RESUMEN
Fungi of the genus Alternaria are ubiquitous plant pathogens and saprophytes which are able to grow under varying temperature and moisture conditions as well as on a large range of substrates. A spectrum of structurally diverse secondary metabolites with toxic potential has been identified, but occurrence and relative proportion of the different metabolites in complex mixtures depend on strain, substrate, and growth conditions. This review compiles the available knowledge on hazard identification and characterization of Alternaria toxins. Alternariol (AOH), its monomethylether AME and the perylene quinones altertoxin I (ATX-I), ATX-II, ATX-III, alterperylenol (ALP), and stemphyltoxin III (STTX-III) showed in vitro genotoxic and mutagenic properties. Of all identified Alternaria toxins, the epoxide-bearing analogs ATX-II, ATX-III, and STTX-III show the highest cytotoxic, genotoxic, and mutagenic potential in vitro. Under hormone-sensitive conditions, AOH and AME act as moderate xenoestrogens, but in silico modeling predicts further Alternaria toxins as potential estrogenic factors. Recent studies indicate also an immunosuppressive role of AOH and ATX-II; however, no data are available for the majority of Alternaria toxins. Overall, hazard characterization of Alternaria toxins focused, so far, primarily on the commercially available dibenzo-α-pyrones AOH and AME and tenuazonic acid (TeA). Limited data sets are available for altersetin (ALS), altenuene (ALT), and tentoxin (TEN). The occurrence and toxicological relevance of perylene quinone-based Alternaria toxins still remain to be fully elucidated. We identified data gaps on hazard identification and characterization crucial to improve risk assessment of Alternaria mycotoxins for consumers and occupationally exposed workers.
Asunto(s)
Micotoxinas , Perileno , Humanos , Alternaria/metabolismo , Micotoxinas/toxicidad , Micotoxinas/análisis , Mutágenos/toxicidad , Mutágenos/metabolismo , Lactonas/toxicidad , Lactonas/metabolismo , Medición de Riesgo , Contaminación de Alimentos/análisisRESUMEN
Bottlebrush polymers are highly promising as unimolecular nanomedicines due to their unique control over the critical parameters of size, shape and chemical function. However, since they are prepared from biopersistent carbon backbones, most known bottlebrush polymers are non-degradable and thus unsuitable for systemic therapeutic administration. Herein, we report the design and synthesis of novel poly(organo)phosphazene-g-poly(α-glutamate) (PPz-g-PGA) bottlebrush polymers with exceptional control over their structure and molecular dimensions (Dh ≈ 15-50 nm). These single macromolecules show outstanding aqueous solubility, ultra-high multivalency and biodegradability, making them ideal as nanomedicines. While well-established in polymer therapeutics, it has hitherto not been possible to prepare defined single macromolecules of PGA in these nanosized dimensions. A direct correlation was observed between the macromolecular dimensions of the bottlebrush polymers and their intracellular uptake in CT26 colon cancer cells. Furthermore, the bottlebrush macromolecular structure visibly enhanced the pharmacokinetics by reducing renal clearance and extending plasma half-lives. Real-time analysis of the biodistribution dynamics showed architecture-driven organ distribution and enhanced tumor accumulation. This work, therefore, introduces a robust, controlled synthesis route to bottlebrush polypeptides, overcoming limitations of current polymer-based nanomedicines and, in doing so, offers valuable insights into the influence of architecture on the in vivo performance of nanomedicines.
