Design and Synthesis of a Transferable Farnesyl Pyrophosphate Analogue to Ras by Protein Farnesyltransferase (original) (raw)
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Inhibition of farnesyltransferase with A-176120, a novel and potent farnesyl pyrophosphate analogue
European Journal of Cancer, 2000
Farnesylation of Ras is required for its transforming activity in human cancer and the reaction is catalysed by the enzyme farnesyltransferase. Recently, we discovered a novel chemical series of potent farnesyl pyrophosphate (FPP) analogues which selectively inhibited farnesyltransferase. Our most potent compound to date in this series, A-176120, selectively inhibited farnesyltransferase activity (IC 50 1.2AE0.3 nM) over the closely related enzymes geranylgeranyltransferase I (GGTaseI) (IC 50 423AE1.8 nM), geranylgeranyltransferase II (GGTaseII) (IC 50 3000 nM) and squalene synthase (SSase) (IC 50 >10 000 nM). A-176120 inhibited ras processing in H-ras-transformed NIH3T3 cells and HCT116 K-ras-mutated cells (ED 50 1.6 and 0.5 mM, respectively). The antiangiogenic potential of A-176120 was demonstrated by a decrease in Ras processing, cell proliferation and capillary structure formation of human umbilical vein endothelial cells (HUVEC), and a decrease in the secretion of vascular endothelial growth factor (VEGF) from HCT116 cells. In vivo, A-176120 reduced H-ras NIH3T3 tumour growth and extended the lifespan of nude mice inoculated with H-or K-ras-transformed NIH3T3 cells. A-176120 also had an additive eect in combination with cyclophosphamide in nude mice inoculated with K-ras NIH3T3 transformed cells. Overall, our results demonstrate that A-176120 is a potent FPP mimetic with both antitumour and anti-angiogenic properties.
Farnesyl Diphosphate-Based Inhibitors of Ras Farnesyl Protein Transferase
Journal of Medicinal Chemistry, 1995
The rational design, synthesis, and biological activity of farnesyl diphosphate (FPP)-based inhibitors of the enzyme Ras farnesyl protein transferase (FPT) is described. Compound 3, wherein a ,&carboxylic phosphonic acid type pyrophosphate (PPI surrogate is connected to the hydrophobic farnesyl group by a n amide linker, was found to be a potent (150(FPT) = 75 nM) and selective inhibitor of FPT, as evidenced by its inferior activity against squalene synthetase (15dSS) = 516 pM) and mevalonate kinase (150(MK) = '200 pM). A systematic structureactivity relationship study involving modifications of the farnesyl group, the amide linker, and the PP surrogate of 3 was undertaken. Both the carboxylic and phosphonic acid groups of the P-carboxylic phosphonic acid PP surrogate are essential for activity, since deletion of either group results in 50-2600-fold loss in activity (6-9, 1 5 0 = 4.6-220 pM). The farnesyl group also displays very stringent requirements and does not tolerate one carbon homologation (12, 1 5 0 = 17.7 pM), substitution by a dodecyl fragment (14,150 = 9 pM), or introduction of a n extra methyl group at the allylic position (18,150 = 55 ,LAM). Modifications around the amide linker group of 3 were more forgiving, as evidenced by the activity ofN-methyl analog (21,150 = 0.53 pM), the one carbon atom shorter farnesoic acid-derived retroamide analog (32,150 = 250 nM), and the exact retroamide analog (49, 1 5 0 = 50 nM). FPP analogs such as 3, 32, and 49 are novel, potent, selective, small-sized, nonpeptidic inhibitors of FPT that may find utility as antitumor agents.
Biochemistry, 2012
Farnesylation is an important post-translational modification essential for proper localization and function of many proteins. Transfer of the farnesyl group from farnesyl diphosphate (FPP) to proteins is catalyzed by protein farnesyltransferase (FTase). We employed a library of FPP analogues with a range of aryl groups substituting for individual isoprene moieties to examine some of the structural and electronic properties of analogue transfer to peptide catalyzed by FTase. Analysis of steady-state kinetics for modification of peptide substrates revealed that the multiple turnover activity depends on the analogue structure. Analogues where the first isoprene is replaced by a benzyl group and an analogue where each isoprene is replaced by an aryl group are good substrates. In sharp contrast with the steady-state reaction, the single turnover rate constant for dansyl-GCVLS alkylation was found to be the same for all analogues, despite the increased chemical reactivity of the benzyl analogues and the increased steric bulk of other analogues. However, the single turnover rate constant for alkylation does depend on the Ca 1 a 2 X peptide sequence. These results suggest that the isoprenoid transition state conformation is preferred over the inactive E•FPP• Ca 1 a 2 X ternary complex conformation. Furthermore, these data suggest that the farnesyl binding site in the exit groove may be significantly more selective for the farnesyl diphosphate substrate than the active site binding pocket and therefore might be a useful site for design of novel inhibitors.
