RESUMO
The role of hypoxic tumour cells in resistance to radiotherapy, and in suppression of immune response, continues to endorse tumour hypoxia as a bona fide, yet largely untapped, drug target. Radiotherapy innovations such as stereotactic body radiotherapy herald new opportunities for classical oxygen-mimetic radiosensitisers. Only nimorazole is used clinically as a radiosensitiser, and there is a dearth of new radiosensitisers in development. In this report, we augment previous work to present new nitroimidazole alkylsulfonamides and we document their cytotoxicity and ability to radiosensitise anoxic tumour cells in vitro. We compare radiosensitisation with etanidazole and earlier nitroimidazole sulfonamide analogues and we identify 2-nitroimidazole and 5-nitroimidazole analogues with marked tumour radiosensitisation in ex vivo assays of surviving clonogens and with in vivo tumour growth inhibition.
Assuntos
Neoplasias , Nitroimidazóis , Radiossensibilizantes , Humanos , Hipóxia Celular , Nitroimidazóis/farmacologia , Radiossensibilizantes/farmacologia , Hipóxia , Neoplasias/tratamento farmacológico , Neoplasias/radioterapiaRESUMO
Hypoxia in tumors results in resistance to both chemotherapy and radiotherapy treatments but affords an environment in which hypoxia-activated prodrugs (HAP) are activated upon bioreduction to release targeted cytotoxins. The benzotriazine 1,4-di-N-oxide (BTO) HAP, tirapazamine (TPZ, 1), has undergone extensive clinical evaluation in combination with radiotherapy to assist in the killing of hypoxic tumor cells. Although compound 1 did not gain approval for clinical use, it has spurred on the development of other BTOs, such as the 3-alkyl analogue, SN30000, 2. There is general agreement that the cytotoxin(s) from BTOs arise from the one-electron reduced form of the compounds. Identifying the cytotoxic radicals, and whether they play a role in the selective killing of hypoxic tumor cells, is important for continued development of the BTO class of anticancer prodrugs. In this study, nitrone spin-traps, combined with electron spin resonance, give evidence for the formation of aryl radicals from compounds 1, 2 and 3-phenyl analogues, compounds 3 and 4, which form carbon C-centered radicals. In addition, high concentrations of DEPMPO (5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide) spin-trap the â¢OH radical. The combination of spin-traps with high concentrations of DMSO and methanol also give evidence for the involvement of strongly oxidizing radicals. The failure to spin-trap methyl radicals with PBN (N-tert-butylphenylnitrone) on the bioreduction of compound 2, in the presence of DMSO, implies that free â¢OH radicals are not released from the protonated radical anions of compound 2. The spin-trapping of â¢OH radicals by high concentrations of DEPMPO, and the radical species arising from DMSO and methanol give both direct and indirect evidence for the scavenging of â¢OH radicals that are involved in an intramolecular process. Hypoxia-selective cytotoxicity is not related to the formation of aryl radicals from the BTO compounds as they are associated with high aerobic cytotoxicity.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Triazinas/química , Triazinas/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Elétrons , Radicais Livres/química , Células HCT116 , Células HT29 , Humanos , Radical Hidroxila/química , Neoplasias/tratamento farmacológico , Detecção de SpinRESUMO
Hypoxia is an adverse prognostic feature of solid cancers that may be overcome with hypoxia-activated prodrugs (HAPs). Tirapazamine (TPZ) is a HAP which has undergone extensive clinical evaluation in this context and stimulated development of optimized analogues. However the subcellular localization of the oxidoreductases responsible for mediating TPZ-dependent DNA damage remains unclear. Some studies conclude only nuclear-localized oxidoreductases can give rise to radical-mediated DNA damage and thus cytotoxicity, whereas others identify a broader role for endoplasmic reticulum and cytosolic oxidoreductases, indicating the subcellular location of TPZ radical formation is not a critical requirement for DNA damage. To explore this question in intact cells we engineered MDA-231 breast cancer cells to express the TPZ reductase human NADPH: cytochrome P450 oxidoreductase (POR) harboring various subcellular localization sequences to guide this flavoenzyme to the nucleus, endoplasmic reticulum, cytosol or inner surface of the plasma membrane. We show that all POR variants are functional, with differences in rates of metabolism reflecting enzyme expression levels rather than intracellular TPZ concentration gradients. Under anoxic conditions, POR expression in all subcellular compartments increased the sensitivity of the cells to TPZ, but with a fall in cytotoxicity per unit of metabolism (termed 'metabolic efficiency') when POR is expressed further from the nucleus. However, under aerobic conditions a much larger increase in cytotoxicity was observed when POR was directed to the nucleus, indicating very high metabolic efficiency. Consequently, nuclear metabolism results in collapse of hypoxic selectivity of TPZ, which was further magnified to the point of reversing O2 dependence (oxic > hypoxic sensitivity) by employing a DNA-affinic TPZ analogue. This aerobic hypersensitivity phenotype was partially rescued by cellular copper depletion, suggesting the possible involvement of Fenton-like chemistry in generating short-range effects mediated by the hydroxyl radical. In addition, the data suggest that under aerobic conditions reoxidation strictly limits the TPZ radical diffusion range resulting in site-specific cytotoxicity. Collectively these novel findings challenge the purported role of intra-nuclear reductases in orchestrating the hypoxia selectivity of TPZ.
Assuntos
Antineoplásicos/química , Hipóxia/tratamento farmacológico , NADPH-Ferri-Hemoproteína Redutase/genética , Pró-Fármacos/química , Tirapazamina/química , Antineoplásicos/farmacologia , Engenharia Celular , Hipóxia Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cobre/metabolismo , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Humanos , Modelos Biológicos , NADPH-Ferri-Hemoproteína Redutase/metabolismo , NADPH-Ferri-Hemoproteína Redutase/ultraestrutura , Oxigênio/metabolismo , Pró-Fármacos/metabolismo , Tirapazamina/metabolismoRESUMO
The development of an improved short and efficient commercial synthesis of the JAK2 inhibitor, a complex pyrrolopyridine, BMS-911543, is described. During the discovery and development of this synthesis, a Pd-catalyzed C-H functionalization was invented which enabled the rapid union of the key pyrrole and imidazole fragments. The synthesis of this complex, nitrogen-rich heterocycle was accomplished in only six steps (longest linear sequence) from readily available materials.
Assuntos
Compostos Heterocíclicos com 3 Anéis/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Catálise , Compostos Heterocíclicos com 3 Anéis/síntese química , Compostos Heterocíclicos com 3 Anéis/química , Humanos , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/metabolismo , Ligantes , Estrutura Molecular , Paládio/química , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/químicaRESUMO
Poly(ADP-ribose)polymerase (PARP) inhibitors (PARPi) have recently been approved for the treatment of breast and ovarian tumors with defects in homologous recombination repair (HRR). Although it has been demonstrated that PARPi also sensitize HRR competent tumors to cytotoxic chemotherapies or radiotherapy, normal cell toxicity has remained an obstacle to their use in this context. Hypoxia-activated prodrugs (HAPs) provide a means to limit exposure of normal cells to active drug, thus adding a layer of tumor selectivity. We have investigated potential HAPs of model PARPi in which we attach a bioreducible "trigger" to the amide nitrogen, thereby blocking key binding interactions. A representative example showed promise in abrogating PARPi enzymatic activity in a biochemical assay, with a ca. 160-fold higher potency of benzyl phthalazinone 4 than the corresponding model HAP 5, but these N-alkylated compounds did not release the PARPi upon one-electron reduction by radiolysis. Therefore, we extended our investigation to include NU1025, a PARPi that contains a phenol distal to the core binding motif. The resulting 2-nitroimidazolyl ether provided modest abrogation of PARPi activity with a ca. seven-fold decrease in potency, but released the PARPi efficiently upon reduction. This investigation of potential prodrug approaches for PARPi has identified a useful prodrug strategy for future exploration.