Asunto(s)
Polímeros , Agua , Distribución Tisular , Polímeros/química , Sustancias Macromoleculares , Agua/química , PéptidosRESUMEN
Glutathione (GSH) is the most abundant thiol in mammalian cells and plays a crucial role in maintaining redox cellular homeostasis. The thiols of two GSH molecules can be oxidized to the disulfide GSSG. The cytosolic GSH/GSSG ratio is very high (>100), and its reduction can lead to apoptosis or necrosis, which are of interest in cancer research. CuII ions are very efficient oxidants of thiols, but with an excess of GSH, CuIn(GS)m clusters are formed, in which CuI is very slowly reoxidized by O2 at pH 7.4 and even more slowly at lower pH. Here, the aerobic oxidation of GSH by CuII was investigated at different pH values in the presence of the anticancer thiosemicarbazone Dp44mT, which accumulates in lysosomes and induces lysosomal membrane permeabilization in a Cu-dependent manner. The results showed that CuII-Dp44mT catalyzes GSH oxidation faster than CuII alone at pH 7.4 and hence accelerates the production of very reactive hydroxyl radicals. Moreover, GSH oxidation and hydroxyl radical production by CuII-Dp44mT were accelerated at the acidic pH found in lysosomes. To decipher this unusually faster thiol oxidation at lower pH, density functional theory (DFT) calculations, electrochemical and spectroscopic studies were performed. The results suggest that the acceleration is due to the protonation of CuII-Dp44mT on the hydrazinic nitrogen, which favors the rate-limiting reduction step without subsequent dissociation of the CuI intermediate. Furthermore, preliminary biological studies in cell culture using the proton pump inhibitor bafilomycin A1 indicated that the lysosomal pH plays a role in the activity of CuII-Dp44mT.
Asunto(s)
Cobre , Tiosemicarbazonas , Animales , Catálisis , Cobre/química , Glutatión/química , Disulfuro de Glutatión/química , Disulfuro de Glutatión/metabolismo , Concentración de Iones de Hidrógeno , Mamíferos/metabolismo , Oxidación-Reducción , Compuestos de Sulfhidrilo/química , Tiosemicarbazonas/química , Tiosemicarbazonas/farmacologíaRESUMEN
Malignant pleural mesothelioma (MPM), an aggressive malignancy affecting pleural surfaces, occurs in three main histological subtypes. The epithelioid and sarcomatoid subtypes are characterized by cuboid and fibroblastoid cells, respectively. The biphasic subtype contains a mixture of both. The sarcomatoid subtype expresses markers of epithelial-mesenchymal transition (EMT) and confers the worst prognosis, but the signals and pathways controlling EMT in MPM are not well understood. We demonstrate that treatment with FGF2 or EGF induced a fibroblastoid morphology in several cell lines from biphasic MPM, accompanied by scattering, decreased cell adhesion and increased invasiveness. This depended on the MAP-kinase pathway but was independent of TGFß or PI3-kinase signaling. In addition to changes in known EMT markers, microarray analysis demonstrated differential expression of MMP1, ESM1, ETV4, PDL1 and BDKR2B in response to both growth factors and in epithelioid versus sarcomatoid MPM. Inhibition of MMP1 prevented FGF2-induced scattering and invasiveness. Moreover, in MPM cells with sarcomatoid morphology, inhibition of FGF/MAP-kinase signaling induced a more epithelioid morphology and gene expression pattern. Our findings suggest a critical role of the MAP-kinase axis in the morphological and behavioral plasticity of mesothelioma.
Asunto(s)
Factor de Crecimiento Epidérmico/metabolismo , Transición Epitelial-Mesenquimal/fisiología , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Neoplasias Pulmonares/patología , Mesotelioma/patología , Neoplasias Pleurales/patología , Línea Celular Tumoral , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Metaloproteinasa 1 de la Matriz/metabolismo , Mesotelioma/metabolismo , Mesotelioma Maligno , Neoplasias Pleurales/metabolismo , Transducción de Señal/fisiologíaRESUMEN
The increasing importance of sigma-2 receptor as target for the diagnosis and therapy of tumors paves the way for the development of innovative optically traceable fluorescent probes as tumor cell contrast and therapeutic agents. Here, a novel hybrid organic-inorganic nanostructure is developed by combining the superior fluorescent properties of inorganic quantum dots (QDs), coated with a hydrophilic silica shell (QD@SiO2 NPs), the versatility of the silica shell, and the high selectivity for sigma-2 receptor of the two synthetic ligands, namely, the 6-[(6-aminohexyl)oxy]-2-(3-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)propyl)-3,4-dihydroisoquinolin-1(2H)-one (MLP66) and 6-[1-[3-(4-cyclohexylpiperazin-1-yl)propyl]-1,2,3,4-tetrahydronaphthalen-5-yloxy]hexylamine (TA6). The proposed nanostructures represent a challenging alternative to all previously studied organic small fluorescent molecules, based on the same sigma-2 receptor affinity moieties. Flow cytometry and confocal fluorescence microscopy experiments, respectively, on fixed and living cancerous MCF7 cells, which overexpress the sigma-2 receptor, prove the ability of functionalized (QD@SiO2-TA6 and QD@SiO2-MLP66) NPs to be internalized and demonstrate their affinity to the sigma-2 receptor, ultimately validating the targeting properties conveyed to the NPs by sigma-2 ligand conjugation. The presented QD-based nanoprobes possess a great potential as in vitro selective sigma-2 receptor imaging agent and, consequently, could provide a significant impact to future theranostic applications.
