RESUMO
The mitogen-activated protein kinase pathway is thought to be essential in cellular growth and differentiation. Here we report the discovery of a highly potent and selective inhibitor of the upstream kinase MEK that is orally active. Tumor growth was inhibited as much as 80% in mice with colon carcinomas of both mouse and human origin after treatment with this inhibitor. Efficacy was achieved with a wide range of doses with no signs of toxicity, and correlated with a reduction in the levels of activated mitogen-activated protein kinase in excised tumors. These data indicate that MEK inhibitors represent a promising, noncytotoxic approach to the clinical management of colon cancer.
Assuntos
Benzamidas/farmacologia , Ciclo Celular/efeitos dos fármacos , Neoplasias do Colo/patologia , Neoplasias do Colo/fisiopatologia , Inibidores Enzimáticos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Benzamidas/uso terapêutico , Caderinas/análise , Proteínas Quinases Dependentes de Cálcio-Calmodulina/antagonistas & inibidores , Divisão Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/enzimologia , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/uso terapêutico , Feminino , Fator de Crescimento de Hepatócito/farmacologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos , Camundongos Nus , Invasividade Neoplásica/prevenção & controle , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
We investigated the constitutive activation of the MEK/ERK pathway in acute myelogenous leukemia (AML) via a flow cytometric technique to quantitate expression of phosphorylated ERK (p-ERK). A total of 42 AML samples (16 newly diagnosed, 26 relapsed/refractory) were analyzed. Normal bone marrow CD34+ cells (n = 10) had little or no expression of p-ERK, while G-CSF-mobilized CD34+ cells exhibited enhanced p-ERK levels. Markedly elevated p-ERK levels were found in 83.3% of the AML samples, with no differences observed between the newly diagnosed and relapsed/refractory samples. Treatment with a MEK inhibitor resulted in significantly decreased p-ERK levels in both the newly diagnosed and relapsed/refractory samples, which was associated with growth arrest, but not apoptosis induction. In summary, we defined conditions for the analysis of MAPK signaling in primary AML samples. Normal CD34+ cells expressed very low levels of p-ERK, and increased p-ERK levels were found in normal G-CSF-stimulated circulating CD34+ cells. Constitutively high p-ERK levels observed in the majority of AML samples suggest deregulation of this pathway that appears to be independent of disease status. The ability of ERK inhibition to promote growth arrest rather than apoptosis suggests that clinical trials of MEK/ERK inhibitors may be more effective when combined with chemotherapy.
Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Leucemia Mieloide/metabolismo , Doença Aguda , Antígenos CD34/metabolismo , Apoptose/efeitos dos fármacos , Benzamidas/farmacologia , Linhagem Celular Tumoral , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , Citometria de Fluxo , Humanos , Fosforilação , RecidivaRESUMO
Since the discovery of the role of ras oncogenes in tumorigenesis, we have witnessed an explosion of research in the signal transduction area. In the quest to understand how Ras transmits extracellular growth signals, the MAP kinase (MAPK) pathway has emerged as the crucial route between membrane-bound Ras and the nucleus. The MAPK pathway encompasses a cascade of phosphorylation events involving three key kinases, namely Raf, MEK (MAP kinase kinase) and ERK (MAP kinase). This kinase cascade presents novel opportunities for the development of new cancer therapies designed to be less toxic than conventional chemotherapeutic drugs. Furthermore, as a signal transduction-based approach to cancer treatment, inhibition of any one of these targets has the potential for translational pharmacodynamic evaluation of target suppression. The rationale for targeting the MAP kinase pathway will be reviewed here along with a discussion of various pharmacological approaches and the promise they hold for a new generation of anticancer drugs.
