One of the major hurdles in cancer chemotherapy is the inability of the agent to selectively target tumor cells. The over expression of the FR-alpha on the surface of a number of tumors including ovarian, endometrial, kidney, lung, mesothelioma, breast and brain and FR-beta on the surface of myeloid leukemia has prompted the development of folic acid and the pteroyl moiety as selective targeting agents to FR-alpha expressing tumors. Thus, conjugates of folic acid and pteroates have been used to selectively deliver toxins, liposomes, imaging and cytotoxic agents to FR-alpha expressing tumors with a high degree of success. The process of utilizing a cytotoxic conjugate with the folic acid or pteroyl moiety as an antitumor agent requires the additional step of cleavage of the conjugate. This cleavage needs to occur selectively inside the tumor cell to release the cytotoxic agent. Premature release of the cytotoxic agent abrogates selectivity and leads to toxicity. The design of a cytotoxic agent that itself selectively targets the FR-alpha and is not appreciably taken up by the RFC would afford highly selective agents against FR-alpha expressing tumors without serious toxicity. We have recently discovered two compounds, AAG366 and AAG344 that are unique and are inhibitors of glycinamide ribonucleotide formyltransferase (GARFTase), are poorly taken up by the RFC and potently inhibits the growth of FR-alpha expressing KB tumor cells with IC50 values of 2.5 and 2.9 nM respectively. AAG366 and AAG344 are remarkable 100-345-fold more inhibitory to tumor cells expressing the FR-a compared to cells that do not express the FR-alpha.
The Specific Aims of this proposal are: 1) to synthesize analogs of AAG366 and AAG344 to provide a structure-activity relationship (SAR) study to optimize the antitumor activity and the inhibitory activity against GARFTase as well as the high affinity binding and uptake by FR-alpha; 2) to test the analogs for cytotoxicity in isogenic (CHO, KB, SKOV3, OVAR3, CCRF-CEM) cell line models with established differences in FR-alpha, RFC and folylpolyglutamate synthetase, to identify the molecular targets by nucleoside and aminoimidazole carboxamide protection from cytotoxicity by in situ metabolic labeling with radiolabeled (glycine, formate) biosynthetic precursors to determine affinities for FR binding, and to determine the inhibition of target enzyme GARFTase and other folate metabolizing enzymes by studies with isolated enzymes; 3) to evaluate the in vivo antitumor activity of AAG366, AAG344 and selected analogs against FR-alpha expressing tumors. This study will provide a comprehensive SAR and should afford optimized analogs with increased antitumor activity against FR-alpha expressing tumors in vitro and in vivo. This study will also further define the mechanism(s) of action of these novel analogs and could provide agents to be used as monotherapy or in combination for clinical use with a different spectrum of antitumor activity and reduced toxicity than those currently in use. ? ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA125153-03
Application #
7475243
Study Section
Drug Discovery and Molecular Pharmacology Study Section (DMP)
Program Officer
Lees, Robert G
Project Start
2006-09-26
Project End
2011-07-31
Budget Start
2008-08-01
Budget End
2009-07-31
Support Year
3
Fiscal Year
2008
Total Cost
$253,704
Indirect Cost
Name
Duquesne University
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
004501193
City
Pittsburgh
State
PA
Country
United States
Zip Code
15282
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