Asunto(s)
Imagen Molecular/métodos , Neoplasias/diagnóstico por imagen , Puntos Cuánticos/química , Receptores sigma/metabolismo , Nanomedicina Teranóstica/métodos , Citometría de Flujo , Colorantes Fluorescentes/química , Colorantes Fluorescentes/metabolismo , Humanos , Ligandos , Células MCF-7 , Microscopía Confocal , Microscopía Fluorescente , Sondas Moleculares/química , Sondas Moleculares/metabolismo , Nanoestructuras/química , Neoplasias/patología , Puntos Cuánticos/metabolismo , Dióxido de Silicio/químicaRESUMEN
Increasing the specificity of cancer therapy, and thereby decreasing damage to normal cells, requires targeting to cancer-cell specific features. The αvß6 integrin is a receptor involved in cell adhesion and is frequently up-regulated in cancer cells compared to normal cells. We have selected a peptide ligand reported to bind specifically to the ß6 integrin and have synthesized a suite of multispecific molecules to explore the potential for targeting of cancer cells. A combination of solid-phase peptide synthesis and chemoselective ligations was used to synthesize multifunctional molecules composed of integrin-targeting peptides, cytotoxic platinum(IV) prodrugs, and fluorescent or affinity probes joined with flexible linkers. The modular synthesis approach facilitates the construction of peptide-drug conjugates with various valencies and properties in a convergent manner. The binding and specificity of the multifunctional peptide conjugates were investigated using a cell line transfected with the ß6 integrin and fluorescence microscopy. This versatile and highly controlled approach to synthesizing labeled peptide-drug conjugates has the potential to target potent cytotoxic drugs specifically to cancer cells, reducing the doses required for effective treatment.
Asunto(s)
Antígenos de Neoplasias/metabolismo , Antineoplásicos/química , Antineoplásicos/farmacología , Integrinas/metabolismo , Compuestos Organoplatinos/química , Compuestos Organoplatinos/farmacología , Péptidos/química , Péptidos/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/farmacocinética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/metabolismo , Sistemas de Liberación de Medicamentos , Humanos , Compuestos Organoplatinos/síntesis química , Compuestos Organoplatinos/farmacocinética , Péptidos/síntesis química , Péptidos/farmacocinética , Técnicas de Síntesis en Fase SólidaRESUMEN
The high mortality rate of lung cancer patients and the frequent occurrence of side effects during cancer therapy demonstrate the need for more selective and targeted drugs. An important and well-established target for lung cancer treatment is the occasionally mutated epidermal growth factor receptor (EGFR). As platinum(II) drugs are still the most important therapeutics against lung cancer, we synthesized in this study the first platinum(IV) complexes coupled to the EGFR-targeting peptide LARLLT (and the shuffled RTALLL as reference). Notably, HPLC-MS measurements revealed two different peaks with the same molecular mass, which turned out to be a transcyclization reaction in the linker between maleimide and the coupled cysteine moiety. With regard to the EGFR specificity, subsequent biological investigations (3-day viability, 14-day clonogenic assays and platinum uptake) on four different cell lines with different verified EGFR expression levels were performed. Unexpectedly, the results showed neither an enhanced activity nor an EGFR expression-dependent uptake of our new compounds. Consequently, fluorophore-coupled peptides were synthesized to re-evaluate the targeting ability of LARLLT itself. However, also with these molecules, flow cytometry measurements showed no correlation of drug uptake with the EGFR expression levels. Taken together, we successfully synthesized the first platinum(IV) complexes coupled to an EGFR-targeting peptide; however, the biological investigations revealed that LARLLT is not an appropriate peptide for enhancing the specific uptake of small-molecule drugs into EGFR-overexpressing cancer cells.