Assuntos
Antineoplásicos/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Animais , Benzamidas/uso terapêutico , Divisão Celular/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Desenho de Fármacos , Previsões , Humanos , MAP Quinase Quinase 1 , MAP Quinase Quinase 2 , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-raf/antagonistas & inibidoresRESUMO
A specific set of proteins in eukaryotic cells contain covalently attached carboxy-terminal prenyl groups (15-carbon farnesyl and 20-carbon geranylgeranyl). Many of them are signaling proteins including Ras, heterotrimeric G proteins and Rab proteins. The protein prenyltransferases which attach prenyl groups to proteins have been well characterized, and an X-ray structure is available for protein farnesyltransferase. Inhibitors of protein farnesyltransferase are showing sufficient promise in preclinical trials as anti-cancer drugs to warrant widespread interest in the pharmaceutical industry.
Assuntos
Alquil e Aril Transferases/metabolismo , Antineoplásicos/metabolismo , Prenilação de Proteína , Antineoplásicos/química , HumanosRESUMO
Pyrazoloacridine (PA), an acridine congener with an unknown mechanism of action, has shown selective activity against solid tumor cells, cytotoxicity in noncycling and hypoxic cells, and promising antitumor activity in Phase I clinical trials. In the present study, the effect of PA on topoisomerase (topo) activity was evaluated using yeast strains lacking functional topo I or II, mammalian cell nuclear extracts, purified samples of mammalian topo I and topo II, and intact mammalian tissue culture cells. Clonogenic assays revealed that PA cytotoxicity in yeast strains was unaffected by selective loss of topo I or topo II activity. On the other hand, enzyme assays revealed that 2-4 microM PA abolished the catalytic activity of both topo I and topo II in vitro. In contrast to topotecan and etoposide, PA did not stabilize covalent topo-DNA complexes. Instead, PA inhibited topotecan-induced stabilization of covalent topo I-DNA complexes and etoposide-induced stabilization of topo II-DNA complexes in vitro and in intact cells. Consistent with these results, colony-forming assays indicated that short-term PA exposure inhibited the cytotoxicity of topotecan and etoposide, whereas prolonged PA exposure was itself toxic to these cells. Accumulation studies revealed that PA was concentrated as much as 250-fold in drug-treated cells, resulting in intranuclear concentrations that far exceeded those required to inhibit topo I and topo II. Collectively, these results not only suggest that PA can target both topo I and topo II at clinically achievable concentrations but also indicate that its mechanism is distinct from topo I and topo II poisons presently licensed for clinical use.
Assuntos
Acridinas/farmacologia , Acridinas/farmacocinética , Antineoplásicos/farmacologia , Antineoplásicos/farmacocinética , Pirazóis/farmacologia , Pirazóis/farmacocinética , Inibidores da Topoisomerase I , Inibidores da Topoisomerase II , Acridinas/toxicidade , Antineoplásicos/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/toxicidade , Etoposídeo/toxicidade , Humanos , Leucemia Mieloide , Pirazóis/toxicidade , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/crescimento & desenvolvimento , Frações Subcelulares/metabolismo , Células Tumorais CultivadasRESUMO
The development of farnesyltransferase inhibitors (FTIs) has been one of the most active areas of anticancer drug development for the past ten years. This review presents a general overview of the developments in this area, along with a critical appraisal of the anticancer activity of FTIs. A historical survey of the protein prenylation field is given, in particular to emphasize the key role played by the Ras oncoprotein in driving the discovery of prenyltransferase enzymes. The different classes of prenylated proteins will be described along with the biochemical characteristics of the key drug target--farnesyltransferase (FTase). Numerous potent farnesyltransferase inhibitors have been developed. The FTIs developed can be separated into three different categories, based on their origin and/or mechanism of action: a) natural products; b) peptidomimetics and other CAAX-competitive inhibitors; c) farnesyl pyrophosphate (FPP) mimetics or analogs and other FPP-competitive inhibitors. Along with a survey of newer FTIs in each class, the development of several representative, potent compounds will be discussed in depth as we discuss the potential advantages and liabilities of each class. Particular emphasis is given to the discovery of new, more potent FPP-competitive FTIs of several diverse structural classes. Testing of different FTIs for their ability to block the growth of various cancer cell types in animal models will be discussed. There are a number of key differences between these compounds and traditional cytotoxic cancer chemotherapeutic agents, with surprising exceptions to their expected modes of action. As some FTIs have entered human clinical trials, answers may soon become available to key mechanistic questions concerning the extent and nature of their antitumor growth properties.
