Iron Labeling and Pre-Clinical MRI Visualization of Therapeutic Human Neural Stem Cells in a Murine Glioma Model

Mya S Thu,Margarita Gutova,Elizabeth Garcia,Karen S Aboody,Marianne Z Metz,Rex A Moats,Seung U Kim,Stephen E Kendall,Richard T Frank,Joseph Najbauer,Ira Harutyunyan,Nicole Sangalang,Pedro R Lowenstein

doi:10.1371/journal.pone.0007218

Abstract

BackgroundTreatment strategies for the highly invasive brain tumor, glioblastoma multiforme, require that cells which have invaded into the surrounding brain be specifically targeted. The inherent tumor-tropism of neural stem cells (NSCs) to primary and invasive tumor foci can be exploited to deliver therapeutics to invasive brain tumor cells in humans. Use of the strategy of converting prodrug to drug via therapeutic transgenes delivered by immortalized therapeutic NSC lines have shown efficacy in animal models. Thus therapeutic NSCs are being proposed for use in human brain tumor clinical trials. In the context of NSC-based therapies, MRI can be used both to non-invasively follow dynamic spatio-temporal patterns of the NSC tumor targeting allowing for the optimization of treatment strategies and to assess efficacy of the therapy. Iron-labeling of cells allows their presence to be visualized and tracked by MRI. Thus we aimed to iron-label therapeutic NSCs without affecting their cellular physiology using a method likely to gain United States Federal Drug Administration (FDA) approval.MethodologyFor human use, the characteristics of therapeutic Neural Stem Cells must be clearly defined with any pertubation to the cell including iron labeling requiring reanalysis of cellular physiology. Here, we studied the effect of iron-loading of the therapeutic NSCs, with ferumoxide-protamine sulfate complex (FE-Pro) on viability, proliferation, migratory properties and transgene expression, when compared to non-labeled cells. FE-Pro labeled NSCs were imaged by MRI at tumor sites, after intracranial administration into the hemisphere contralateral to the tumor, in an orthotopic human glioma xenograft mouse model.ConclusionFE-Pro labeled NSCs retain their proliferative status, tumor tropism, and maintain stem cell character, while allowing in vivo cellular MRI tracking at 7 Tesla, to monitor their real-time migration and distribution at brain tumor sites. Of significance, this work directly supports the use of FE-Pro-labeled NSCs for real-time tracking in the clinical trial under development: “A Pilot Feasibility Study of Oral 5-Fluorocytosine and Genetically modified Neural Stem Cells Expressing Escherichia coli Cytosine Deaminase for Treatment of Recurrent High-Grade Gliomas”.

Highlights

Glioblastoma multiforme (GBM) is the most common primary malignant brain tumor in adults, with a mean survival of less than one year following diagnosis, despite advances in surgical, radiation and chemotherapeutic approaches [1,2,3]
This work directly supports the use of ferumoxide-protamine sulfate complex (FE-Protamine Sulfate (Pro))-labeled neural stem cells (NSCs) for real-time tracking in the clinical trial under development: ‘‘A Pilot Feasibility Study of Oral 5-Fluorocytosine and Genetically modified Neural Stem Cells Expressing Escherichia coli Cytosine Deaminase for Treatment of Recurrent High-Grade Gliomas’’
Adherent NSCs in a 96-well culture plate were labeled at various concentrations of FE-Pro complex (FE at 50, 100, 200 and 250 mg/ml, each of which was complexed with increasing Pro at 3, 10, and 25 mg/ml)

Summary

Introduction

Glioblastoma multiforme (GBM) is the most common primary malignant brain tumor in adults, with a mean survival of less than one year following diagnosis, despite advances in surgical, radiation and chemotherapeutic approaches [1,2,3]. New treatment modalities, including immunotherapy, gene therapy and drug delivery across the blood-brain barrier (BBB), have yet to achieve significant clinical success due to shortcomings in effectively targeting these invasive tumor cells, while minimizing toxicity to normal tissue [2]. Neural stem cells (NSCs) have inherent tumor-tropic properties to primary and invasive tumor foci, and offer much promise as cellular delivery vehicles to effectively target therapeutic gene products to these invasive glioma cells [4,5,6]. This provides the basis for developing novel NSC-mediated treatment approaches to localize therapeutic gene products to malignant primary and invasive tumor foci. We aimed to iron-label therapeutic NSCs without affecting their cellular physiology using a method likely to gain United States Federal Drug Administration (FDA) approval

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