Current Pharmaceutical Biotechnology - Volume 5, Issue 6, 2004

Molecular and cellular imaging is a relatively young field that is rapidly changing our approach towards understanding and solving problems in biology and medicine. Among the different imaging modalities, MR imaging offers both whole body penetration and high spatial and temporal resolution. Now that the hardware necessary to perform MR imaging in sufficient detail can be found in many places across the world, it is anticipated that future advances in molecular and cellular imaging will heavily depend on further development of MR probes and contrast agents. With this in mind, this special issue of Current Pharmaceutical Biotechnology offers a sampling of the current state-of-the-art of MR contrast agents that can be used to tag molecules and cells, as well as probe the biochemical microenvironment. The article by Artemov et al. describes different strategies that can be followed to target contrast agents to cell surface receptors. Lanza et al. report on the use of paramagnetic nanoparticles that can be derivatized for specific molecular imaging as well as drug delivery. Next is the review by Aime et al. on intracellular magnetic labeling using paramagnetic chelates and different uptake mechanisms. Lowe then follows and outlines the current status of so-called smart or activated contrast agents that can respond to changes in the microenvironment. Koretsky reviews the new emerging application of manganese to trace neuronal connextions in the brain. Kobayashi et al. are using dendrimers as macromolecular contrast agents and describe their various applications in MR imaging. Ho et al. employ superparamagnetic iron oxides to label macrophages in vivo for detection of these cells in various models of transplanted organ rejection. Finally, a contribution from our own Institute on the use of magnetic nanoparticles to monitor (stem) cell therapy closes this issue. I hope you will enjoy this issue as much as I did editing it.

Over the past decade MR (magnetic resonance) imaging has emerged as one of the major modalities for noninvasive functional imaging. Recent advances in the development of targeted MR contrast agents have added significantly to the capabilities of MR imaging. In particular, the use of targeted contrast agents to report on the expression of cell surface receptors, combined with the functional capabilities of MR imaging, together provide unique opportunities to understand receptor-mediated pathways. In this article we have reviewed current MRI strategies used to visualize receptor expression, the potential advantages and drawbacks of these strategies, and novel areas of focus for the future.

Molecular biology and genomic sciences are revealing the early biological signatures for many diseases. In response, the Molecular Imaging community is rapidly developing contrast agents to visualize the nascent pathological changes and to concomitantly deliver treatment directly to the site of disease. The evaluation, development and use of these new agents require a complementary understanding of contrast chemistry and imaging techniques. The fundamental issues surrounding magnetic contrast agent development, rational drug delivery, MR molecular imaging, and their interdependence are elucidated.

The low sensitivity is the major disadvantage of MRI as compared to PET. Therefore, amplification strategies are necessary for specific pathway labeling. This survey is aimed at exploring different routes to the entrapment of Gd(III) chelates in various type of cells at amounts sufficiently large to allow MRI visualization. Namely, the obtained results have been summarized in terms of internalization via i) pinocytosis; ii) phagocytosis; iii) receptors; iv) receptor mediated endocytosis; v) transporters; vi) transmembrane carrier peptides. MRI visualization of cells appears possible when the number of internalized Gd(III) chelates is of the order of 10 7- 10 8/cell. Pinocytosis shows to be particularly useful for labeling cells that can be incubated for several hours in the presence of high concentrations of Gd-agent. This approach appears very effective for labeling stem cells. Nanoparticles filled with Gd-chelates can be used for an efficient loading of cells endowed with a good phagocytic activity. Entrapment via receptors most often results in receptor mediated endocytosis. Suitably functionalized monomeric and multimeric Gdchelates can be considered for being internalized by this route as well as supramolecular systems such as those formed between Avidin and biotinylated Gd-complexes. Exploitation of up-regulated transporters of nutrients in tumor cells appears to be a promising route for their differentiation from healthy cells. Finally, properly designed systems entering the cells by means of penetrin-like peptides deserve great attention.

The relative non-specificity of the first generation MR contrast agents has meant that a new approach to their design is required. This review focuses on a new class of more specific or functional agents. These are the so-called “activated”, “smart” or “responsive” contrast agents. The relaxivity of an activated contrast agent is responsive to (or can be modulated by) a particular in vivo stimulus such as a change in biological environment or activity. More specifically, a “switching on” of contrast in response to an event such as a change in physiological pH, metal ion concentration, enzyme activity or partial pressure of oxygen is sought. The current generation of activated MR contrast agents is discussed herein.