Assuntos
Antineoplásicos/farmacologia , Dimetilaliltranstransferase/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Neoplasias/tratamento farmacológico , Ensaios Clínicos como Assunto , Humanos , Neoplasias/metabolismo , Neoplasias/patologiaRESUMO
The ability of DNA repair inhibitors to potentiate alkylating agent cytotoxicity was explored with PD 128763, a dihydroisoquinolinone known to effectively inhibit poly(ADP-ribose) synthetase. The cytotoxic activity of streptozotocin in L1210 leukemia cells was maximally potentiated (7-fold decrease in IC50) under conditions of 24 hr exposure to PD 128763 following treatment with the alkylating agent for 1 hr. Similar treatment conditions resulted in a much greater effect (36-fold enhancement in activity) for the 2-nitroimidazole RSU 1069. In contrast, 3-aminobenzamide was only weakly effective at enhancing activity of either streptozotocin or RSU 1069 (2-3 fold potentiation). However, PD 128763 was ineffective at potentiating the cytotoxicity of the bifunctional alkylating agents carmustine (BCNU) and lomustine (CCNU). Our results are consistent with a role for (poly-ADP) ribosylation in the repair of monofunctional alkylating agent damage. This study supports further exploration of the combination of PD 128763 and RSU 1069 as a potentially useful chemotherapeutic regimen.
Assuntos
Alquilantes/farmacologia , Isoquinolinas/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases , Animais , Carmustina/farmacologia , Sinergismo Farmacológico , Técnicas In Vitro , Leucemia L1210 , Misonidazol/análogos & derivados , Misonidazol/farmacologia , Estreptozocina/farmacologia , Células Tumorais CultivadasRESUMO
Following the toxicity and synthetic difficulties encountered with the hypoxic cell radiosensitizer RSU 1069, efforts have focused on development of a superior analogue. Two compounds, RB 6145 and PD 130908, have emerged from this program which overcome the instability and synthetic problems associated with RSU 1069 while retaining favorable biological activity. Both agents show comparable radiosensitizing activity to RSU 1069 following oral or i.p. administration to mice bearing the KHT or RIF-1 tumors. Sensitizing efficiency is about 10 X greater than that observed for misonidazole or etanidazole. Toxicity toward hypoxic tumor cells in vivo is demonstrated by clamping tumors (for 60 min) following administration of PD 130908 or RB 6145. Both are effective hypoxic cytotoxins, but less potent than RSU 1069. Systemic toxicity is substantially reduced following oral drug administration. Further, doses achievable following fractionated drug treatments are sufficiently high to produce significant levels of radiosensitization.
Assuntos
Misonidazol/análogos & derivados , Nitroimidazóis/uso terapêutico , Radiossensibilizantes/uso terapêutico , Sarcoma Experimental/radioterapia , Animais , Hipóxia Celular/efeitos dos fármacos , Hipóxia Celular/fisiologia , Hipóxia Celular/efeitos da radiação , Terapia Combinada , Avaliação de Medicamentos , Camundongos , Camundongos Endogâmicos C3H , Misonidazol/uso terapêutico , Misonidazol/toxicidade , Transplante de Neoplasias , Nitroimidazóis/toxicidade , Radiossensibilizantes/toxicidade , Sarcoma Experimental/tratamento farmacológicoRESUMO
RB 6145, the ring-opened analog of RSU 1069, and PD 130908, the desoxy ring-opened analog of RSU 1069, were compared to RSU 1069 for their emetic potential in dogs. When RB 6145 and PD 130908 were administered intravenously at doses ranging from 20% to 50% of the mouse equivalent maximum tolerated dose (MTD), both analogs were less emetic than RSU 1069 on a molar basis. Furthermore, the 5HT3 antagonist ondansetron prevented emesis at doses as high as 75% of the MTD. The reactivity, and hence the emetic liability of these compounds, is thought to be mediated by formation of the corresponding aziridine intermediate. In mouse plasma, both analogs rapidly converted to two products, the reactive aziridine and a stable oxiazolidinone species formed upon reaction with bicarbonate in the blood. A positive correlation exists between the amounts of aziridine formed by these analogs and their emetic potential.