Assuntos
Inibidores de Poli(ADP-Ribose) Polimerases/química , Antineoplásicos/química , Cromatografia Líquida , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Pró-Fármacos/química , Quinazolinas/químicaRESUMO
Extracellular acidification is an important feature of tumor microenvironments but has yet to be successfully exploited in cancer therapy. The reversal of the pH gradient across the plasma membrane in cells that regulate intracellular pH (pHi) has potential to drive the selective uptake of weak acids at low extracellular pH (pHe). Here, we investigate the dual targeting of low pHe and hypoxia, another key feature of tumor microenvironments. We prepared eight bioreductive prodrugs based on the benzotriazine di-oxide (BTO) nucleus by appending alkanoic or aminoalkanoic acid sidechains. The BTO acids showed modest selectivity for both low pHe (pH 6.5 versus 7.4, ratios 2 to 5-fold) and anoxia (ratios 2 to 8-fold) in SiHa and FaDu cell cultures. Related neutral BTOs were not selective for acidosis, but had greater cytotoxic potency and hypoxic selectivity than the BTO acids. Investigation of the uptake and metabolism of representative BTO acids confirmed enhanced uptake at low pHe, but lower intracellular concentrations than expected for passive diffusion. Further, the modulation of intracellular reductase activity and competition by the cell-excluded electron acceptor WST-1 suggests that the majority of metabolic reductions of BTO acids occur at the cell surface, compromising the engagement of the resulting free radicals with intracellular targets. Thus, the present study provides support for designing bioreductive prodrugs that exploit pH-dependent partitioning, suggesting, however, that that the approach should be applied to prodrugs with obligate intracellular activation.
Assuntos
Hipóxia Celular/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Neoplasias/metabolismo , Pró-Fármacos , Triazinas/química , Triazinas/farmacologia , Linhagem Celular Tumoral , Fenômenos Químicos , Relação Dose-Resposta a Droga , Desenho de Fármacos , Humanos , Modelos Biológicos , Estrutura Molecular , Oxirredução/efeitos dos fármacos , ÓxidosRESUMO
We describe a new library generation method, Machine-based Identification of Molecules Inside Characterized Space (MIMICS), that generates sets of molecules inspired by a text-based input. MIMICS-generated libraries were found to preserve distributions of properties while simultaneously increasing structural diversity. Newly identified MIMICS-generated compounds were found to be bioactive as inhibitors of specific components of the unfolded protein response (UPR) and the VEGFR2 pathway in cell-based assays, thus confirming the applicability of this methodology toward drug design applications. Wider application of MIMICS could facilitate the efficient utilization of chemical space.
Assuntos
Descoberta de Drogas/métodos , Redes Neurais de Computação , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidoresRESUMO
The radical chemistry and cytotoxicity of a series of quinoxaline di-N-oxide (QDO) compounds has been investigated to explore the mechanism of action of this class of bioreductive drugs. A series of water-soluble 3-trifluoromethyl (4-10), 3-phenyl (11-19), and 3-methyl (20-21) substituted QDO compounds were designed to span a range of electron affinities consistent with bioreduction. The stoichiometry of loss of QDOs by steady-state radiolysis of anaerobic aqueous formate buffer indicated that one-electron reduction of QDOs generates radicals able to initiate chain reactions by oxidation of formate. The 3-trifluoromethyl analogues exhibited long chain reactions consistent with the release of the HO(â¢), as identified in EPR spin trapping experiments. Several carbon-centered radical intermediates, produced by anaerobic incubation of the QDO compounds with N-terminal truncated cytochrome P450 reductase (POR), were characterized using N-tert-butyl-α-phenylnitrone (PBN) and 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO) spin traps and were observed by EPR. Experimental data were well simulated for the production of strongly oxidizing radicals, capable of H atom abstraction from methyl groups. The kinetics of formation and decay of the radicals produced following one-electron reduction of the parent compounds, both in oxic and anoxic solutions, were determined using pulse radiolysis. Back oxidation of the initially formed radical anions by molecular oxygen did not compete effectively with the breakdown of the radical anions to form oxidizing radicals. The QDO compounds displayed low hypoxic selectivity when tested against oxic and hypoxic cancer cell lines in vitro. The results from this study form a kinetic description and explanation of the low hypoxia-selective cytotoxicity of QDOs against cancer cells compared to the related benzotriazine 1,4-dioxide (BTO) class of compounds.
