Our work is directed toward developing peptides, peptide mimetics, and bivalent nucleotides that will be potent and selective inhibitors of mammalian ribonucleotide reductase (mRR), a type 1 RR, and by extension, effective inhibitor of tumor cell proliferation and viral replication. The reaction catalyzed by RR, the reduction of nucleoside diphosphates (NDPs) to deoxynucleosides diphosphates (dNDPs), is the rate-limiting step in the de novo synthesis of dNTPs, and hence of DNA, and as such is a clear target for therapeutic agents directed against these diseases. Active RR depends on the association of two different subunits, mR1 and mR2. The oligomeric structures of the active enzyme (mR1mR2)n is dependent on allosteric effectors, with n ranging from two to six. The C-terminal peptide of mR2 inhibits RR activity. Thus, a lead molecule for an mRR inhibitor is the acetylated heptapeptide AcFTLDADF, denoted P7, corresponding to the C-terminus of mR2. Recently, we have found that a cyclic peptide, mimicking P7 and denoted cycP7, and the simple protected amino acids FmocTrp and FmocPhe, are also effective inhibitors. In addition, within the R1 dimer, the active site of RR lies close to an allosteric specificity site, raising the possibility that these sites can be effectively bridged by bivalent nucleotide inhibitors. We propose to continue efforts begun in the current grant period to develop peptide-based and bivalent nucleotide inhibitors that will not only have high affinity for mR1 but also be effective inhibitors in tissue cell culture. Such molecules would be promising candidates for new therapeutic agents. In carrying out this work we will make extensive use of trNOE NMR and modeling, employing a feedback logic to the design of new peptides and peptidomimetics and the evolving model for the mR1 peptide binding site. We will also attempt to obtain crystals of mR1 and of the mR1.mR2 complex suitable for high resolution structure determination as an aid to inhibitor design. Finally, high affinity peptides and peptidomimetics will be converted into inhibitors of tumor cell proliferation and viral replication in cell culture by conjugation with carriers mediating delivery into intact cells. Such carriers, along with bivalent nucleosides, and phosphorylated bivalent nucleoside prodrugs, will permit in vivo testing of our compounds.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA058567-07
Application #
6729170
Study Section
Bio-Organic and Natural Products Chemistry Study Section (BNP)
Program Officer
Fu, Yali
Project Start
1997-06-19
Project End
2006-04-30
Budget Start
2004-05-01
Budget End
2005-04-30
Support Year
7
Fiscal Year
2004
Total Cost
$313,124
Indirect Cost
Name
University of Pennsylvania
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Xu, Hai; Fairman, James W; Wijerathna, Sanath R et al. (2008) The structural basis for peptidomimetic inhibition of eukaryotic ribonucleotide reductase: a conformationally flexible pharmacophore. J Med Chem 51:4653-9
Fuertes, Michael J; Kaur, Jaskiran; Deb, Prasant et al. (2005) Design, synthesis, and evaluation of octahydropyranopyrrole-based inhibitors of mammalian ribonucleotide reductase. Bioorg Med Chem Lett 15:5146-9
Gao, Ying; Kashlan, Ossama B; Kaur, Jaskiran et al. (2005) Mechanisms of action of peptide inhibitors of mammalian ribonucleotide reductase targeting quaternary structure. Biopolymers 80:9-17
Cooperman, Barry S; Gao, Ying; Tan, Chiheng et al. (2005) Peptide inhibitors of mammalian ribonucleotide reductase. Adv Enzyme Regul 45:112-25
Radivoyevitch, Tomas; Kashlan, Ossama B; Cooperman, Barry S (2005) Rational polynomial representation of ribonucleotide reductase activity. BMC Biochem 6:8
He, Jian; Roy, Beatrice; Perigaud, Christian et al. (2005) The enantioselectivities of the active and allosteric sites of mammalian ribonucleotide reductase. FEBS J 272:1236-42
Tan, Chiheng; Gao, Ying; Kaur, Jaskiran et al. (2004) More potent linear peptide inhibitors of mammalian ribonucleotide reductase. Bioorg Med Chem Lett 14:5301-4
Kashlan, Ossama B; Cooperman, Barry S (2003) Comprehensive model for allosteric regulation of mammalian ribonucleotide reductase: refinements and consequences. Biochemistry 42:1696-706
Cooperman, Barry S (2003) Oligopeptide inhibition of class I ribonucleotide reductases. Biopolymers 71:117-31
Kashlan, Ossama B; Scott, Charles P; Lear, James D et al. (2002) A comprehensive model for the allosteric regulation of mammalian ribonucleotide reductase. Functional consequences of ATP- and dATP-induced oligomerization of the large subunit. Biochemistry 41:462-74

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