Assuntos
Antieméticos/uso terapêutico , Misonidazol/análogos & derivados , Nitroimidazóis/toxicidade , Radiossensibilizantes/toxicidade , Vômito/induzido quimicamente , Animais , Cães , Avaliação de Medicamentos , Imidazóis/uso terapêutico , Camundongos , Misonidazol/sangue , Misonidazol/farmacocinética , Misonidazol/toxicidade , Nitroimidazóis/sangue , Nitroimidazóis/farmacocinética , Ondansetron , Radiossensibilizantes/farmacocinética , Vômito/prevenção & controleRESUMO
Protein farnesyltransferase (FTase), the enzyme responsible for protein farnesylation, has become a key target for the rational design of cancer chemotherapeutic agents. Herein it is shown that certain novel prenyl diphosphate analogues are potent inhibitors of mammalian FTase. Furthermore, the alcohol precursors of two of these compounds are able to block anchorage-independent growth of ras-transformed cells. While 3-allylfarnesol inhibits protein farnesylation, 3-vinylfarnesol instead leads to abnormal prenylation of proteins with the 3-vinylfarnesyl group. In a similar manner, 3-allylgeranylgeraniol acts as a highly specific inhibitor of protein geranylgeranylation, while 3-vinylgeranylgeraniol restores protein geranylgeranylation in cells. This study indicates that certain prenyl alcohol analogues can act as prenyltransferase inhibitors in situ, via a novel prodrug mechanism. These analogues may prove to be valuable tools for investigating the therapeutic consequences of inhibiting geranylgeranylation relative to farnesylation. Furthermore, the 3-vinyl alcohol analogues can inhibit transformed cell growth through a mechanism not involving prenyltransferase inhibition.
Assuntos
Antineoplásicos/farmacologia , Diterpenos/química , Farneseno Álcool/análogos & derivados , Prenilação de Proteína/efeitos dos fármacos , Alquil e Aril Transferases/metabolismo , Animais , Catálise , Eletroforese em Gel de Poliacrilamida , Cinética , Modelos Químicos , Células Tumorais CultivadasRESUMO
A series of 3,5-disubstituted-2,1-benzisoxazole-4,7-diones was synthesized and evaluated as radiosensitizers both in vitro and in vivo. These compounds were designed as non-nitro electron-affinic agents in an effort to alleviate some of the toxicities seen with the 2-nitroimidazole radiosensitizers evaluated in the clinic. Several compounds in this series were potent radiosensitizers in vitro, with sensitizer enhancement ratios of 2.0-2.3 at concentrations less than 0.5 mM. Compounds with potent in vitro activity were also evaluated in vivo. However, none of these compounds showed radiosensitizing activity in vivo. The reduction potentials of these compounds were determined by cyclic voltammetry and compared to other electron-affinic radiosensitizers. In general, the reduction potentials of this series of compounds was slightly more positive than the 2-nitroimidazoles, but they fell within the range postulated as acceptable to yield in vivo activity. The results suggest that factors other than reduction potential may be responsible for the lack of in vivo radiosensitizing activity observed for this class of radiosensitizers.
