The long-term goal of our research is to establish a model of the invasion of the normal brain by malignant astrocytoma tumors that permits the development of specific and effective therapies. The goal of this proposal is to test the hypothesis that focal adhesion kinase (p125FAK) and the associated adaptor protein p130 CAS mediate malignant astrocytoma cell migration/invasion of normal brain. We have established that integrins alphavbeta3 and alphavbeta5 mark the malignant astrocyte phenotype and mediate attachment and migration of these cells toward relevant ligands, such as vitronectin, vitronectin attachment of malignant astrocytoma cells result in activation of p125FAK, a cytoplasmic tyrosine kinase that is a key attachment of malignant astrocytoma cells result in activation of p125FAK, a cytoplasmic tyrosine kinase that is a key component of integrin-associated signalling cascades in other cell types; and over-expression of p125FAK in malignant astrocytoma cells results in increased vitronectin-directed migration. We propose to 1) determine the role of p125FAK in the migration and invasion of malignant astrocytoma cells in vitro, 2) determine the role of p130CAS in the migration and invasion of malignant astrocytoma cells in vitro, and 3) determine the biologic role of p125FAK and p130 CAS in the intracerebral acid mouse model of malignant astrocytoma cells by injecting and propagating malignant astrocytoma cells by injecting and propagating malignant astrocytoma cells over- expressing wild type and mutant constructs of p125FAK and p130CAS. The expression and level of phosphorylation of endogenous p125FAK and p130CAS in U-251 MG and U-87 MG human malignant astrocytoma cells, and rat C6 glioma cells, as well as normal neonatal rat astrocytes will be determined, followed by determination of the binding of p125FAK to Src, p130 CAS and Grb2, and the binding of p130CAS to Crk, Nck, Src and p125 FAK. Wild-type and mutant constructs of p125 FAK and p130 CAS will be over-expressed in these neoplastic and normal astrocytes followed by evaluation of their adhesive, migratory, invasive and proliferative properties in vitro, and their proliferative and invasive properties in vivo. The expression and level of phosphorylation of exogenous (transfected) p125 FAK and p130CAS, as well as the binding of p125FAK to p130CAS, Src, and Grb2, and the binding of p130 CAS to Src, Nck and p125FAK will be determined in these cells. Completion of this work will establish a model of p125 FAK and p130CAS signaling after ligation of integrins on astrocytoma cells, which will lead to the development of therapy directed toward specific steps in the invasion process.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA059958-10
Application #
6628303
Study Section
Special Emphasis Panel (ZRG1-MEP (01))
Program Officer
Sussman, Daniel J
Project Start
1994-01-14
Project End
2005-01-31
Budget Start
2003-04-15
Budget End
2005-01-31
Support Year
10
Fiscal Year
2003
Total Cost
$226,282
Indirect Cost
Name
University of Alabama Birmingham
Department
Pathology
Type
Schools of Medicine
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Ding, Qiang; Grammer, J Robert; Nelson, Mark A et al. (2005) p27Kip1 and cyclin D1 are necessary for focal adhesion kinase regulation of cell cycle progression in glioblastoma cells propagated in vitro and in vivo in the scid mouse brain. J Biol Chem 280:6802-15
Hecker, Timothy P; Ding, Qiang; Rege, Tanya A et al. (2004) Overexpression of FAK promotes Ras activity through the formation of a FAK/p120RasGAP complex in malignant astrocytoma cells. Oncogene 23:3962-71
Repovic, Pavle; Fears, Constance Y; Gladson, Candece L et al. (2003) Oncostatin-M induction of vascular endothelial growth factor expression in astroglioma cells. Oncogene 22:8117-24
Ding, Qiang; Stewart Jr, Jerry; Olman, Mitchell A et al. (2003) The pattern of enhancement of Src kinase activity on platelet-derived growth factor stimulation of glioblastoma cells is affected by the integrin engaged. J Biol Chem 278:39882-91
Natarajan, Meera; Hecker, Timothy P; Gladson, Candece L (2003) FAK signaling in anaplastic astrocytoma and glioblastoma tumors. Cancer J 9:126-33
Hecker, Timothy P; Grammer, J Robert; Gillespie, G Yancey et al. (2002) Focal adhesion kinase enhances signaling through the Shc/extracellular signal-regulated kinase pathway in anaplastic astrocytoma tumor biopsy samples. Cancer Res 62:2699-707
Ding, Qiang; Stewart Jr, Jerry; Prince, Charles W et al. (2002) Promotion of malignant astrocytoma cell migration by osteopontin expressed in the normal brain: differences in integrin signaling during cell adhesion to osteopontin versus vitronectin. Cancer Res 62:5336-43
Han, X; Stewart Jr, J E; Bellis, S L et al. (2001) TGF-beta1 up-regulates paxillin protein expression in malignant astrocytoma cells: requirement for a fibronectin substrate. Oncogene 20:7976-86
Gladson, C L; Stewart, J E; Olman, M A et al. (2000) Attachment of primary neonatal rat astrocytes to vitronectin is mediated by integrins alphavbeta5 and alpha8beta1: modulation by the type 1 plasminogen activator inhibitor. Neurosci Lett 283:157-61
Wang, D; Yamamoto, S; Hijiya, N et al. (2000) Transcriptional regulation of the human osteopontin promoter: functional analysis and DNA-protein interactions. Oncogene 19:5801-9

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