PET-based reporter gene imaging was developed approximately 12 years ago, and the technology is beginning to be translated into specific clinical applications. Although viral genes were initially used in radiotracer-based reporter systems (eg, herpes simplex thymidine kinase, HSV1-tk), human reporter genes increasingly will play a more important role in the future.
The first patient applications have already been reported, where imaging was used to track and monitor “suicide” gene therapy (see Figure 1). These vector-tracking studies were able to take advantage of the fact that the therapeutic gene (HSV1-tk or HSV-sr39tk) also serves as a reporter gene. In the years to come, it is likely that human reporter genes will begin to supplement and may even replace HSV1-tk or HSV1-sr39tk as the optimal constitutively-expressed vector-tracking genes. However, one distinct advantage of HSV1-tk and HSV1-sr39tk is their ability to function as a suicide gene as well as a reporter gene. This dual reporter-suicide feature provides “added value” to gene therapy protocols; it adds an important vector-safety feature as well as a vector-monitoring feature.
Three classes of human reporter genes have been developed and were recently compared; they include receptors, transporters and enzymes. An example of highly expressed cell membrane receptors include the somatostatin family of receptors (hSSTRs). The transporter group includes the sodium-iodide symporter (hNIS), and the norepinephrine transporter (hNET). The endogenous enzyme classification includes human mitochochodrial thymidine kinase 2 (hTK2).
The initial applications of reporter gene imaging in patients will focus on tracking studies using reporters that are constitutively expressed. These studies will involve at least two different clinical disciplines: a) gene therapy and b) adoptive cell-based therapies. They will benefit from the availability of efficient human reporter systems that can provide critical monitoring information for adeno-, retro-, and lenteviral-based gene therapy, oncolytic bacterial and viral therapy, and adoptive cell-based therapies.
Figure 1. HSV1-tk reporter gene imaging in patients after Liposome-HSV-1-tk-complex transduction (Panel A). Coregistration of [124I]FIAU-PET and MRI before (left column) and after (right column) HSV1-tk vector injection. A region of specific [124I]FIAU retention (at 68 h) within the tumor is visualized (white arrow). This tumor region showed signs of necrosis (cross hairs, right column after ganciclovir treatment).
Figure adapted from Jacobs et al. (2001b)
Adenoviral transgene (HSV1-tksr39) expression in patients with liver cancer (Panel B). Coronal PET images 1.5 and 6.5 hours after injection of [18F]-FHBG (48 h after 2 x1012 Adv-tk). Localization of [18F]-FHBG in the treated lesion was variable in the early images, but could be seen at 6.5 hours in all patients (see arrows).
Figure adapted from Penuelas, et al. (2005)
The translational applications of noninvasive in vivo reporter gene imaging are likely to include: (i) quantitative monitoring of the gene therapy vector and transduction efficacy in clinical protocols by imaging the location, extent and duration of transgene expression; (ii) monitoring cell trafficking, targeting, replication and activation in adoptive T cell and stem/progenitor cell therapies; (iii) assessments of endogenous molecular events using different inducible reporter gene imaging systems.
The second application of reporter genes imaging that is likely to be translated into clinical studies involves transduction vectors that contain at least two different reporter constructs. One will be a “constitutive” reporter that will be used to identify the site, extent and duration of vector delivery, and can be used to monitor the efficiency of tissue transduction (the normalizing term) for subsequent image and data analysis.
A second reporter will be “inducible” and sensitive to endogenous transcription factors, as well as to post-transcriptional processing, modulation of reporter protein translation, protein-protein interactions, and reporter protein ubiquitination. The “inducible” reporters will act as “sensors” and will be used to monitor the functional status and characteristics of the transduced cells. It is now recognized that the readout from “inducible” reporters requires coupling with the readout from a “constitutive” reporter in order to appropriately interpret the data.
We are optimistic. The tools and resources largely exist and we should be able to perform more gene imaging studies in patients in the near future. The advantages and benefits of noninvasive imaging to monitor transgene expression in gene therapy protocols are obvious. The ability to visualize transcriptional and post-transcriptional regulation of endogenous target gene expression, as well as specific intracellular protein-protein interactions in patients, will provide the opportunity for new experimental venues in patients. They include the potential to image the malignant phenotype of an individual patient’s tumor at a molecular level and to monitor changes in the phenotype over time, as well as to image a drug’s effect on a specific signal transduction pathway in an individual patient’s tumor.
For more information:
- Ronald Blasberg, MD, is a Member in the Departments of Neurology and Radiology, and in the Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center, New York.
- Jacobs A, Voges J, Reszka R, et al. Positron-emission tomography of vector-mediated gene expression in gene therapy for gliomas. Lancet. 2001;358:727-729.
- Penuelas I, Mazzolini G, Boan JF, et al. Positron emission tomography imaging of adenoviral-mediated transgene expression in liver cancer patients. Gastroenterology. 2005;128:1787-1795.
- Serganova I, Ponomarev V, Blasberg R. Human Reporter Genes: Potential Use in Clinical Studies. Nucl Med Biol. 2007;34:791-807.