Gene Therapy Imaging Research Articles

Positron emission tomography reporter gene strategy for use in the central nervous system

There is a growing need for monitoring or imaging gene therapy in the central nervous system (CNS). This can be achieved with a positron emission tomography (PET) reporter gene strategy. Here we report the development of a PET reporter gene system using the PKM2 gene with its associated radiotracer [18F]DASA-23. The PKM2 reporter gene was delivered to the brains of mice by adeno-associated virus (AAV9) via stereotactic injection. Serial PET imaging was carried out over 8 wk to assess PKM2 expression. After 8 wk, the brains were excised for further mRNA and protein analysis. PET imaging at 8 wk post-AAV delivery showed an increase in [18F]DASA-23 brain uptake in the transduced site of mice injected with the AAV mice over all controls. We believe PKM2 shows great promise as a PET reporter gene and to date is the only example that can be used in all areas of the CNS without breaking the blood-brain barrier, to monitor gene and cell therapy.

Facile synthesis of carbon-11-labeled cholesterol-based cationic lipids as new potential PET probes for imaging of gene delivery in cancer

Gene therapy based on gene delivery is a promising strategy for the treatment of various human diseases such as cancer. Cationic lipids represent one of the important synthetic gene delivery systems. There is a great interest in imaging of gene therapy using the biomedical imaging technique positron emission tomography (PET). Carbon-11-labeled cholesterol-based cationic lipids were first designed and synthesized as new potential PET probes for imaging of gene delivery in cancer. The [ 11C-methyl]quaternary amine target tracers, N-[ 11C]methyl- N-[4-(cholest-5-en-3β-yloxycarbonyl)butyl]pyrrolidinium iodide ([ 11C] 4a), N-[ 11C]methyl- N′-[4-(cholest-5-en-3β-yloxycarbonyl)butyl]imidazolium iodide ([ 11C] 4b), N-[ 11C]methyl- N-[4-(cholest-5-en-3β-yloxycarbonyl)butyl]piperidinium iodide ([ 11C] 4c), N-[ 11C]methyl- N-[4-(cholest-5-en-3β-yloxycarbonyl)butyl]-4-methylpiperidinium iodide ([ 11C] 4d), and N-[ 11C]methyl- N-[4-(cholest-5-en-3β-yloxycarbonyl)butyl]morpholinium iodide ([ 11C] 4e), were prepared from their corresponding tertiary amine precursors with [ 11C]methyl iodide ([ 11C]CH 3I) through N-[ 11C]methylation and isolated by a simplified solid-phase extraction (SPE) method using a Silica Sep-Pak cartridge in 50–60% radiochemical yields decay corrected to end-of-bombardment (EOB), based on [ 11C]CO 2, and 111–185 GBq/μmol specific activity at the end of synthesis (EOS).

501. Novel Receptor-Targeted Nanoparticles for Delivery and Magnetic Resonance Imaging of Gene Therapy In Vivo

501. Novel Receptor-Targeted Nanoparticles for Delivery and Magnetic Resonance Imaging of Gene Therapy In Vivo

GENIS: Gene Expression of Sodium Iodide Symporter for Noninvasive Imaging of Gene Therapy (GT) in Human Prostate Carcinoma

GENIS: Gene Expression of Sodium Iodide Symporter for Noninvasive Imaging of Gene Therapy (GT) in Human Prostate Carcinoma

Gene expression and gene therapy imaging

The fast growing field of molecular imaging has achieved major advances in imaging gene expression, an important element of gene therapy. Gene expression imaging is based on specific probes or contrast agents that allow either direct or indirect spatio-temporal evaluation of gene expression. Direct evaluation is possible with, for example, contrast agents that bind directly to a specific target (e.g., receptor). Indirect evaluation may be achieved by using specific substrate probes for a target enzyme. The use of marker genes, also called reporter genes, is an essential element of MI approaches for gene expression in gene therapy. The marker gene may not have a therapeutic role itself, but by coupling the marker gene to a therapeutic gene, expression of the marker gene reports on the expression of the therapeutic gene. Nuclear medicine and optical approaches are highly sensitive (detection of probes in the picomolar range), whereas MRI and ultrasound imaging are less sensitive and require amplification techniques and/or accumulation of contrast agents in enlarged contrast particles. Recently developed MI techniques are particularly relevant for gene therapy. Amongst these are the possibility to track gene therapy vectors such as stem cells, and the techniques that allow spatiotemporal control of gene expression by non-invasive heating (with MRI guided focused ultrasound) and the use of temperature sensitive promoters.

Hyperthermia and gene therapy: Potential use of MicroPET imaging

In recent years, both hyperthermia and gene-therapy have been evaluated as approaches to improve cancer radiotherapy. In addition, potential exists to combine these approaches to increase the overall therapeutic efficacy. For example, it has been reported that adenovirus-mediated heat-inducible gene expression may reduce the normal tissue toxicity associated with constitutively controlled expression of therapeutic genes. In our laboratory, we have shown that adenovirus-mediated, heat-activated antisense Ku70 expression radiosensitizes tumor cells in vitro and in vivo, suggesting a novel approach to use heat-activated gene-radiotherapy to radiosensitize human tumors.However, to optimize the application of heat-activated gene-radiotherapy in the clinic, we need to develop techniques to improve the delivery of the therapeutic adenovirus and to verify/monitor the delivery non-invasively. In an ongoing study we test the effect of mild hyperthermia in improving adenovirus-medicated vector delivery in a mouse tumor model. In addition, we evaluate the use of non-invasive microPET imaging to monitor the spread of the adenoviral vector. Our preliminary results show that (1) microPET imaging can be used to monitor non-invasively the viral vector delivery and dissemination, and (2) mild heat shock leads to significantly improved viral vector distribution, in other words, a wider spatial spread, in vivo.Here, we will present a short review on the current status of hyperthermia and heat-activated gene-radiotherapy, and the potential use of PET imaging in gene therapy.

