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A D Hamilton

Publications and source records attributed to A D Hamilton.

At least 73 records · Page 4Linked to original sources
PubMed1997

Farnesyltransferase as a target for anticancer drug design.

The currently understood function for Ras in signal transduction is in mediating the transmission of signals from external growth factors to the cell nucleus. Mutated forms of this GTP-binding protein are found in 30% of human cancers with particularly high prevalence in colon and pancreatic carcinomas. These mutations destroy the GTPase activity of Ras and cause the protein to be locked in its active, GTP bound form. As a result, the signaling pathways are activated, leading to uncontrolled tumor growth. Ras function in signaling requires its association with the plasma membrane. This is achieved by posttranslational farnesylation of a cysteine residue present as part of the CA1A2X carboxyl terminal tetrapeptide of all Ras proteins. The enzyme that recognizes and farnesylates the CA1A2X sequence, Ras farnesyltransferase (FTase), has become an important target for the design of inhibitors that might be interesting as antitumor agents. Several approaches have been taken in the search for in vivo active inhibitors of farnesyltransferase. These include the identification of natural products such as the chaetomellic and zaragozic acids that mimic farnesylpyrophosphate, bisubstrate transition state analogs combining elements of the farnesyl and tetrapeptide substrates and peptidomimetics that reproduce features of the carboxyl terminal tetrapeptide CA1A2X sequence. This last group of compounds has been most successful in showing highly potent inhibition of FTase and selective blocking of Ras processing in a range of Ras transformed tumor cell lines at concentrations as low as 10 nM. Certain peptidomimetics will also block tumor growth in various mouse models, with apparently few toxic side effects. These results suggest that farnesyltransferase inhibitors hold considerable promise as anticancer drugs in the clinic.

Alkyl and Aryl Transferases↗
PubMed1997

Inhibition of Ras prenylation: a novel approach to cancer chemotherapy.

The demonstration that Ras requires prenylation for its cancer-causing activity led several groups of investigators to an intense search for farnesyltransferase and geranylgeranyltransferase inhibitors as potential anticancer drugs. Rational design of small organic molecules that mimic the carboxyl terminal tetrapeptide prenylation site on Ras resulted in pharmacological agents capable of inhibiting Ras processing and selectively antagonizing oncogenic signaling, and suppressing human tumor growth in mouse models without side effects. These agents presently are undergoing advanced preclinical studies. This review describes the efforts of several groups to design, synthesize and evaluate the biological activities of several classes of prenyltransferase inhibitors. Several important issues, such as mechanism of action of prenyltransferase inhibitors and potential mechanisms of resistance to inhibition of K-Ras farnesylation, are also discussed.

Alkyl and Aryl Transferases↗
PubMed1997

Design, synthesis and evaluation of synthetic receptors for the recognition of aspartate pairs in an alpha-helical conformation.

The specific targeting of protein surface functional groups remains a largely unexplored aspect in molecular recognition. In this study, a series of zwitterionic, 16-mer peptides serve as models for the recognition of carboxylate pairs in proteins. A receptor is described that contains two guanidinium groups separated by 4-5 A by a rigid bicyclo[3.3.0]octane spacer. Modeling studies indicate that such a receptor would be suitable for binding with two aspartate carboxylates when the amino acids are separated by two (i + 3) or three (i + 4) other amino acids in an alpha-helical peptide. Studies employing circular dichroism spectroscopy demonstrated that the addition of the receptor to the i + 3 peptide substrate caused a 23% enhancement of helical structure in 15% water/methanol at 25 degrees C. Other substrate peptides [(i + 1), (i + 4), (i + 7), (i + 10)] showed lower helical induction. Similar, but weaker binding and helical induction were observed under buffered conditions (10 mM Tris-Mes, pH 7.0). These results, along with studies employing a series of related di-cationic receptors, suggest a 1:1 binding model composed of specific hydrogen interactions between each receptor guanidinium with each substrate carboxylate when the peptide adopts a helical conformation.

Amino Acid Sequence↗
PubMed1997

Recognition of neutral species with synthetic receptors.

The design of synthetic receptors for neutral molecules is an area of intense current interest. The area has grown from early work on cyclodextrin or single crown ether complexation to encompass a wide array of receptor shapes and structures. Furthermore, the range of substrate selectivities has increased from simple aromatic or metal ion substrates to include key biological components such as peptides and carbohydrates. Recent advances have included the application of split bead combinatorial methods for the identification of receptors for oligopeptides, the design of self-assembling spherical receptors, the use of multiple hydrogen bonding groups for carboxylic acid and carbohydrate recognition and the achievement of impressive catalytic effects with synthetic receptors.

