We propose to develop chip-based microarray devices for protease activity profiles of cancer specimens for protease discovery and a new molecular diagnosis paradigm. By marrying the strengths of the Craik and Majumdar labs, we intend to develop a device that could be used to rapidly monitor the proteolytic activities of proteases in a multiplex fashion allowing large numbers of samples to be tested simultaneously in a rapid, multiplexed, quantitative determination of protease activity in nanoliter-scale cell lysates. Since early detection of cancer is closely related to cancer survival we are very encouraged by the prospects of using protease activity as a marker for cancer detection.
The goal of this proposal is to develop a novel microarray platform to simultaneously and quantitatively detect active serine proteases from biological specimens. The immediate application will be to detect three important proteases that are dys-regulated during the various stages of cancer progression. This will be achieved by integrating the knowledge and expertise in the engineering of microarrays with basic science and clinical oncology. By leveraging our understanding of the role of extracellular proteases in cancer we will target MTSP-1/matriptase, urokinase type plasminogen activator and prostate specific antigen for functional monitoring of their activity for both preclinical and eventually, clinical use. In particular, the proposal will focus on: 1) developing molecular probes that specifically and covalently bind only to active proteases; 2) developing and optimizing the surface chemistry for attaching these probes on solid surfaces that minimizes non-specific binding and maximizes protease capture efficiency; 3) developing a microarray for highthroughput and multiplexed analysis of active proteases from cancer samples to be interrogated optically using fluorescent or chemiluminescent labeling of antibodies. The proposal takes advantage of a collaboration between the Craik laboratory at UC San Francisco and the Majumdar laboratory at UC Berkeley and will be greatly facilitated by QB3 (California Institute for Quantitative Biomedical Research), that is a cooperative effort between the state of California and the UC campuses at Berkeley, San Francisco, and Santa Cruz. The proposal will exploit molecular oncology expertise and samples provided by the UCSF Comprehensive Cancer Center. Support functions include the Molecular Foundry, the Microfabrication Laboratory, the Biomolecular Nanotechnology Center, facilities for manufacture of microarrays, and data management and integration.