Asunto(s)
Antineoplásicos/farmacología , Receptores ErbB/antagonistas & inhibidores , Compuestos Organoplatinos/farmacología , Péptidos/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Receptores ErbB/metabolismo , Humanos , Estructura Molecular , Compuestos Organoplatinos/síntesis química , Compuestos Organoplatinos/química , Péptidos/química , Relación Estructura-ActividadRESUMEN
A controversial relationship between sigma-2 and progesterone receptor membrane component 1 (PGRMC1) proteins, both representing promising targets for the therapy and diagnosis of tumors, exists since 2011, when the sigma-2 receptor was reported to be identical to PGRMC1. Because a misidentification of these proteins will lead to biased future research hampering the possible diagnostic and therapeutic exploitation of the two targets, there is the need to solve the debate on their identity. With this aim, we have herein investigated uptake and distribution of structurally different fluorescent sigma-2 receptor ligands by flow cytometry and confocal microscopy in MCF7 cells, where together with intrinsic sigma-2 receptors, PGRMC1 was constitutively present or alternatively silenced or overexpressed. HCT116 cells, with constitutive or silenced PGRMC1, were also studied. These experiments showed that the fluorescent sigma-2 ligands bind to their receptor irrespective of PGRMC1 expression. Furthermore, isothermal titration calorimetry was conducted to examine if DTG and PB28, two structurally distinct nanomolar affinity sigma-2 ligands, bind to purified PGRMC1 proteins that have recently been revealed to form both apo-monomeric and heme-mediated dimeric forms. While no binding to apo-PGRMC1 monomer was detected, a micromolar affinity to heme-mediated dimerized PGRMC1 was demonstrated in DTG but not in PB28. The current data provide evidence that sigma-2 receptor and PGRMC1 are not identical, paving the pathway for future unbiased research in which these two attractive targets are treated as different proteins while the identification of the true sigma-2 protein further needs to be pursued.
Asunto(s)
Proteínas de la Membrana/metabolismo , Receptores de Progesterona/metabolismo , Receptores sigma/metabolismo , Línea Celular Tumoral , Colorantes Fluorescentes/metabolismo , Células HCT116 , Humanos , Ligandos , Células MCF-7RESUMEN
α-N-Heterocyclic thiosemicarbazones are among the most promising ribonucleotide reductase inhibitors identified so far. Triapine, the most prominent representative of this class of substances, has been investigated in multiple phase I and II clinical trials. With regard to clinical practice, Triapine showed activity against hematological diseases, but ineffectiveness against a variety of solid tumors. However, the reasons are still vague and the amount of ADME (absorption, distribution, metabolism and excretion) data for Triapine available in the literature is very limited. Therefore, different analytical tools were used to investigate the metabolism of Triapine including electrochemical oxidations, liver microsomes and in vivo samples from mice. The main metabolic reactions, observed by all three methods, were dehydrogenation and hydroxylations, confirming that electrochemistry, as a purely instrumental approach, can be applied for the simulation of metabolic pathways. The dehydrogenated metabolite M1 was identified as a thiadiazole ring-closed oxidation product of Triapine. From a biological point of view, M1, as a key metabolite, is of interest since the crucial chemical property of α-N-heterocyclic thiosemicarbazones to bind metal ions is lost and cytotoxicity studies showed no anticancer activity of M1. The in vivo data of the urine samples revealed very high levels of the metabolites and Triapine itself already 15 min after treatment. This clearly indicates that Triapine is rapidly metabolised and excreted, which represents an important step forward to understand the possible reason for the inefficiency of Triapine against solid tumors.
Asunto(s)
Antineoplásicos/metabolismo , Piridinas/metabolismo , Tiosemicarbazonas/metabolismo , Animales , Línea Celular Tumoral , Humanos , Ratones , Ratones Endogámicos BALB C , Microsomas Hepáticos/metabolismo , Oxidación-ReducciónRESUMEN
Phosphoglycerate dehydrogenase (PHGDH) has emerged as a crucial factor in macromolecule synthesis, neutralizing oxidative stress, and regulating methylation reactions in cancer cells, lymphocytes, and endothelial cells. However, the role of PHGDH in tumor-associated macrophages (TAMs) is poorly understood. Here, we found that the T helper 2 (Th2) cytokine interleukin-4 and tumor-conditioned media upregulate the expression of PHGDH in macrophages and promote immunosuppressive M2 macrophage activation and proliferation. Loss of PHGDH disrupts cellular metabolism and mitochondrial respiration, which are essential for immunosuppressive macrophages. Mechanistically, PHGDH-mediated serine biosynthesis promotes α-ketoglutarate production, which activates mTORC1 signaling and contributes to the maintenance of an M2-like macrophage phenotype in the tumor microenvironment. Genetic ablation of PHGDH in macrophages from tumor-bearing mice results in attenuated tumor growth, reduced TAM infiltration, a phenotypic shift of M2-like TAMs toward an M1-like phenotype, downregulated PD-L1 expression and enhanced antitumor T-cell immunity. Our study provides a strong basis for further exploration of PHGDH as a potential target to counteract TAM-mediated immunosuppression and hinder tumor progression.