Binding Sites↗
PubMed1997

Inhibition of the prenylation of K-Ras, but not H- or N-Ras, is highly resistant to CAAX peptidomimetics and requires both a farnesyltransferase and a geranylgeranyltransferase I inhibitor in human tumor cell lines.

The farnesyltransferase (FTase) inhibitor FTI-277 is highly effective at blocking oncogenic H-Ras but not K-Ras4B processing and signaling. While inhibition of processing and signaling of oncogenic K-Ras4B is more sensitive to the geranylgeranyltransferase I (GGTase I) inhibitor GGTI-286 than it is to FTI-277 in K-Ras4B-transformed NIH3T3 cells, the sensitivity of K-Ras as well as H- and N-Ras to the CAAX peptidomimetics in human tumor cell lines is not known. Here, we report that a panel of five human carcinoma cell lines from pancreatic, pulmonary, and bladder origins all express H-, N-, and K-Ras, and their respective prenylation sensitivities to the FTase and GGTase I inhibitors is variable. In all of the cell lines investigated, the prenylation of N-Ras was highly sensitive to FTI-277, and in two of the cell lines, N-Ras showed slight sensitivity to GGTI-298, an analog of GGTI-286. Although the prenylation of H-Ras was also sensitive to FTI-277, complete inhibition of H-Ras processing even at high concentrations of FTI-277 and/or GGTI-298 was never achieved. The prenylation of K-Ras, on the other hand, was highly resistant to FTI-277 and GGTI-298. Most significantly, treatment of human tumor cell lines with both inhibitors was required for inhibition of K-Ras prenylation. In one cell line, the human lung adenocarcinoma A-549, prenylation of K-Ras was highly resistant even when co-treated with both inhibitors. Furthermore, soft agar experiments demonstrated that in all the human tumor cell lines tested inhibition of K-Ras prenylation was not necessary for inhibition of anchorage-independent growth. In addition, although GGTI-298 had very little effect on soft agar growth, the combination of FTI-277 and GGTI-298 resulted in significant growth inhibition. Therefore, the results demonstrate that while FTI-277 inhibits N-Ras and H-Ras processing in the human tumor cell lines evaluated, inhibition of K-Ras processing requires both an FTase inhibitor as well as a GGTase I inhibitor, and that inhibition of human tumor growth in soft agar does not require inhibition of oncogenic K-Ras processing.

Alkyl and Aryl Transferases↗
PubMed1997

Inhibitors of prenyl transferases.

Because farnesylation of Ras is required for its cancer-causing activity, several classes of farnesyl transferase inhibitors have recently been developed as potential anticancer drugs. During the last 12 months, important advances have been made in this field. In this review, we focus on three topics: targets of farnesyl transferase inhibitors other than Ras, alternative prenylation of K-Ras by the closely related prenyl transferase, geranyl geranyl transferase I, and the effects of geranyl geranyl transferase I inhibitors on cell cycle, apoptosis, and human tumor growth.

Animals↗
PubMed1997

Anticancer activity of farnesyltransferase and geranylgeranyltransferase I inhibitors: prospects for drug development.

Inhibition of farnesyltransferase (FTase) has been thoroughly investigated as a strategy to discover novel anticancer drugs because the oncoprotein Ras, requires farnesylation for its cancer-causing activity. Several highly potent and selective FTase inhibitors have been made and show excellent antitumour activity against human tumours in animal models without toxicity to normal cells. However, resistance of the most frequently mutated form of Ras, K-Ras, to FTase inhibitors and its alternative prenylation by geranylgeranyltransferase I (GGTase I), has cast doubts on whether K-Ras is the target for FTase inhibitors. This monthly update focuses on issues of critical importance to the further development of FTase inhibitors as anticancer agents. Alternative prenylation of K-Ras by GGTase I as a mechanism of resistance to FTase inhibitors, targets for FTase inhibitors other than K-Ras and the relevance of GGTase I inhibitors as antitumour agents will be discussed.

Journal Article↗
PubMed1997

Inhibition of Ras prenylation: a signaling target for novel anti-cancer drug design.

