University of Pennsylvania · Department of Bioengineering
The broad goal of our research program is to develop novel imaging agents that can be used to elucidate the molecular mechanisms of disease in living cells and/or subjects. We are developing agents that allow us to monitor a wide range of biological processes including gene regulation, RNA localization, protein expression, and enzymatic activity. We believe that molecular imaging will provide a major advancement over most current diagnostic imaging approaches since, in many cases, malignancies can be detected by an altered molecular profile well before any visual signs of disease. Advancement in the design of imaging agents could also have many far-reaching implications in the development of pharmaceuticals by facilitating the high-throughput screening of various compounds and by simplifying the evaluation of various therapeutics in living subjects (i.e. allow for quantification of efficacy and dosing). A broad spectrum of disciplines are combined in our lab to bring ideas for molecular probes from the initial chemical or biological synthesis, to the in vitro validation and evaluation, and in many cases to the imaging in living subjects. Further, a variety of imaging modalities are used including fluorescence, magnetic resonance, and bioluminescence. Several of the major thrusts in our lab are described here.
Contrast-Enhanced Magnetic Resonance Imaging and Drug Delivery with Targeted Nanoparticles
The non-invasive imaging of cancer biomarkers in living subjects could provide a powerful technique for locating metastatic disease, staging tumors, evaluating the availability of therapeutic targets, and monitoring the efficacy of treatment. Magnetic resonance (MR) imaging is a particularly attractive platform for such molecular imaging applications due to its ability to acquire high-resolution anatomical images in conjunction with measures of biomarker expression. However, a major obstacle faced by MR is overcoming the relatively low sensitivity of targeted MR contrast agents. In general, the number of cell receptors at a disease site is too low to recruit enough MR contrast agents to generate sufficient contrast. Therefore, there remains a need to develop new imaging agents capable of generating higher contrast and/or novel amplification strategies that will result in improved targeting. Our lab is pursuing both of these avenues. Specifically, we are developing new formulations of both iron oxide- and gadolinium-based nanoparticles, to improve the contrast-enhancing capabilities per nanoparticle. In parallel, we are developing new amplification schemes that allow for an improvement in the accumulation of contrast agents at the target site.
In addition to our projects aimed at imaging disease sites in vivo, we are also using our nanoparticle carriers as drug delivery devices. We believe that the same techniques that we are using to improve the targeting of imaging agents will also benefit the delivery of therapeutic compounds.
Absolute Quantification of RNA Expression
Variations in gene expression are commonly considered the major determinants for dictating cell behavior. Accordingly, methods to measure gene expression, such as reverse-transcriptase (RT) PCR and DNA microarrays, have proven to be invaluable in regards to understanding cell regulatory processes and disease mechanisms. However, these methods generally provide only the relative change in gene expression for a population of cells and not an absolute measure of RNA copies at the single cell level. Under many circumstances it is the aberrant behavior of only a few cells or the stochasticity of RNA expression within a population that leads to disease evolution. Currently, we are developing optical imaging probes that are capable of the real-time visualization and absolute quantification of gene expression in single living cells. We envision that the development of an optical probe that can provide a more complete profile of gene expression, with spatial and temporal resolution in single living cells, will lead to significant advancements in molecular medicine, clinical diagnosis and biotechnology, and will also facilitate our understanding of human health and disease.
Monitoring Intracellular Pathways with Bioluminescent Reporters
In healthy individuals, homeostasis is maintained through a balance between cell proliferation and cell death. Therefore, it is not surprising that defects in the regulation of cell proliferation and/or cell death have been associated with a number of human diseases, including cancer, viral infections, autoimmune diseases, and neurodegenerative disorders. It is envisioned that the ability to non-invasively image these processes in living subjects would not only provide insight into diverse disease mechanisms but would also facilitate the development of drugs to treat these and other diseases, and allow one to monitor the effectiveness of therapy. Bioluminescent imaging is a particularly attractive platform for such molecular imaging applications due to its simplicity, sensitivity, the ability to obtain temporal information, and the potential to conduct high-throughput screening. Currently, we are genetically engineering bioluminescent probes that will allow us to non-invasively image programmed cell death and cell proliferations in living subjects.
Diagnostic Assays Designed Around Magnetic Switches
An interesting characteristic of superparamagnetic iron oxide nanoparticles (SPIO) is that they become more effective MR contrast agents when self-assembled into larger complexes. This mechanism of contrast enhancement during SPIO self-assembly (or vice versa during SPIO disassembly) is generally referred to as magnetic relaxation switching (MRSW) and can be detected by MR imaging. Currently, we are trying to exploit MRSW as a tool for detecting biomolecules in homogeneous assays. Specifically, ligands targeted by iron oxide nanoparticles (e.g. oligonucleotides, enzymes, proteins, viruses, etc.) are used to trigger SPIO self-assembly (or disassembly). The presence of these ligands can then be easily detected by a change in the MR signal intensity. A unique advantage of MRSW over many alternative detection technologies is that changes in MR signal can easily be measured in turbid media, whole-cell lysates, and potentially in vivo.