Asunto(s)
Ácidos Cetoglutáricos , Diana Mecanicista del Complejo 1 de la Rapamicina , Fosfoglicerato-Deshidrogenasa , Transducción de Señal , Microambiente Tumoral , Macrófagos Asociados a Tumores , Animales , Macrófagos Asociados a Tumores/inmunología , Macrófagos Asociados a Tumores/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Fosfoglicerato-Deshidrogenasa/metabolismo , Ratones , Ácidos Cetoglutáricos/metabolismo , Humanos , Ratones Endogámicos C57BL , Fenotipo , Línea Celular Tumoral , Activación de MacrófagosRESUMEN
Thiosemicarbazones (TSCs) are a class of biologically active compounds with promising anticancer activity. Their typical mechanism, especially of the clinically far developed representative Triapine, is chelation of iron (Fe), with the Fe-containing enzyme ribonucleotide reductase as primary intracellular target. However, for the subclass of terminally disubstituted, nanomolar-active derivatives like Dp44mT and Me2NNMe2, recent findings suggest that the chelation, stability, and reduction properties of the copper(II) (Cu) complexes are essential for their modes of action. Consequently, it is important to elucidate whether blood serum Cu(II) is a potential metal source for these TSCs. To gain more insights, the interaction of Triapine, Dp44mT or Me2NNMe2 with purified human serum albumin (HSA) as the main pool of labile Cu(II) was investigated by UV-vis and electron paramagnetic resonance measurements. Subsequently, a size-exclusion chromatography inductively coupled plasma mass spectrometry method for the differentiation of Cu species in serum was developed, especially separating the non-labile Cu enzyme ceruloplasmin from HSA. The results indicate that the TSCs specifically chelate copper from the N-terminal Cu-binding site of HSA. Furthermore, the Cu(II)-TSC complexes were shown to form ternary HSA conjugates, most likely via histidine. Noteworthy, Fe-chelation from transferrin was not overserved, even not for Triapine. In summary, the labile Cu pool of HSA is a potential source for Cu-TSC complex formation and, consequently, distinctly influences the anticancer activity and pharmacological behavior of TSCs.
Asunto(s)
Antineoplásicos , Tiosemicarbazonas , Humanos , Albúmina Sérica Humana , Cobre/química , Tiosemicarbazonas/farmacología , Tiosemicarbazonas/química , Quelantes/química , Antineoplásicos/farmacología , Antineoplásicos/químicaRESUMEN
COTI-2 is currently being evaluated in a phase I clinical trial for the treatment of gynecological and other solid cancers. As a thiosemicarbazone, this compound contains an N,N,S-chelating moiety and is, therefore, expected to bind endogenous metal ions. However, besides zinc, the metal interaction properties of COTI-2 have not been investigated in detail so far. This is unexpected, as we have recently shown that COTI-2 forms stable ternary complexes with copper and glutathione, which renders this drug a substrate for the resistance efflux transporter ABCC1. Herein, the complex formation of COTI-2, two novel terminal N-disubstituted derivatives (COTI-NMe2 and COTI-NMeCy), and the non-substituted analogue (COTI-NH2) with iron, copper, and zinc ions was characterized in detail. Furthermore, their activities against drug-resistant cancer cells was investigated in comparison to COTI-2 and Triapine. These data revealed that, besides zinc, also iron and copper ions need to be considered to play a role in the mode of action and resistance development of these thiosemicarbazones. Moreover, we identified COTI-NMe2 as an interesting new drug candidate with improved anticancer activity and resistance profile.
