cord. Knowledge of delivered cell dose and survival are also crucial. Departments of a Neurosurgery and d Radiology and Imaging Sciences, School of Medicine, Emory University, Atlanta, Georgia, USA; b Department of Biomedical Engineering, Georgia Institute of Technology and Emory University, Atlanta, Georgia, USA; c Board of Governors Regenerative Medicine Institute, Cedars-Sinai Medical Center, Los Angeles, California, USA Correspondence: Nicholas M. Boulis, M.D., 101 Woodruff Circle, Room 6339, Atlanta, Georgia 30322, USA. Telephone: 404-727-2247; E-Mail: nboulis@ emory.edu; or Jason J. Lamanna, Ph.D., 101 Woodruff Circle, Room 6339, Atlanta, Georgia 30322, USA. Telephone: 404-727-2247; E-Mail: jlamann@emory.edu Received January 4, 2016; accepted for publication July 11, 2016. ©AlphaMed Press 1066-5099/2016/$20.00/0 http://dx.doi.org/ 10.5966/sctm.2015-0422 STEM CELLS TRANSLATIONAL MEDICINE 2016;5:1–12 www.StemCellsTM.com ©AlphaMed Press 2016 ENABLING TECHNOLOGIES FOR CELL -BASED CLINICAL TRANSLATION This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. 2017;6:139–150 with Received January 4, 2016; accepted for publication July 11, 2016; published Online First on August 29, 2016. SIGNIFICANCE STATEMENT Correspondence:Nicholas M. Boulis, M.D., 101 Woodruff Circle, Room 6339, Atlanta, Georgia 30322, USA. Telephone: 404-727-2247; e-mail : nboulis@ emory.edu; or Jason J. Lamanna, Ph.D., 101 Woodruff Circle, Room 6339, Atlanta, Georgia 30322, USA. Telephone: 404-727-2247; e-mail :j lamann@emory.edu STEM CELLS TRANSLATIONAL MEDICINE 2017;6:139–150 www.StemCellsTM.com O c 2016 The Authors STEM CELLS TRANSLATIONAL MEDICINE published by Wiley Periodicals, Inc. on behalf of AlphaMed Press