Biochemistry, 2001
Farnesylation is a posttranslational lipid modification in which a 15-carbon farnesyl isoprenoid is linked via a thioether bond to specific cysteine residues of proteins in a reaction catalyzed by protein farnesyltransferase (FTase). We synthesized the benzyloxyisoprenyl pyrophosphate (BnPP) series of transferable farnesyl pyrophosphate (FPP) analogues (1a-e) to test the length dependence of the isoprenoid substrate on the FTase-catalyzed transfer of lipid to protein substrate. Kinetic analyses show that pyrophosphates 1a-e and geranyl pyrophosphate (GPP) transfer with a lower efficiency than FPP whereas geranylgeranyl pyrophosphate (GGPP) does not transfer at all. While a correlation was found between K m and analogue hydrophobicity and length, there was no correlation between k cat and these properties. Potential binding geometries of FPP, GPP, GGPP, and analogues 1a-e were examined by modeling the molecules into the active site of the FTase crystal structure. We found that analogue 1d displaces approximately the same volume of the active site as does FPP, whereas GPP and analogues 1a-c occupy lesser volumes and 1e occupies a slightly larger volume. Modeling also indicated that GGPP adopts a different conformation than the farnesyl chain of FPP, partially occluding the space occupied by the Ca 1 a 2 X peptide in the ternary X-ray crystal structure. Within the confines of the FTase pocket, the double bonds and branched methyl groups of the geranylgeranyl chain significantly restrict the number of possible conformations relative to the more flexible lipid chain of analogues 1a-e. The modeling results also provide a molecular explanation for the observation that an aromatic ring is a good isostere for the terminal isoprene of FPP.
2018
The post-translational addition of a farnesyl moiety to the Ras oncoprotein is essential for its membrane localization and is required for both its biological activity and ability to induce malignant transformation. The present invention describes design and synthesis of a farnesylpyrophosphate (FPP) analog, 8-anilinogeranyl pyrophosphate (AGPP) that is transferred to Ras by farnesyltransferase (FTase), in Which the (Jo-terminal isoprene unit of the farnesyl group has been replaced With an aniline functionality. AGPP potently inhib
New Biotechnology, 2013
As isoprenoid biosynthetic pathway has gained importance since last few years, key enzymes of this pathway have been characterized and their functional roles in the cell metabolism have been explored using molecular biology approaches. A key enzyme in this pathway is farnesyl pyrophosphate (EC 2.5.1.10) synthase (FPPS) which supplies precursors for the biosynthesis of essential isoprenoids like carotenoids, withanolides, ubiquinones, dolichols, sterols, among others and also helps in farnesylation and geranylation of proteins. It is a chain elongation enzyme which catalyzes head to tail condensation of two molecules of isopentenyl diphosphate with dimethylallyl diphosphate to form farnesyl pyrophosphate (FPP). Recent studies have validated FPPS as a molecular target of bisphosphonates for drug development against tumors as well as human pathogens. The present paper synthesizes the information on characterization, structural and functional relationships, evolution, localization as well as advances on FPPS enzyme as a target for drug development.
International Journal of Peptide Research and Therapeutics, 2007
The development of tools for proteomic analysis is an active area of research. Here, we report on the synthesis of 12-propargoxyfarnesyl diphosphate (1), an alkyne-containing analogue of farnesyl diphosphate (FPP), and its enzymatic incorporation into peptide substrates by both protein-farnesyltransferase (PFTase) and protein-geranylgeranyltransferase type I (PGGTase-I). Compound 1 was prepared from farnesol in 6 steps. Kinetic analyses indicate that 1 is incorporated into cognate peptide substrates by PFTase or PGGTase at concentrations and rates comparable to those of the natural lipid substrates for these enzymes, and mass spectrometric analyses proved the structures of the prenylated peptide products. Incubation of 1 in the presence of PFTase and PGGTase peptide substrates, and the cognate transferases, results in the simultaneous prenylation of both peptides emphasizing the dual substrate nature of 1. Thus, because 1 is a substrate for both enzymes, it can be used to introduce alkyne functionality into proteins that are normally either farnesylated or geranylgeranylated. This approach should be useful for a broad range of applications ranging from selective protein labeling to proteomic analysis.
Inhibitors of farnesyl:protein transferase—A possible cancer chemotherapeutic
1996
The recent interest in inhibitors of farnesyl:protein transferase (FPTase) has resulted in a better understanding of the enzymology of this protein. Rationally designed inhibitors of prenyl transfer have emerged as potential new drug candidates because of the insight gained over bow a prenyl group is enz3'matically transferred onto a peptide thiol. This paper will explore how advances in our understanding of FPTase mediated catalysis has affected the design of FPTase inhibitors as possible cancer therapeutic agents. Without structural information of the enzyme, substrate analogues comprise the first area of drug design: these include peptidomimetics of the four C-terminal amino acids of rasP21 as well as farnesyl diphosphate analogs. In addition, phosphate anion was found to enhance the inhibitory potency of certain compounds known to be competitive with respect to farnesyl diphosphate and therefore incorporation of the phosphate anion may also provide a basis for improved inhibitor design.