RESUMEN
α-N-Heterocyclic thiosemicarbazones such as triapine and COTI-2 are currently investigated as anticancer therapeutics in clinical trials. However, triapine was widely inactive against solid tumor types. A likely explanation is the short plasma half-life time and fast metabolism. One promising approach to overcome these drawbacks is the encapsulation of the drug into nanoparticles (passive drug-targeting). In a previous work we showed that it was not possible to stably encapsulate free triapine into liposomes. Hence, in this manuscript we present the successful preparation of liposomal formulations of the copper(II) complexes of triapine and COTI-2. To this end, various drug-loading strategies were examined and the resulting liposomes were physico-chemically characterized. Especially for liposomal Cu-triapine, a decent encapsulation efficacy and a slow drug release behavior could be observed. In contrast, for COTI-2 and its copper(II) complex no stable loading could be achieved. Subsequent in vitro studies in different cell lines with liposomal Cu-triapine showed the expected strongly reduced cytotoxicity and DNA damage induction. Also in vivo distinctly higher copper plasma levels and a continuous release could be observed for the liposomal formulation compared to free Cu-triapine. Taken together, the here presented nanoformulation of Cu-triapine is an important step further to increase the plasma half-life time and tumor targeting properties of anticancer thiosemicarbazones.
Asunto(s)
Antineoplásicos , Complejos de Coordinación , Cobre , Tiosemicarbazonas , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/química , Antineoplásicos/farmacocinética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Complejos de Coordinación/administración & dosificación , Complejos de Coordinación/química , Complejos de Coordinación/farmacocinética , Cobre/administración & dosificación , Cobre/química , Cobre/farmacocinética , Liberación de Fármacos , Femenino , Humanos , Liposomas , Metahemoglobina/metabolismo , Ratones Endogámicos BALB C , Tiosemicarbazonas/administración & dosificación , Tiosemicarbazonas/química , Tiosemicarbazonas/farmacocinéticaRESUMEN
Chemotherapy with platinum complexes is essential for clinical anticancer therapy. However, due to side effects and drug resistance, further drug improvement is urgently needed. Herein, we report on triple-action platinum(IV) prodrugs, which, in addition to tumor targeting via maleimide-mediated albumin binding, release the immunomodulatory ligand 1-methyl-d-tryptophan (1-MDT). Unexpectedly, structure-activity relationship analysis showed that the mode of 1-MDT conjugation distinctly impacts the reducibility and thus activation of the prodrugs. This in turn affected ligand release, pharmacokinetic properties, efficiency of immunomodulation, and the anticancer activity in vitro and in a mouse model in vivo. Moreover, we could demonstrate that the design of albumin-targeted multi-modal prodrugs using platinum(IV) is a promising strategy to enhance the cellular uptake of bioactive ligands with low cell permeability (1-MDT) and to improve their selective delivery into the malignant tissue. This will allow tumor-specific anticancer therapy supported by a favorably tuned immune microenvironment.
Asunto(s)
Antineoplásicos/uso terapéutico , Complejos de Coordinación/uso terapéutico , Factores Inmunológicos/uso terapéutico , Maleimidas/uso terapéutico , Neoplasias/tratamiento farmacológico , Profármacos/uso terapéutico , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Línea Celular Tumoral , Complejos de Coordinación/síntesis química , Complejos de Coordinación/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Factores Inmunológicos/síntesis química , Factores Inmunológicos/farmacología , Indolamina-Pirrol 2,3,-Dioxigenasa/antagonistas & inhibidores , Masculino , Maleimidas/síntesis química , Maleimidas/farmacología , Ratones Endogámicos BALB C , Ratones SCID , Estructura Molecular , Platino (Metal)/química , Profármacos/síntesis química , Profármacos/farmacología , Relación Estructura-Actividad , Succinimidas/síntesis química , Succinimidas/farmacología , Succinimidas/uso terapéuticoRESUMEN
COTI-2 is a novel anticancer thiosemicarbazone in phase I clinical trial. However, the effects of metal complexation (a main characteristic of thiosemicarbazones) and acquired resistance mechanisms are widely unknown. Therefore, in this study, the copper and iron complexes of COTI-2 were synthesized and evaluated for their anticancer activity and impact on drug resistance in comparison to metal-free thiosemicarbazones. Investigations using Triapine-resistant SW480/Tria and newly established COTI-2-resistant SW480/Coti cells revealed distinct structure-activity relationships. SW480/Coti cells were found to overexpress ABCC1, and COTI-2 being a substrate for this efflux pump. This was unexpected, as ABCC1 has strong selectivity for glutathione adducts. The recognition by ABCC1 could be explained by the reduction kinetics of a ternary Cu-COTI-2 complex with glutathione. Thus, only thiosemicarbazones forming stable, nonreducible copper(II)-glutathione adducts are recognized and, in turn, effluxed by ABCC1. This reveals a crucial connection between copper complex chemistry, glutathione interaction, and the resistance profile of clinically relevant thiosemicarbazones.