Manganese is an essential metal that participates as a co-factor in a number of critical biological functions such as electron transport, detoxification of free radicals, and synthesis of neurotransmitters. Like other heavy metals, high concentrations of manganese are toxic. For example, chronic overexposure to manganese leads to movement disorders. In order to maintain this balance between being an essential participant in enzyme function and being a toxic heavy metal, a rich biology has evolved to transport and store manganese. Paramagnetic forms of manganese ions are potent MRI relaxation agents. Indeed, Mn2+ was the first contrast agent proposed for use in MRI. Recently, there is renewed interest in combining the strong MRI relaxation effects of Mn2+ with its unique biology in order to expand the range of information that can be measured by MRI. Manganese Enhanced MRI is being developed to give unique tissue contrast, assess tissue viability, act as a surrogate marker of calcium influx into cells and trace neuronal connections. In this article we review recent work and point out prospects for the future uses of manganese enhanced MRI.

Paramagnetic metals can induce T1 shortening by interaction with free water molecules. Two metal ions, Gadolinium and Manganese, are currently available for human use. Gadolinium-based MRI contrast agents (CAs) can operate using a ∼100-fold lower concentration of Gadolinium ions in comparison to the necessary concentration of Iodine atoms employed in CT imaging in the tissues. Therefore, numerous macromolecular MRI CAs prepared employing relatively simple chemistry are readily available that can provide sufficient enhancement for multiple applications. Herein, we describe the synthesis, characteristics, and potential applications of dendrimer-based macromolecular MRI CAs in our recently reported libraries. This entire series of dendrimer-based macromolecular MRI CAs have a spherical shape and possess similar surface charges. Changes in molecular size altered the route of excretion. Smaller sized contrast agents, of less than 60 kD molecular weight, were excreted through the kidney resulting in these agents being potentially suitable as functional renal contrast agents. Less hydrophilic and larger sized contrast agents were found better suited for use as blood pool contrast agents. Hydrophobic variants of CAs formed with polypropylenimine diaminobutane dendrimer cores quickly accumulated in the liver and can function as liver contrast agents. Larger hydrophilic agents are also useful for lymphatic imaging. Finally, contrast agents conjugated with either monoclonal antibodies or with avidin are able to function as tumor-specific contrast agents and might also be employed as therapeutic drugs for either gadolinium neutron capture therapy or in conjunction with radioimmunotherapy.

Organ transplantation is the generally preferred medical procedure of treatment for patients with end-stage organ failure. The immunological reaction of rejection is a major cause of functional failure in transplant patients. The current “gold standard” for detecting or confirming graft rejection following solid organ transplantation requires biopsy samples in order to detect immune cell (e.g., T-cells, macrophages, etc.) infiltration into the graft and other pathological changes. This procedure is not only invasive, having associated risks, but is also prone to sampling errors that can yield false negative results. To circumvent the need for biopsies, we are developing magnetic resonance imaging (MRI) techniques to monitor the accumulation of immune cells at the transplanted organ as a means to detect graft rejection. By labeling immune cells with an MRI contrast agent, dextran-coated ultrasmall superparamagnetic iron oxide (USPIO) particles, we can monitor the accumulation of these labeled immune cells at the rejecting graft as a non-invasive method to detect graft rejection. Cells can be labeled ex vivo and then infused into the animal, or MRI contrast agents can be introduced directly into the animal in vivo. Our results show excellent correlation among the MRI signal intensity due to the USPIO-labeled macrophages at the rejecting graft, immuno-staining for macrophages, histo-pathology for graft rejection, and the iron staining of tissue samples. In this article, we shall give a summary of our progress from detecting single immune cells in vitro to monitoring the accumulation of immune cells in vivo at the transplanted kidneys, hearts, and lungs in our rat models for organ transplantation by MRI.

Given the remarkable progress that has recently been obtained in animal studies, the clinical use of stem and progenitor cells to correct or replace defective cell populations may soon become a reality. In order to develop effective cell therapies, the location and distribution of these cells must be determined in a non-invasive manner. Magnetic resonance (MR) tracking of magnetically labeled cells following transplantation or transfusion may fulfill this requirement. Indeed, a series of recent studies indicate that MRI cell tracking has great potential for further evaluation and optimization of cell therapy. Due to its biocompatibility and strong effects on T2(*) relaxation, iron oxide nanoparticles appear to be the contrast agent of choice, and several methods now exist to shuttle sufficient amount of these compounds into cells. Most of the tracking work has been carried out in disease models of the central nervous system, but, recently, the infarcted heart has also received attention. With its excellent spatial resolution and the ability to track labeled cells over prolonged periods of time, MR monitoring of cell therapy is likely to become an important technique in the foreseeable future.