Research Interests
Research in the Rao lab uses physical, chemical and biological tools to develop novel imaging strategies. We develop new molecular probes to monitor specific biological targets under physiological settings, and in general our projects fit into one or more of the following three interconnected lines.
1. Enzyme activity-based in vivo imaging
Many receptors and enzymes play crucial roles in cellular and physiological functions. While receptors generally function by binding ligands, enzymes catalyze biochemical transformations. Employing a common strategy for imaging receptors, enzymes may also be imaged by binding labeled and irreversible inhibitors. This approach does not take advantage of an enzyme's biological function, catalysis. Enzyme activity-based approaches rely upon and benefit from enzymatic function, creating signal amplification and producing high sensitivity with little perturbation to the biological functions of targeted enzymes. We have long-standing interests in imaging real-time enzyme activity in vivo in caspases, MMP, ribozymes, and beta-lactamases. The protease on which we currently focus is granzyme B.
Granzyme B
Granzyme B has been implicated in a host of immune diseases, and is important in immunotherapies of cancer. We develop sensors and probes for in vitro detection and in vivo imaging of granzyme B activity. Publications in this area use our TESLA (target-enabled in-situ ligand aggregation) strategy to image the granzyme activity in antigen receptor T-cells and in checkpoint blockade therapies (ACS Cent. Sci., 2022). We have also used a bioluminescence turn-on probe to image granzyme B activity in responder- and nonresponder populations towards checkpoint blockade therapy (Cell Chem. Biol., 2022).
2. Pathogen detection strategies
Detection of bacteria-based pathogens is a difficult undertaking that requires exquisite sensitivity and specificity. Not only do the strains of pathogenic bacteria need to be correctly identified, but strains of non-pathogenic bacteria must be ignored. The utility of diagnostic tests is often limited by stability in storage, price considerations, sensitivity, and time-to-result. Taking these factors into consideration, the development of point-of-care detection protocols is an important step in eradicating diseases.
Tuberculosis
Tuberculosis infects more than 10 million people per year with 1.5 million deaths worldwide, making it the deadliest infectious disease in the world. The standard of care for tuberculosis patients relies on a cocktail of antibiotics to kill the bacteria. However, due to inadequate diagnostic capabilities in developing countries, many asymptomatic patients do not receive timely treatment and eventually develop fatal infections. We are developing a tuberculosis detection strategy specifically for use in resource-poor communities. This means using cheap and readily available materials in the detection procotol while employing stable and facile detection kit components. We hope this research will be directly translatable to the communities most in need.
3. Metals in biology and therapeutics
Metal ions are fundamental elements for the maintenance of life. Their absence can cause growth disorders, severe malfunction, carcinogenesis, or death. They are essential in several structural and functional roles: intra- and intercellular communications, maintaining electrical charges and osmotic pressure, electron transfer processes, in the regulation of DNA transcription, among others. Currently, the detection of metals relies on chelators bound to reporters. The localization and activity of metals has been studied by fluorescence microscopy, ICP-MS, EPR spectroscopy, and other ground-breaking instrumentation. We aim to understand the role of copper in diseases such as cancer.
Copper
Copper is an essential metal in respiration, as it is a cofactor in the active site of enzymes like cytochrome oxidase C and superoxide dismutase. The bioinorganic community understands that there is almost no labile copper ions in cells. When this homeostatis is disturbed, the potential for disease increases dramatically. The disregulation of copper in cancers is also an active area of research. We are investigating the complexation of copper in cancer models by designing new chelator-reporters. We have previously used a copper-complexing molecule to treat triple-negative breast cancer in mice (Nature Biotech., 2020). We hope to extend this research by developing more copper reporters, understanding the biochemical implications of copper complexation, and broadening treatment strategies for triple-negative breast cancers.