The cancer-causing activity of Ras requires the prenylation of a cysteine fourth from its carboxyl terminus. Rational design of peptidomimetics of the carboxyl terminal tetrapeptide prenylation site on Ras resulted in pharmacological agents capable of inhibiting Ras processing, selectively antagonizing oncogenic signaling and suppressing human tumor growth in mouse models without side effects. This mini-review describes the efforts of several groups to design, synthesize and evaluate the biological activities of farnesyltransferase and geranylgeranyltransferase I inhibitors. Among the important issues that will be discussed are the mechanism of action of these inhibitors and the potential mechanisms of resistance to inhibition of K-Ras farnesylation.

Alkyl and Aryl Transferases↗
PubMed1996

Protein geranylgeranylation, not farnesylation, is required for the G1 to S phase transition in mouse fibroblasts.

In order to assess the relative contributions of farnesylated and/or geranylgeranylated proteins on cell cycle progression from G1 to S phase we designed potent and selective farnesyltransferase (FTI-277) and geranylgeranyltransferase-I (GGTI-298) inhibitors. Flow cytometry studies showed that treatment of NIH3T3 cells with GGTI-298 or lovastatin, which inhibits both protein farnesylation and geranylgeranylation, arrested cells in G0/G1 whereas cells treated with FTI-277 progressed normally through the cell cycle. [3H]thymidine incorporation studies showed that mevalonate and geranylgeraniol, but not farnesol, released the lovastatin G1 block. Furthermore, mevalonate release of the lovastatin G1 block was inhibited by GGTI-298 but not by FTI-277. These results demonstrate that geranylgeranylated proteins are required for cells to proceed from G1 to S phase, and that farnesylated proteins do not play an essential role in the G1 to S phase transition

Actins↗
PubMed1996

Platelet-derived growth factor receptor tyrosine phosphorylation requires protein geranylgeranylation but not farnesylation.

We have used specific inhibitors for farnesyltransferase (FTase) and geranylgeranyltransferase (GGTase) I as well as combinations of lovastatin with geranylgeraniol (GGOH) or farnesol (FOH) to investigate the role of protein prenylation in platelet-derived growth factor (PDGF)-induced PDGF receptor tyrosine phosphorylation. NIH-3T3 cells treated with the highly specific FTase inhibitor FTI-277 had no effect on PDGF receptor tyrosine phosphorylation or PDGF activation of mitogen-activated protein kinase (MAPK) at doses that completely inhibit FTase-dependent processing. In contrast, treatment of these cells with GGTase I inhibitor GGTI-298 strongly inhibited receptor tyrosine phosphorylation, and co-treatment with FTI-277 had no additional effect. Interestingly, the inhibitory effect of GGTI-298 on PDGF activation of MAPK was only partial. Furthermore, although lovastatin, which inhibits both protein geranylgeranylation and protein farnesylation, blocked PDGF receptor tyrosine phosphorylation, co-treatment with GGOH, but not FOH, reversed the lovastatin block. In addition, although lovastatin was observed to block MAPK activation by PDGF, co-treatment with GGOH, but not FOH, restored its activation. Further investigations indicated that inhibition of receptor tyrosine phosphorylation was not due to decreased expression of the receptor or to inhibition of GGTase II. Thus, these results demonstrate that PDGF receptor tyrosine phosphorylation requires protein geranylgeranylation but not protein farnesylation and that the tyrosine phosphorylation levels of the receptor are modulated by a protein that is a substrate for GGTase I.

3T3 Cells↗
PubMed1996

The farnesyltransferase inhibitor FTI-277 radiosensitizes H-ras-transformed rat embryo fibroblasts.

Many tumor cells have a greater resistance to ionizing radiation than their normal counterparts, suggesting that the development of drugs that can reduce that radioresistance would potentiate the efficacy of radiation therapy. Because activated H-ras expression has been shown to markedly increase radiation resistance in some transformed cells, the inactivation of H-ras would then be predicted to radiosensitize these tumor cells, while leaving normal cells unaffected. H-ras depends for activity upon farnesylation, which can be blocked by farnesylation inhibitors, including the compound FTI-277. In keeping with this prediction, inhibition of H-ras processing using FTI-277 resulted in higher levels of apoptosis after irradiation and increased radiosensitivity in H-ras-transformed rat embryo cells but did not affect control cells. These experiments suggest that farnesylation inhibitors may prove clinically useful as radiosensitizers of tumors that depend on ras function.