Assuntos
Quinoxalinas/química , Espectroscopia de Ressonância de Spin Eletrônica , Espectrometria de Massas , NADPH-Ferri-Hemoproteína Redutase/química , Ressonância Magnética Nuclear Biomolecular , Oxirredução , Óxidos/químicaRESUMO
An evolution in radiotherapy practice is leading to greater use of stereotactic body radiotherapy (SBRT), raising the prospect of increased hypoxic cell radioresistance. New clinical interest in nitroimidazole radiosensitisers, combined with appropriate biomarkers, signals a revival for radiosensitisers in the context of SBRT. Our interest in modifiers of radiation therapy led us to revisit this area and we have identified a new class of nitroimidazole radiosensitiser. We have developed an abbreviated screening protocol suitable for an academic drug discovery laboratory which allows expeditious triage of compounds with poor physicochemical and in vitro properties and combines in vitro radiosensitisation data with tumour pharmacokinetic data to efficiently select candidates for further evaluation.
Assuntos
Radiossensibilizantes/análise , Radiossensibilizantes/farmacologia , Animais , Transporte Biológico/efeitos dos fármacos , Biomarcadores/metabolismo , Hipóxia Celular/efeitos dos fármacos , Feminino , Células HCT116 , Humanos , Camundongos Nus , Neoplasias/patologia , Radiossensibilizantes/farmacocinética , RadiocirurgiaRESUMO
Kinetic, spectroscopic, crystallographic, and computational studies probing a Pd-catalyzed C-H arylation reaction reveal that mono-oxidation of the bis-phosphine ligand is critical for the formation of the active catalyst. The bis-phosphine mono-oxide is shown to be a hemilabile, bidentate ligand for palladium. Isolation of the oxidative addition adduct, with structural elucidation by X-ray analysis, showed that the mono-oxide was catalytically competent, giving the same reaction rate in the productive reaction as the Pd(II)/xantphos precursor. A dual role for the carboxylate base in both catalyst activation and reaction turnover was demonstrated, along with the inhibiting effect of excess phosphine ligand. The generality of the role of phosphine mono-oxide complexes in Pd-catalyzed coupling processes is discussed.
Assuntos
Paládio/química , Fosfinas/química , Xantenos/química , Catálise , Cristalografia por Raios X , Cinética , Espectroscopia de Ressonância Magnética , OxirreduçãoRESUMO
In Trifolium repens the decline in bud outgrowth that occurs with distance from basal root systems is due to correlative inhibition by the first-formed basal branches. The apical and axillary buds on these basal branches are the source of the inhibitory effect, but their mode of action is unclear. Inhibition might occur via basal branches being a sink for xylem-transported branching stimulants or alternatively by providing a source of inhibitory signals, or by both mechanisms. To distinguish between these mechanisms, four experiments were conducted on plants varying in initial growth stage from 10 to 19 nodes along their main stems to determine any variation in the relative importance of the operative mechanisms of correlative inhibition. Inhibitory signal exported into the main stem, detected as a branching response to girdling of basal branches, was relatively more significant in smaller (initially with 10-15 nodes on the main stem) than in larger (>19 nodes on main stem) plants. This signal was shown not to involve auxin fluxes, and is unidentified. However, across all stages of growth, the predominant mechanism driving correlative inhibition was the action of axillary and apical buds of basal branches as sinks for the stimulatory signal. This study indicates that the relative importance of the mechanisms regulating bud outgrowth in T. repens varies with growth stage and that, during intermediate stages, regulation has some similarity to that in Pisum.
