Cancer's intractable nature is driving the search for better, more effective anti-neoplastic agents. The search typically begins with high throughput screening (HTS) of combinatorial chemical libraries. Technological advances in HTS technology have greatly increased the efficiency of these screens, leading to an explosion of drug candidates. The increased production of HTS campaigns has uncovered a new bottleneck in the drug discovery process: the biological characterization of hit compounds. The larger volume of candidate compounds and the desire for more informative assays necessitates higher-throughput, high-content cell-based secondary assays. The design of assays for cancer therapeutics faces even larger obstacles due to the inherent diversity of this disease. Within a specific cancer type, thousands of different combinations of mutations and gene expression profiles can lead to oncogenic transformation making no specific cell type representative. In this Phase I SBIR proposal we outline a solution to these problems with the implementation of a novel cell-based assay that multiplexes experimental readouts and libraries of cell types. By multiplexing the cell lines in a high-content screening format we are able to obtain a systems level view of compound action while reducing the number of samples, providing a higher-throughput and more informative secondary screening platform. The foundation of the work is a combination of selective labeling, or barcoding, of cells and multiparameter cell-based assays.
In Aim I, 27 breast cancer cell lines will be labeled with unique fluorescent barcode signatures. The cell lines are manipulated as one sample; however the data can be separated upon analysis to determine the effects of compounds on each individual cell type.
In Aim II, five cellular assays are optimized to be used in conjunction with the barcoding and provide the simultaneous analysis of phosphoproteins, cell cycle, and apoptosis for each of the 27 cell lines. As an example of the power of this system and to validate the technology, the breast cancer panel will be profiled against eight commonly used chemotherapy drugs using the five assay readouts in Aim III. The completion of these studies will provide a unique method of high-content secondary screening across multiple cell lines that radically reduces the number of samples, enabling these type of screens to be performed earlier thereby facilitating the selection of higher quality lead compounds.
The platform technology described in this proposal gives scientists the power to assess the effects of drugs on dozens of cell lines simultaneously, greatly expanding the amount of pertinent biological information available on drug candidates. This will lead to the selection of better drugs, faster, drastically reducing the resources needed to find therapeutic molecules and increasing the chances of success. ? ? ?