Gene therapy imaging in patients for oncological applications

Thus far, traditional methods for evaluating gene transfer and expression have been shown to be of limited value in the clinical arena. Consequently there is a real need to develop new methods that could be repeatedly and safely performed in patients for such purposes. Molecular imaging techniques for gene expression monitoring have been developed and successfully used in animal models, but their sensitivity and reproducibility need to be tested and validated in human studies. In this review, we present the current status of gene therapy-based anticancer strategies and show how molecular imaging, and more specifically radionuclide-based approaches, can be used in gene therapy procedures for oncological applications in humans. The basis of gene expression imaging is described and specific uses of these non-invasive procedures for gene therapy monitoring illustrated. Molecular imaging of transgene expression in humans and evaluation of response to gene-based therapeutic procedures are considered. The advantages of molecular imaging for whole-body monitoring of transgene expression as a way to permit measurement of important parameters in both target and non-target organs are also analyzed. The relevance of this technology for evaluation of the necessary vector dose and how it can be used to improve vector design are also examined. Finally, the advantages of designing a gene therapy-based clinical trial with imaging fully integrated from the very beginning are discussed and future perspectives for the development of these applications outlined.

In vivo imaging of gene therapy

In vivo imaging of gene therapy

Enzyme-targeted, nucleoside-based radiopharmaceuticals for scintigraphic monitoring of gene transfer and expression.

Enzymes, the expression products of transferred or native genes, offer unique windows of opportunity for clinical diagnosis and therapy. Although some expression products can be monitored in plasma, nuclear medicine imaging (SPECT and PET) offers the unique ability to selectively measure the intensity and regional/spatial distribution of gene expression both in vivo, in situ. Importantly, the superior sensitivity and moderate spatial resolution of the nuclear techniques also enable in vivo kinetic characterization of enzyme-substrate interaction. Indeed, the non-invasive, whole-body assessment of gene expression can only be achieved through imaging techniques. Given today's technology, nuclear imaging techniques uniquely provide the necessary sensitivity required to evaluate the success of the gene delivery and expression (transcription and translation), and to detect unwanted expression by non-target tissues. Enzymes are a special class of proteinacious gene expression products that selectively bind specific substrates for the purpose of molecular biotransformation rather than for signal transduction. In general, enzymes have received much less attention for imaging than receptors and antibodies, despite the enzymes' high substrate specificity and the potential for kinetic evaluation. Enzymes are attractive targets for diagnostic imaging and radioisotope radiotherapy because they convert multiple molecular copies of the substrate (radiotracer) per molecule of enzyme, thereby greatly increasing the ultimate sensitivity relative to the sensitivity offered by receptors that bind with 1:1 stoichiometry. Not surprisingly, enzymes have been the preferred molecular targets to date for scintigraphic imaging of gene therapy. This overview describes opportunities and advances in the utilization of radiolabelled nucleosides and nucleoside bases for imaging in gene therapy, with emphasis on the exploitation of enzyme systems for scintigraphic imaging of gene expression in gene therapy of cancer. Herpes simplex virus type-1 thymidine kinase and bacterial/fungal cytosine deaminase are discussed within the context of gene therapy issues such as gene vectors for targeting and delivery, the bystander effect, and radionucleoside delivery. The utilization of nucleosides as markers of tissue proliferation is discussed with respect to selected enzyme targets.

New approaches for imaging in gene therapy

Gene therapy is increasingly used experimentally and clinically to replace defective genes and/or impart new functions to cells and tissues. With the recent advances in vector design, improvements in transgene and prodrug activation strategies, gene therapy has been applied to a wide variety of diseases, tissues and organ systems. It is now clear that our specialty will play a critical role in gene therapy research and its clinical applications. Three aspects of gene therapy are of particular interest to imaging. The first is in delivering genes and vector products by minimally invasive interventional techniques. The second is in quantitating gene and DNA deliveries, for example, by nuclear imaging. Finally, imaging can be used to monitor the levels of transgene expression in vivo. A variety of imaging techniques including PET imaging, nuclear imaging, MR imaging and optical imaging can potentially be used to achieve the latter. This brief introductory overview is intended to summarize current strategies and illustrate the role that radiology will play in this field.

HSV-thymidine kinase: a molecular target for transfection imaging in gene therapy of cancer: L.I. Wiebe, E.E. Knaus and K.W. Morin. Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada T6G 2N8

HSV-thymidine kinase: a molecular target for transfection imaging in gene therapy of cancer: L.I. Wiebe, E.E. Knaus and K.W. Morin. Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada T6G 2N8