Assuntos
Isoxazóis/síntese química , Radiossensibilizantes/síntese química , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cricetinae , Cricetulus , Isoxazóis/farmacologia , Camundongos , Radiossensibilizantes/farmacologia , Relação Estrutura-AtividadeRESUMO
In vitro screening of a number of 2-(aminoalkyl)-5-nitropyrazolo[3,4,5- kl]acridines has previously indicated (Sebolt, J.S.; et al. Cancer Res. 1987, 47, 4299-4304) that these compounds, in general, exhibit selective cytotoxicity against the human colon adenocarcinoma, HCT-8, cell line, relative to mouse leukemia L1210 cells. Comparative molecular field analysis (CoMFA) was applied to HCT-8 and L1210 growth inhibition assays (IC50s) of a series (44) of the pyrazoloacridine derivatives with the objective of predicting improved solid tumor selectivity. In the absence of crystallographic data, the 9-methoxy derivative (15), which is currently in clinical study, was selected as the template molecular model. Two different structural alignments were tested: an alignment of structures based on root mean square (RMS)-fitting of each structure to 15 was compared with an alternative strategy, steric and electrostatic alignment (SEAL). Somewhat better predictive cross-validation correlations (r2) were obtained with models based on RMS vis-à-vis SEAL alignment for both sets of assays. A large change in lattice spacing, e.g., 2 to 1 A, causes significant variations in the CoMFA results. A shift in the lattice of half of its spacing had a much smaller effect on the CoMFA data for a lattice of 1 A than one of 2 A. The relative contribution of steric and electrostatic fields to both models were about equal, underscoring the importance of both terms. Neither calculated log P nor HOMO and/or LUMO energies contribute to the model. Steric and electrostatic fields of the pyrazoloacridines are the sole relevant descriptors to the structure-activity (cross-validated and conventional) correlations obtained with the cytotoxic data for both the L1210 and HCT-8 cell lines. The cross-validated r2, derived from partial least-squares calculations, indicated considerable predictive capacity for growth inhibition of both the leukemia and solid-tumor data. Evidence for the predictive performance of the CoMFA-derived models is provided in the form of plots of actual vs predicted growth inhibition of L1210 and HCT-8 cells, respectively, by the pyrazoloacridines. The steric and electrostatic features of the QSAR are presented in the form of standard deviation coefficient contour maps of steric and electrostatic fields. The maps indicate that increases or decreases in steric bulk that would enhance growth inhibition of HCT-8 cells would likewise promote growth inhibition of L1210 cells. Contour maps generated to analyze the electrostatic field contributions of the pyrazoloacridines to growth inhibition provide an essentially similar set of results.(ABSTRACT TRUNCATED AT 400 WORDS)
Assuntos
Acridinas/uso terapêutico , Adenocarcinoma/tratamento farmacológico , Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Leucemia L1210/tratamento farmacológico , Pirazóis/uso terapêutico , Acridinas/química , Animais , Eletroquímica , Humanos , Substâncias Intercalantes , Camundongos , Modelos Moleculares , Estrutura Molecular , Pirazóis/química , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
A series of compounds related to alpha-(1-aziridinylmethyl)-2-nitro-1H-imidazole-1-ethanol (RSU 1069, 1) were synthesized and evaluated as selective hypoxic cell cytotoxic agents and as radiosensitizers. The aziridine moiety was replaced with a number of other potential alkylating groups including cycloalkylaziridines and azetidines. The data indicated that modification of the aziridine of 1 resulted in a substantial decrease in the ability of the compounds to selectively kill hypoxic cells. However, these modifications did not affect the compounds' in vitro radiosensitizing activity since many of the derivatives were as potent as 1. All of the compounds that were evaluated in vivo were less toxic than 1, and several members of this series had significant activity. The best compound was trans-alpha-[[(4-bromotetrahydro-2H-pyran-3-yl) amino]methyl]-2-nitro-1H-imidazole-1-ethanol (18), which, due to its activity and log P value, is a candidate for additional in vivo studies.