Assuntos
Brotos de Planta/crescimento & desenvolvimento , Trifolium/crescimento & desenvolvimento , Biomassa , Caules de Planta/crescimento & desenvolvimentoRESUMO
A review of literature published in 2014 focusing on analytical methods for pesticides in the water environment is presented here. The review includes publications on extraction methods and on analytical methods such as biochemical assays and immunoassays, electrochemical methods, chromatographic or mass spectrometric techniques, spectrophotometric techniques, fluorescence, and chemiluminescence techniques.
RESUMO
The radical species underlying the activity of the bioreductive anticancer prodrug, SN30000, have been identified by electron paramagnetic resonance and pulse radiolysis techniques. Spin-trapping experiments indicate both an aryl-type radical and an oxidising radical, trapped as a carbon-centred radical, are formed from the protonated radical anion of SN30000. The carbon-centred radical, produced upon the one-electron oxidation of the 2-electron reduced metabolite of SN30000, oxidises 2-deoxyribose, a model for the site of damage on DNA which leads to double strand breaks. Calculations using density functional theory support the assignments made.
Assuntos
Óxidos N-Cíclicos/química , Radicais Livres/química , Pró-Fármacos/química , Triazinas/química , Hipóxia Celular , Quebras de DNA de Cadeia Dupla , Espectroscopia de Ressonância de Spin Eletrônica , Humanos , Concentração de Íons de Hidrogênio , Cinética , Radiólise de Impulso , Teoria Quântica , Temperatura , TirapazaminaRESUMO
Two novel scaffolds, 4-pyridylanilinothiazoles (PAT) and 3-pyridylphenylsulfonyl benzamides (PPB), previously identified as selective cytotoxins for von Hippel-Lindau-deficient Renal Carcinoma cells, were used as templates to prepare affinity chromatography reagents to aid the identification of the molecular targets of these two classes. Structure-activity data and computational models were used to predict possible points of attachment for linker chains. In the PAT class, Click coupling of long chain azides with 2- and 3-pyridylanilinothiazoleacetylenes gave triazole-linked pyridylanilinothiazoles which did not retain the VHL-dependent selectivity of parent analogues. For the PPB class, Sonagashira coupling of 4-iodo-(3-pyridylphenylsulfonyl)benzamide with a propargyl hexaethylene glycol carbamate gave an acetylene which was reduced to the corresponding alkyl 3-pyridylphenylsulfonylbenzamide. This reagent retained the VHL-dependent selectivity of the parent analogues and was successfully utilized as an affinity reagent.
Assuntos
Antineoplásicos/farmacologia , Benzamidas/farmacologia , Carcinoma de Células Renais/tratamento farmacológico , Desenho de Fármacos , Neoplasias Renais/tratamento farmacológico , Piridinas/farmacologia , Sulfonas/farmacologia , Tiazóis/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Benzamidas/síntese química , Benzamidas/química , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cromatografia de Afinidade , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Neoplasias Renais/patologia , Modelos Moleculares , Estrutura Molecular , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade , Sulfonas/síntese química , Sulfonas/química , Tiazóis/síntese química , Tiazóis/químicaRESUMO
A novel class of nitroimidazole alkylsulfonamides have been prepared and evaluated as hypoxia-selective cytotoxins and radiosensitisers. The sulfonamide side chain markedly influences the physicochemical properties of the analogues: lowering aqueous solubility and raising the electron affinity of the nitroimidazole group. The addition of hydroxyl or basic amine groups increased aqueous solubility, with charged amine groups contributing to increased electron affinity. The analogues covered the range of electron affinity for effective radiosensitisation with one-electron reduction potentials ranging from -503 to -342mV. Cytotoxicity under normoxia or anoxia against a panel of human tumour cell lines was determined using a proliferation assay. 2-Nitroimidazole sulfonamides displayed significant hypoxia-selective cytotoxicity (6 to 64-fold), while 4- and 5-nitroimidazole analogues did not display hypoxia-selective cytotoxicity. All analogues sensitised anoxic HCT-116 human colorectal cells to radiation at non-toxic concentrations. 2-Nitroimidazole analogues provided modest sensitisation due to the relatively low concentrations used while several 5-nitroimidazole analogues provided equivalent sensitisation to misonidazole and etanidazole at similar molar concentrations.