Alkyl and Aryl Transferases↗
PubMed1996

Design and synthesis of non-peptide Ras CAAX mimetics as potent farnesyltransferase inhibitors.

Cysteine farnesylation of the ras oncogene product Ras is required for its transforming activity and is catalyzed by farnesyltransferase (FTase). The Ras carboxyl terminal tetrapeptide CAAX (C is cysteine, A is any aliphatic amino acid, X is methionine or serine) is the minimum sequence for FTase recognition. We report here the design, synthesis, and biological characterization of Ras CAAX non-peptide mimetics in which the cysteine is linked through a reduced pseudopeptide bond to 4-amino-3'-carboxybiphenyl. These non-peptide mimetics are potent inhibitors of FTase (IC50 = 40 nM for the most potent inhibitor) and are highly selective for FTase over GGTase I (geranylgeranyltransferase I). They are not substrates for farnesylation, do not have peptidic features, and have no hydrolyzable bonds. Structure-activity studies reveal the importance of the position of the carboxylic acid on the aryl ring as well as the reduction of the cysteine amide bond. Substitution at the 2-position of 4-amino-3'-carboxybiphenyl increases inhibitory potency, while the removal of the carboxylic acid results in a 10-fold loss of inhibitory activity.

Alkyl and Aryl Transferases↗
PubMed1996

Synthesis and structure of chiral macrocycles containing 2,2'-bipyridine subunits.

A series of macrocyclic receptors has been prepared containing bipyridine groups linked to two amino acids. Variations in both the amino acid and the linking spacer have been made. The structure of the resulting macrocycles has been investigated using 1H NMR spectroscopy and X-ray crystallography. The use of L-valine leads to an open conformation for the macrocycle in which the 2-propyl substituents are directed perpendicular to the plane of the ring leaving the bipyridine and amide groups accessible for binding to a metal or complementary substrate. Proline-based macrocycles take up a twisted arrangement with the linking chain stretched across the face of the bipyridine which takes up a trans conformation. The metal ion binding properties of these derivatives have been investigated and shown to occur only to the valine macrocyles which have the two pyridine rings preorganized for complexation. These macrocycles have also been shown to bind to phenolic hydroxyl groups by using hydrogen-bond donors and acceptors from the amide groups in the linking chain.

2,2'-Dipyridyl↗
PubMed1995

Activated Drosophila Ras1 is selectively suppressed by isoprenyl transferase inhibitors.

Ras CAAX (C = cysteine, A = aliphatic amino acid, and X = any amino acid) peptidomimetic inhibitors of farnesyl protein transferase suppress Ras-dependent cell transformation by preventing farnesylation of the Ras oncoprotein. These compounds are potential anticancer agents for tumors associated with Ras mutations. The peptidomimetic FTI-254 was tested for Ras1-inhibiting activity in whole animals by injection of activated Ras1val12 Drosophila larvae. FTI-254 decreased the ability of Ras1val12 to form supernumerary R7 photoreceptor cells in the compound eye of transformed flies. In contrast, it had no effect on the related supernumerary R7 phenotypes of flies transformed with either the activated sevenless receptor tyrosine kinase, Raf kinase, or a chimeric Ras1val12 protein that is membrane associated through myristylation instead of isoprenylation. Therefore, FTI-254 acts as an isoprenylation inhibitor to selectively inhibit Ras1val12 signaling activity in a whole-animal model system.

Alkyl and Aryl Transferases↗
PubMed1995

Disruption of oncogenic K-Ras4B processing and signaling by a potent geranylgeranyltransferase I inhibitor.