Assuntos
Aziridinas/síntese química , Misonidazol/análogos & derivados , Nitroimidazóis/síntese química , Radiossensibilizantes/síntese química , Animais , Aziridinas/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Fenômenos Químicos , Química , Cricetinae , Fibrossarcoma/radioterapia , Camundongos , Misonidazol/química , Misonidazol/farmacologia , Estrutura Molecular , Nitroimidazóis/farmacologia , Nitroimidazóis/uso terapêutico , Oxigênio/administração & dosagem , Piranos/síntese química , Piranos/farmacologia , Piranos/uso terapêutico , Radiossensibilizantes/farmacologia , Radiossensibilizantes/uso terapêutico , Relação Estrutura-AtividadeRESUMO
Mutational activation of ras has been found in many types of human cancers, including a greater than 50% incidence in colon and about 90% in pancreatic carcinomas. The activity of both native and oncogenic ras proteins requires a series of post-translational processing steps. The first event in this process is the farnesylation of a cysteine residue located in the fourth position from the carboxyl terminus of the ras protein, catalyzed by the enzyme farnesyltransferase (FTase). Inhibitors of FTase are potential candidates for development as antitumor agents. Through a high-volume screening program, the pentapeptide derivative PD083176 (1), Cbz-His-Tyr(OBn)-Ser(OBn)-Trp-DAla-NH2, was identified as an inhibitor of rat brain FTase, with an IC50 of 20 nM. Structure-activity relationships were carried out to determine the importance of the side chain and chirality of each residue. This investigation led to a series of potent FTase inhibitors which lack a cysteine residue as found in the ras peptide substrate. The parent compound (1) inhibited the insulin-induced maturation of Xenopus oocytes (concentration: 5 pmol/oocyte), a process which is dependent on the activation of the ras pathway.
Assuntos
Alquil e Aril Transferases , Cisteína/química , Cisteína/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Oligopeptídeos/síntese química , Oligopeptídeos/farmacologia , Transferases/antagonistas & inibidores , Aminoácidos/química , Animais , Sítios de Ligação , Antagonistas da Insulina/farmacologia , Oócitos/citologia , Oócitos/efeitos dos fármacos , Fosfatos/química , Ratos , Relação Estrutura-Atividade , XenopusRESUMO
The modifying effects of PD 128763 (3,4-dihydro-5-methyl-1(2H)-isoquinolinone), a potent inhibitor of poly(adenosine-diphosphate (ADP)-ribose) polymerase, on radiation-induced cell killing were examined in Chinese hamster V79 cells. This compound has an IC50 value against the purified enzyme approximately 50X lower than 3-aminobenzamide (3-AB), a widely used specific inhibitor of the enzyme. Exposure of exponentially growing cells to a noncytotoxic concentration (0.5 mM) of PD 128763 for 2 h immediately following X irradiation increased their radiation sensitivity, modifying both the shoulder and the slope of the survival curve. When recovery from sublethal damage and potentially lethal damage was examined in exponential and plateau-phase cells, respectively, postirradiation incubation with 0.5 mM PD 128763 was found not only to inhibit both these processes fully, but also to enhance further the level of radiation-induced cell killing. This is in contrast to the slight effect seen with the less potent inhibitor, 3-AB. The results presented suggest that the mechanism of radiosensitization by PD 128763 is related to the potent inhibition of poly(ADP-ribose) polymerase by this compound.
Assuntos
Sobrevivência Celular/efeitos dos fármacos , Isoquinolinas/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases , Animais , Benzamidas/farmacologia , Sobrevivência Celular/efeitos da radiação , Células Cultivadas/efeitos da radiação , Cricetinae , CricetulusAssuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antineoplásicos/síntese química , Dipeptídeos/síntese química , Inibidores Enzimáticos/síntese química , Proteínas ras/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular , Dipeptídeos/química , Dipeptídeos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Camundongos , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Prenilação de Proteína , Ratos , Células Tumorais CultivadasRESUMO
A series of dihydroisoquinolinones, formally rigid analogs of 3-substituted benzamides, and a series of 2,3-disubstituted benzamides were synthesized and evaluated as inhibitors of poly(ADP-ribose) polymerase. The results indicated that the orientation of the amide with respect to the substituent on the aromatic ring was critical for optimum inhibitory activity. Selected compounds were also evaluated for their ability to modify the radiation response of mammalian cells to ionizing radiation. A number of the 5-substituted dihydroisoquinolinones, which were very potent inhibitors of the enzyme, were able to enhance the lethal effects of ionizing radiation in mammalian cells, as measured by changes in the survival curve parameters Do and/or Dq.
