There is an increasing appreciation of the role of angiogenesis in the growth and metastasis of solid tumors, and data suggest that antiangiogenesis therapy could be a viable approach to treatment of advanced disease. Thymidine phosphorylase (TP) has been recently found to be an angiogenic factor, an observation that was based on its identity to a previously described angiogenic factor, platelet-derived endothelial cell growth factor (PD-ECGF). TP is expressed at high levels in many human solid tumors including colorectal cancers, and the level of its expression has been correlated in numerous clinical studies, including one we conducted, with neovascularization, tumor aggressiveness, and poor patient prognosis. Immunohistochemical evaluation of TP by us and others has demonstrated that elevated TP expression in many, but not all, colon tumors occurs mainly in tumor-associated macrophages (TAMs), rather than in the tumor cells. We have found that a human monocytic cell line (THP1) expressed a higher level of TP than human colon carcinoma cell lines. We also found that TNF-alpha stimulated TP expression in THP1 cells and in WiDr colon carcinoma cells, and we will use these two cell lines as well as primary cultures of TAMs to determine the molecular basis for the regulation of TP expression (i.e. transcriptional, post-transcriptional). TP's angiogenic activity appears to be dependent on its catalytic activity, with 2-D-deoxyribose (2dR) serving as the putative mediator of endothelial cell activation. We have established an in vitro model in which human cancer cells or monocytes can be co-cultured with normal human endothelial cells (HUVEC), and can induce their migration in a TP-dependent manner. We will use this model to begin to determine the mechanisms by which TP stimulates HUVEC and human microvascular endothelial cell (HMEC) migration, using analogs of thymidine and 2dR we will synthesize to test the hypothesis that migration is solely related to 2dR formation, and by using a TP inhibitor we have made and a neutralizing TP antibody to test the hypothesis that TP's intracellular actions are sufficient to stimulate HUVEC migration. Little further is known of the mechanism of action of TP. Our studies showing differences in the specific integrins which mediate TP vs. VEGF-induced HUVEC migration are the first to explore the cellular actions of TP on endothelial cells. We have also found that TP induces tyrosine phosphorylation of FAK (focal adhesion kinase), a protein that plays a central role in cell attachment, migration, and signaling. We will continue our studies to define the integrin-associated signal transduction pathways that are activated by TP in HUVEC and HMEC, testing the hypothesis that these differ initially from those occurring in response to angiogenic factors such as VEGF and bFGF, which have clearly defined cell-surface receptors. In addition to providing a basic understanding of the mechanisms of angiogenic action of TP colon cancers, these studies could provide insights to optimize the use of current and future anti-angiogenic agents which target integrins.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA089352-03
Application #
6633905
Study Section
Experimental Therapeutics Subcommittee 1 (ET)
Program Officer
Ault, Grace S
Project Start
2001-06-01
Project End
2005-05-31
Budget Start
2003-06-13
Budget End
2004-05-31
Support Year
3
Fiscal Year
2003
Total Cost
$237,718
Indirect Cost
Name
Montefiore Medical Center (Bronx, NY)
Department
Type
DUNS #
041581026
City
New York
State
NY
Country
United States
Zip Code
10467
Schwartz, Edward L (2009) Antivascular actions of microtubule-binding drugs. Clin Cancer Res 15:2594-601
Lu, Haiyan; Klein, Robert S; Schwartz, Edward L (2009) Antiangiogenic and antitumor activity of 6-(2-aminoethyl)amino-5-chlorouracil, a novel small-molecule inhibitor of thymidine phosphorylase, in combination with the vascular endothelial growth factor-trap. Clin Cancer Res 15:5136-44
Dalyot-Herman, Nava; Delgado-Lopez, Fernando; Gewirtz, David A et al. (2009) Interference with endothelial cell function by JG-03-14, an agent that binds to the colchicine site on microtubules. Biochem Pharmacol 78:1167-77
Hotchkiss, Kylie A; Ashton, Anthony W; Schwartz, Edward L (2003) Thymidine phosphorylase and 2-deoxyribose stimulate human endothelial cell migration by specific activation of the integrins alpha 5 beta 1 and alpha V beta 3. J Biol Chem 278:19272-9
Zhu, Geng Hui; Schwartz, Edward L (2003) Expression of the angiogenic factor thymidine phosphorylase in THP-1 monocytes: induction by autocrine tumor necrosis factor-alpha and inhibition by aspirin. Mol Pharmacol 64:1251-8
Zhu, Geng Hui; Lenzi, Michelle; Schwartz, Edward L (2002) The Sp1 transcription factor contributes to the tumor necrosis factor-induced expression of the angiogenic factor thymidine phosphorylase in human colon carcinoma cells. Oncogene 21:8477-85
Hotchkiss, Kylie A; Ashton, Anthony W; Mahmood, Radma et al. (2002) Inhibition of endothelial cell function in vitro and angiogenesis in vivo by docetaxel (Taxotere): association with impaired repositioning of the microtubule organizing center. Mol Cancer Ther 1:1191-200