Assuntos
Hipóxia Celular/efeitos dos fármacos , Nitroimidazóis/farmacologia , Radiossensibilizantes/farmacologia , Sulfonamidas/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Células HCT116 , Células HT29 , Humanos , Estrutura Molecular , Nitroimidazóis/síntese química , Nitroimidazóis/química , Radiossensibilizantes/síntese química , Radiossensibilizantes/química , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/químicaRESUMO
Hypoxia is an important component of the tumor microenvironment and has been the target of drug discovery efforts for almost half a century. These efforts have evolved from offsetting the impact of hypoxia on radiotherapy with oxygen-mimetic radiosensitizers to using hypoxia as a means to selectively target tumors. The more recent description of hypoxia-inducible factors and their role in the hypoxia response network has revealed a host of new drug targets to selectively target tumors. We are developing hypoxia-directed drugs in each of the following areas: novel radiosensitizers for hypofractionated radiotherapy, a second-generation benzotriazine di-N-oxide hypoxia-activated prodrug, and a hypoxia-inducible factor-1-dependent cytotoxin that targets glucose transport. These projects are discussed in the context of hypoxia-directed drug discovery.
Assuntos
Descoberta de Drogas/métodos , Terapia de Alvo Molecular/métodos , Neoplasias/tratamento farmacológico , Microambiente Tumoral/efeitos dos fármacos , Animais , Antineoplásicos/uso terapêutico , Hipóxia Celular/genética , Reparo do DNA/fisiologia , Humanos , Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Radiossensibilizantes/uso terapêutico , Microambiente Tumoral/fisiologiaRESUMO
The emergence of drug-resistant strains of Mycobacterium tuberculosis (M.tb), the causative agent of tuberculosis, is on the rise and increasing antimicrobial resistance is a global threat. This phenomenon necessitates new drug design methods such as a prodrug strategy to develop novel antitubercular agents. The prodrug strategy is a viable and useful means to improve the absorption, distribution, metabolism, excretion and toxicity (ADMET) profiles of pharmacologically active agents. Granulomas are a pathological hallmark of M.tb infection and bear a remarkable resemblance to the tumour microenvironment, including regions of hypoxia. The hypoxic environment observed in the two structures offer an exceptional opportunity to deliver antitubercular agents selectively in a similar manner to hypoxia activated prodrugs in cancer therapy. Nitroimidazoles have been studied extensively as bioactivated prodrugs of cancer, and their suitability as substrates for mammalian reductases highlight their huge potential. This review will discuss the mechanism of action and resistance mechanisms of the current prodrugs used for the treatment of tuberculosis. It will also highlight the potential advantages and challenges of using hypoxia activated prodrugs as a viable strategy to target latent M.tb in hypoxic regions of granulomas.
RESUMO
The dominant role of non-homologous end-joining in the repair of radiation-induced double-strand breaks identifies DNA-dependent protein kinase (DNA-PK) as an excellent target for the development of radiosensitizers. We report the discovery of a new class of imidazo[4,5-c]pyridine-2-one DNA-PK inhibitors. Structure-activity studies culminated in the identification of 78 as a nM DNA-PK inhibitor with excellent selectivity for DNA-PK compared to related phosphoinositide 3-kinase (PI3K) and PI3K-like kinase (PIKK) families and the broader kinome, and displayed DNA-PK-dependent radiosensitization of HAP1 cells. Compound 78 demonstrated robust radiosensitization of a broad range of cancer cells in vitro, displayed high oral bioavailability, and sensitized colorectal carcinoma (HCT116/54C) and head and neck squamous cell carcinoma (UT-SCC-74B) tumor xenografts to radiation. Compound 78 also provided substantial tumor growth inhibition of HCT116/54C tumor xenografts in combination with radiation. Compound 78 represents a new, potent, and selective class of DNA-PK inhibitors with significant potential as radiosensitizers for cancer treatment.