Assuntos
Benzamidas/síntese química , Isoquinolinas/síntese química , Inibidores de Poli(ADP-Ribose) Polimerases , Animais , Benzamidas/química , Linhagem Celular/efeitos dos fármacos , Linhagem Celular/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Cricetinae , Desenho de Fármacos , Isoquinolinas/química , Radiossensibilizantes/síntese química , Relação Estrutura-AtividadeRESUMO
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 how a prenyl group is enzymatically 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.
Assuntos
Alquil e Aril Transferases , Antineoplásicos/química , Inibidores Enzimáticos/química , Prenilação de Proteína , Transferases/antagonistas & inibidores , Magnésio/metabolismo , Fosfatos de Poli-Isoprenil/química , Sesquiterpenos , Zinco/metabolismoRESUMO
Limonene and its metabolite perillyl alcohol are naturally-occurring isoprenoids that block the growth of cancer cells both in vitro and in vivo. This cytostatic effect appears to be due, at least in part, to the fact that these compounds are weak yet selective and non-toxic inhibitors of protein prenylation. Protein-farnesyl transferase (FTase), the enzyme responsible for protein farnesylation, has become a key target for the rational design of cancer chemotherapeutic agents. Therefore, several alpha-hydroxyphosphonate derivatives of limonene were designed and synthesized as potentially more potent FTase inhibitors. A noteworthy feature of the synthesis was the use of trimethylsilyl triflate as a mild, neutral deprotection method for the preparation of sensitive phosphonates from the corresponding tert-butyl phosphonate esters. Evaluation of these compounds demonstrates that they are exceptionally poor FTase inhibitors in vitro (IC50 > or = 3 mM) and they have no effect on protein farnesylation in cells. In contrast, farnesyl phosphonyl(methyl)phosphinate, a diphosphate-modified derivative of the natural substrate farnesyl diphosphate, is a very potent FTase inhibitor in vitro (Ki=23 nM).
Assuntos
Fosfatos de Poli-Isoprenil/síntese química , Terpenos/síntese química , Alquil e Aril Transferases/antagonistas & inibidores , Animais , Antineoplásicos Fitogênicos/farmacologia , Cicloexenos , Relação Dose-Resposta a Droga , Concentração Inibidora 50 , Cinética , Limoneno , Modelos Químicos , Fosfatos de Poli-Isoprenil/farmacologia , Sesquiterpenos , Terpenos/farmacologiaRESUMO
Investigation of the comparative activities of various inhibitors of farnesyl:protein transferase (FPTase) has led to the observation that the presence of phosphate or pyrophosphate ions in the assay buffer increases the potency of farnesyl diphosphate (FPP) competitive inhibitors. In addition to exploring the phenomenon of phosphate synergy, we report here the effects of various other ions including sulfate, bicarbonate, and chloride on the inhibitory ability of three FPP competitive compounds: Cbz-His-Tyr-Ser(OBn)TrpNH2 (2), Cbz-HisTyr(OPO42-)-Ser(OBn)TrpNH2 (3), and alpha-hydroxyfarnesyl phosphonic acid (4). Detailed kinetic analysis of FPTase inhibition revealed a high degree of synergy for compound 2 and each of these ions. Phosphorylation of 2 to give 3 completely eliminated any ionic synergistic effect. Moreover, these ions have an antagonistic effect on the inhibitory potency of compound 4. The anions in the absence of inhibitor exhibit non-competitive inhibition with respect to FPP. These results suggest that phosphate, pyrophosphate, bicarbonate, sulfate, and chloride ions may be binding at the active site of both free enzyme and product-bound enzyme with normal substrates. These bound complexes increase the potency of FPP competitive inhibitors and mimic an enzyme:product form of the enzyme. None of the anions studied here proved to be synergistic with respect to inhibition of geranylgeranyl transferase I. These findings provide insight into the mechanism of action of FPP competitive inhibitors for FPTase and point to enzymatic differences between FPTase and geranylgeranyl transferase I that may facilitate the design of more potent and specific inhibitors for these therapeutically relevant target enzymes.