Prenylation of the carboxyl-terminal CAAX (C, cysteine; A, aliphatic acid; and X, any amino acid) of Ras is required for its biological activity. We have designed a CAAX peptidomimetic, GGTI-287, which is 10 times more potent toward inhibiting geranylgeranyltransferase I (GGTase I) in vitro (IC50 = 5 nM) than our previously reported farnesyltransferase inhibitor, FTI-276. In whole cells, the methyl ester derivative of GGTI-287, GGTI-286, was 25-fold more potent (IC50 = 2 microM) than the corresponding methyl ester of FTI-276, FTI-277, toward inhibiting the processing of the geranylgeranylated protein Rap1A. Furthermore, GGTI-286 is highly selective for geranylgeranylation over farnesylation since it inhibited the processing of farnesylated H-Ras only at much higher concentrations (IC50 > 30 microM). While the processing of H-Ras was very sensitive to inhibition by FTI-277 (IC50 = 100 nM), that of K-Ras4B was highly resistant (IC50 = 10 microM). In contrast, we found the processing of K-Ras4B to be much more sensitive to GGTI-286 (IC50 = 2 microM). Furthermore, oncogenic K-Ras4B stimulation inhibited potently by GGTI-286 (IC50 = 1 microM) but weakly by FTI-277 (IC50 = 30 microM). Significant inhibition of oncogenic K-Ras4B stimulation of MAP kinase by GGTI-286 occurred at concentrations (1-3 microM) that did not inhibit oncogenic H-Ras stimulation of MAP kinase. The data presented in this study provide the first demonstration of selective disruption of oncogenic K-Ras4B processing and signaling by a CAAX peptidomimetic. The higher sensitivity of K-Ras4B toward a GGTase I inhibitor has a tremendous impact on future research directions targeting Ras in anticancer therapy.

3T3 Cells↗
PubMed1995

Ras CAAX peptidomimetic FTI-277 selectively blocks oncogenic Ras signaling by inducing cytoplasmic accumulation of inactive Ras-Raf complexes.

Ras-induced malignant transformation requires Ras farnesylation, a lipid posttranslational modification catalyzed by farnesyltransferase (FTase). Inhibitors of this enzyme have been shown to block Ras-dependent transformation, but the mechanism by which this occurs remains largely unknown. We have designed FTI-276, a peptide mimetic of the COOH-terminal Cys-Val-Ile-Met of K-Ras4B that inhibited potently FTase in vitro (IC50 = 500 pM) and was highly selective for FTase over geranylgeranyltransferase I (GGTase I) (IC50 = 50 nM). FTI-277, the methyl ester derivative of FTI-276, was extremely potent (IC50 = 100 nM) at inhibiting H-Ras, but not the geranylgeranylated Rap1A processing in whole cells. Treatment of H-Ras oncogene-transformed NIH 3T3 cells with FTI-277 blocked recruitment to the plasma membrane and subsequent activation of the serine/threonine kinase c-Raf-1 in cells transformed by farnesylated Ras (H-RasF), but not geranylgeranylated, Ras (H-RasGG). FTI-277 induced accumulation of cytoplasmic non-farnesylated H-Ras that was able to bind Raf and form cytoplasmic Ras/Raf complexes in which Raf kinase was not activated. Furthermore, FTI-277 blocked constitutive activation of mitogen-activated protein kinase (MAPK) in H-RasF, but not H-RasGG, or Raf-transformed cells. FTI-277 also inhibited oncogenic K-Ras4B processing and constitutive activation of MAPK, but the concentrations required were 100-fold higher than those needed for H-Ras inhibition. The results demonstrate that FTI-277 blocks Ras oncogenic signaling by accumulating inactive Ras/Raf complexes in the cytoplasm, hence preventing constitutive activation of the MAPK cascade.

Alkyl and Aryl Transferases↗
PubMed1995

Ras CAAX peptidomimetic FTI 276 selectively blocks tumor growth in nude mice of a human lung carcinoma with K-Ras mutation and p53 deletion.

Farnesylation of the oncoprotein Ras is required for its cancer-causing activity. We have designed farnesyltransferase inhibitor (FTI)-276, a tetrapeptide mimetic of the carboxyl terminus of K-Ras4B, as a highly potent and selective inhibitor of Ras farnesylation in vitro and in vivo. FTI-276 blocked the growth in nude mice of a human lung carcinoma that expresses the two most prevalent genetic alterations in human cancers (K-Ras oncogenic mutation and deletion in the tumor suppressor gene p53). In contrast, FTI-276 did not inhibit tumor growth of a human lung carcinoma that harbors no Ras mutations. Furthermore, FTI-276 inhibited oncogenic signaling and tumor growth of NIH 3T3 cells transformed with the ras but not the raf oncogene. Inhibition of tumor growth in vivo was dose dependent and correlated with inhibition of Ras processing in tumors in vivo. The work described here identifies FTI-276 as a highly selective suppressor of Ras-dependent oncogenicity and suggests that a broad spectrum of human cancers with aberrant Ras function could benefit from farnesyltransferase inhibitor treatment.

3T3 Cells↗