Assuntos
Proteína Quinase Ativada por DNA , Inibidores de Proteínas Quinases , Radiossensibilizantes , Humanos , Proteína Quinase Ativada por DNA/antagonistas & inibidores , Proteína Quinase Ativada por DNA/metabolismo , Animais , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacocinética , Inibidores de Proteínas Quinases/síntese química , Radiossensibilizantes/farmacologia , Radiossensibilizantes/química , Radiossensibilizantes/farmacocinética , Radiossensibilizantes/síntese química , Relação Estrutura-Atividade , Camundongos , Linhagem Celular Tumoral , Imidazóis/farmacologia , Imidazóis/química , Imidazóis/síntese química , Imidazóis/farmacocinética , Piridonas/farmacologia , Piridonas/química , Piridonas/síntese química , Piridonas/farmacocinética , Ensaios Antitumorais Modelo de Xenoenxerto , Camundongos Nus , RatosRESUMO
BACKGROUND: The nitro-chloromethylbenzindoline prodrug nitro-CBI-DEI appears a promising candidate for the anti-cancer strategy gene-directed enzyme prodrug therapy, based on its ability to be converted to a highly cytotoxic cell-permeable derivative by the nitroreductase NfsB from Escherichia coli. However, relative to some other nitroaromatic prodrugs, nitro-CBI-DEI is a poor substrate for E. coli NfsB. To address this limitation we evaluated other nitroreductase candidates from E. coli and Pseudomonas aeruginosa. FINDINGS: Initial screens of candidate genes in the E. coli reporter strain SOS-R2 identified two additional nitroreductases, E. coli NfsA and P. aeruginosa NfsB, as being more effective activators of nitro-CBI-DEI than E. coli NfsB. In monolayer cytotoxicity assays, human colon carcinoma (HCT-116) cells transfected with P. aeruginosa NfsB were >4.5-fold more sensitive to nitro-CBI-DEI than cells expressing either E. coli enzyme, and 23.5-fold more sensitive than untransfected HCT-116. In three dimensional mixed cell cultures, not only were the P. aeruginosa NfsB expressing cells 540-fold more sensitive to nitro-CBI-DEI than pure cultures of untransfected HCT-116, the activated drug that they generated also displayed an unprecedented local bystander effect. CONCLUSION: We posit that the discrepancy in the fold-sensitivity to nitro-CBI-DEI between the two and three dimensional cytotoxicity assays stems from loss of activated drug into the media in the monolayer cultures. This emphasises the importance of evaluating high-bystander GDEPT prodrugs in three dimensional models. The high cytotoxicity and bystander effect exhibited by the NfsB_Pa/nitro-CBI-DEI combination suggest that further preclinical development of this GDEPT pairing is warranted.
Assuntos
Nitrorredutases/metabolismo , Pró-Fármacos/metabolismo , Pseudomonas aeruginosa/enzimologia , Efeito Espectador , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Terapia Enzimática , Expressão Gênica , Terapia Genética , Células HCT116 , Humanos , Concentração Inibidora 50 , Nitrorredutases/genética , Pró-Fármacos/farmacologia , Pró-Fármacos/toxicidade , Pseudomonas aeruginosa/genética , Ensaio Tumoral de Célula-TroncoRESUMO
Assembling a freestanding, sharp-edged slab of homogeneous material that is much denser than gas, but much more rarefied than a solid, is an outstanding technological challenge. The solution may lie in focusing a dense aerosol to assume this geometry. However, whereas the geometrical optics of dilute aerosols is a well-developed field, the dense aerosol limit is mostly unexplored. Yet controlling the geometrical optics of dense aerosols is necessary in preparing such a material slab. Focusing dense aerosols is shown here to be possible, but the finite particle density reduces the effective Stokes number of the flow, a critical result for controlled focusing.