This proposal provides a means of laying the groundwork for the candidate's career in scleroderma research. Her formal Ph.D. training was primarily in pharmacology and receptor signaling. She has developed an interest in proteomics technologies and wants to apply them to scleroderma research. In the course of this award period, the candidate expects to develop skills in the areas of rheumatology, biology of scleroderma fibroblasts, mass spectrometry, and proteomics that are much needed, through both didactic and laboratory experience. Then, using these new skills in combination with her receptor pharmacology and signal transduction background, she will develop independent research projects investigating the fundamental mechanisms of scleroderma fibrosis. Toward these ends, a proposal has been prepared which should provide ample opportunity to develop new skills and to obtain the theoretical background in scleroderma research required for the candidate's long-term goals. The research plan of the proposal is based on the observations that connective tissue growth factor (CTGF) Specifically binds numerous proteins in lung fibroblasts derived from scleroderma patients. CTGF is highly expressed in many fibrotic disorders and is considered to be a molecular marker of fibrosis. Interstitial lung fibrosis strikes up to 80% of scleroderma patients and is now the leading cause of death. Since scleroderma lung fibroblasts not only synthesize increased amounts of CTGF, but also contain proteins that specifically interact with CTGF, we postulate that such interactions are critical for the pathogenesis and expansion of scleroderma pulmonary fibrosis. Our research plan aims to identify the CTGF-interacting proteins and to define the components of a signal transduction complex for CTGF in scleroderma lung fibroblasts. This project will involve a combination of biochemical and proteomic techniques applied to the detailed reconstruction of CTGF's microenvironment in scleroderma lung fibroblasts. Part of the proposed studies will be carried out with BIAcore 3000, which allows detection of even transient binding of proteins. The use of this commercial instrument in this manner is a novel technique, which is not commonly available. An additional factor that makes this proposal particularly unique is the availability of the well-equipped Mass Spectrometry Facility, which offers instrumentation to sequence proteins at picomolar levels and lower. Cutting-edge proteomics technologies will allow us to identify CTGF-interacting proteins resolved via ID or 2D gel electrophoresis or eluted from the BIAcore sensor chip, as well as to spot the complex of protein cross linked to CTGF. The identified protein(s) might serve then as potential target(s) in the treatment of scleroderma pulmonary fibrosis for which no proven, effective therapy currently exists.
