Nanoparticle libraries to identify novel ligands for interrogation of cellular pathways
Small molecules derived from diversity oriented synthesis (DOS) are commonly used as a discovery platform to identify new drugs, to approximate ligand/receptor binding kinetics and resultant hits have been used for systems wide perturbations in cell based assays. This project uses nanoparticle platforms to immobilize DOS compounds to increase target avidity through multivalency, to create new chemical entities, to visualize biological interactions in vivo and to impart more optimal pharmacokinetics to small molecules. Based on our prior work with novel (magneto)fluorescent nanoparticles, we have begun to test different library methods and differentially screen thousands of molecules against known (and unknown) biological targets. The library methods employed include chemical ones (DOS compounds, natural compounds, approved drugs, curated small molecule collections, peptide libraries and aptamer libraries) and biological ones (phage display, mRNA display, SELEX, Bacterial display, ribosomal display, cell display, covalent DNA display, yeast 2 hybrid, sh/si RNA). These methods and materials will facilitate the development of functional nanomaterials for applications such as differentiating cell lines, detecting distinct cellular states and targeting specific cell types.
Identification of key genes and gene function involved in complex human diseases
Genomic profiling has identified large data sets on many complex diseases in the mouse and humans. However, the variability of such data sets, importance of network nodes and clinically imageable targets are much more difficult to discern. This project uses text and data mining tools as well as network analyses to identify novel targets for specific cancer types, Type 1 diabetes and atherosclerosis. A second complementary approach has been the use of in vivo library displays to identify "hits" and then discern the cellular binding partner. Using these methods we are currently working on several unique leads including cell surface proteases.
Development of novel miniaturized sensing technologies (chips) for real-time high-throughput analysis
Multiplexed, platform independent and rapid analysis of biological specimen remains a major bottleneck in unravelling complex biological phenomena. A number of established techniques provide accurate measurements, facilitate early disease detection and have been used to gain valuable insights into biology at the systems level. However, many of the techniques rely on extensive and time consuming purification of samples, typically followed by a set of amplification strategies and often do not allow rapid multiplexed measurements required in complex diseases. We have recently developed a chip based micro-NMR (µNMR) system to perform highly sensitive measurements in complex biological samples. NMR devices allow measurements in turbid samples, characterization of chemical species and form the basis of clinical imaging systems. Using microfabrication technology we miniaturized elements of an NMR system to perform proton T2 measurements of aqueous biological samples. Using readily available magnetic nanoparticles as sensors for proximity assays the technique allows parallel measurements of nucleic acids, proteins, peptides, metabolites and cells. Importantly, the spatial proximity of two nanoparticles by an analyte of choice results in large-scale amplification allowing rapid, parallel detection of biological targets in unprocessed samples. The integration of the sensor with microfluidic system allows facile control and manipulation of small volumes of liquid, and additional magnetic separation and concentration of targets from a complex parent specimen. In this project we apply the chip for parallel measurements of cellular proteins.