Intimal hyperplasia and/or constrictive remodeling are major causes of recurrent disease followingvascular reconstruction. Transforming growth factor-beta (TGF-P) is an important cytokine that has been found to be integrally involved in both of these processes. The goal of this proposal is to explore the role of TGF-P signalingintermediates in the pathophysiology of intimal hyperplasia and constrictive remodeling. We will specifically examine a family of intracellular signalingproteins, termed SMADs, and their role in these two processes. Our laboratory and others have shown that TGF-P inhibits smooth muscle cell (SMC) proliferation and stimulates apoptosis;functionsthat would limit intimal hyperplasia. Conversely, TGF-P stimulates extracellular matrix production (collagen I being the predominant type), an effect that enhances the formation of hyperplastic lesions. In preliminary experiments, we have found that Smads selectively mediate the effects of TGF-p. Specifically, SmadS stimulates collagen expression but has no effect on SMC proliferation.These differential functions of SmadS in SMCs may allow us to design molecular tools that can selectively preserve the """"""""inhibitory"""""""" effects but block the """"""""stimulatory"""""""" effects of TGF-P on intimal hyperplasia. In the studies detailed in this proposal, we will further define the role of SmadS and other Smad proteins in vascular SMC function.
In specific aim I we will evaluate the function of Smads in SMC by transiently transfecting wild type and dominant negative mutant Smads as well as reporter genes for SMC function into SMC lines. We will then create adenoviral vectors that express selected Smads that appear to have a significant impact on SMC physiology and test their effect in human primary vascular SMC.
In specific aim II, we will explore the interaction of TGF-P with the promoter of the gene of type I collagen. The ultimate goal of these experiments will be to design an oligonucleotide decoy that can act as a selectivegenetic inhibitor of collagen transcription. Finally, in specific aim III we will test the hypothesis that stimulation and/or inhibition of Smads or selective inhibition of the collagen promoter can limit the formation of intimal hyperplasia in a rat model of vascular injury. We anticipatethat the results of these studies will both enhance our knowledge of the pathophysiology of intimal hyperplasia and also allow for the design of novel therapies to inhibit this complex human disease process which affects thousands of patients each year. PERFORMANCE SITE(S) (organization, city, state) Weill MedicalCollege CornellUniversity 1300 York Avenue New York, NY 10021 KEY PERSONNEL. See instructions on Page 11. Use continuation pages as needed to provide the required Name Organization K. Craig Kent, MD Weill Medical College Cornell University Bo Liu, PhD Weill Medical College Cornell University Otway Louie, MD Weill Medical College Cornell University Shi-Qin Xu Weill Medical College Cornell University Eseng Lai MD, PhD Memorial Sloan-Ketterin Cancer Center Timothy McCafrey, PhD Weill Medical College Cornell University Michel Mann, MD Harvard Medical School Manikkam Suthanthiran,MD Weill Medical College Cornell University Ron Crystal, MD Weill Medical College Cornell University Joan Massague, Ph.D. Memorial Sloan-Ketterin Cancer Center information in the format shown below. Role on Project PrincipalInvestigator Co-Investigator Research Fellow Technician Collaborator Collaborator Collaborator Collaborator Consultant Consultant PHS 398 (Rev. 4/98) Page 2 BB Number pages consecutively at the bottom throughout the application.Do not use suffixes such as 3a, 3b. CC Principal Investigator/Program Director (Last, first, middle): Kent. K. Craig Type the name of the principal investigatorA^^fcn director at the top of each printed page and ea^Bhntinuation page. (For type specifications, see instructions onpage 6.) ^B^P ^^B^ RESEARCH GRANT TABLE OF CONTENTS Page Numbers Face Page Description, Performance Sites, and Personnel Table of Contents Detailed Budget for Initial Budget Period Budget for Entire Proposed Period of Support Budgets Pertaining to Consortium/ContractualArrangements NA Biographical Sketch-Principal Investigator/Program Director (Not to exceed two pages) Other Biographical Sketches (Not to exceed two pages for each) Other Support Resources 20 Research Plan Introduction to Revised Application (Not to exceed 3 pages) Introduction to Supplemental Application (Not to exceed 1page) a.
Specific Aims 2 1 b. Background and Significance 22-25 c. Preliminary Studies/Progress Report .J (Items a-d: not toexceed 25 pages"""""""") I 26-31 d. Research Design and Methods ^ / 32-45 e. Human Subjects I I 45 f. Vertebrate Animals X. 45 g. Literature Cited 47-51 h. Consortium/Contractual Arrangements i. Consultants Checklist Type density and type size of the entire application must conform to limits provided in instructions on page 6. | x | Check if Appendix (Five collated sets. No page numbering necessary for Appendix.) Appendix is Number of publications and manuscripts accepted or submitted for publication (not to exceed 10) 10 included Other items (list): PHS 398 (Rev. 4/98) (Form Page 3) Page 3 CC Number pages consecutively at the bottom throughout the application. Do not use suffixes such as 3a, 3b.
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