Asunto(s)
Aminoquinolinas/metabolismo , Cobre/metabolismo , Resistencia a Antineoplásicos/fisiología , Glutatión/metabolismo , Líquido Intracelular/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Tiosemicarbazonas/metabolismo , Aminoquinolinas/química , Aminoquinolinas/farmacología , Antineoplásicos/química , Antineoplásicos/metabolismo , Antineoplásicos/farmacología , Línea Celular Tumoral , Cobre/química , Resistencia a Antineoplásicos/efectos de los fármacos , Glutatión/química , Humanos , Líquido Intracelular/efectos de los fármacos , Tiosemicarbazonas/química , Tiosemicarbazonas/farmacología , Difracción de Rayos XRESUMEN
Aims: Due to their significant biological activity, thiosemicarbazones (TSCs) are promising candidates for anticancer therapy. In part, the efficacy of TSCs is linked to their ability to chelate essential metal ions such as copper and iron. Triapine, the best-studied anticancer TSC, has been tested clinically with promising results in hematological diseases. During the past few years, a novel subclass of TSCs with improved anticancer activity was found to induce paraptosis, a recently characterized form of cell death. The aim of this study was to identify structural and chemical properties associated with anticancer activity and paraptosis induction of TSCs. Results: When testing a panel of structurally related TSCs, compounds with nanomolar anticancer activity and paraptosis-inducing properties showed higher copper(II) complex solution stability and a slower reduction rate, which resulted in reduced redox activity. In contrast, TSCs with lower anticancer activity induced higher levels of superoxide that rapidly stimulated superoxide dismutase expression in treated cells, effectively protecting the cells from drug-induced redox stress. Innovation: Consequently, we hypothesize that in the case of close Triapine derivatives, intracellular reduction leads to rapid dissociation of intracellularly formed copper complexes. In contrast, TSCs characterized by highly stable, slowly reducible copper(II) complexes are able to reach new intracellular targets such as the endoplasmic reticulum-resident protein disulfide isomerase. Conclusion: The additional modes of actions observed with highly active TSC derivatives are based on intracellular formation of stable copper complexes, offering a new approach to combat (drug-resistant) cancer cells.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Cobre/química , Resistencia a Antineoplásicos/efectos de los fármacos , Tiosemicarbazonas/química , Tiosemicarbazonas/farmacología , Antioxidantes/química , Antioxidantes/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Retículo Endoplásmico/metabolismo , Humanos , Oxidación-Reducción/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , SolubilidadRESUMEN
We have recently demonstrated that riboflavin (Rf) functions as unconventional bioorthogonal photocatalyst for the activation of PtIV prodrugs. In this study, we show how the combination of light and Rf with two PtIV prodrugs is a feasible strategy for light-mediated pancreatic cancer cell death induction. In Capan-1 cells, which have high tolerance against photodynamic therapy, Rf-mediated activation of the cisplatin and carboplatin prodrugs cis,cis,trans-[Pt(NH3)2(Cl)2(O2CCH2CH2CO2H)2] (1) and cis,cis,trans-[Pt(NH3)2(CBDCA)(O2CCH2CH2CO2H)2] (2, where CBDCA = cyclobutane dicarboxylate) resulted in pronounced reduction of the cell viability, including under hypoxia conditions. Such photoactivation mode occurs to a considerable extent intracellularly, as demonstrated for 1 by uptake and cell viability experiments. 195Pt NMR, DNA binding studies using circular dichroism, mass spectrometry and immunofluorescence microscopy were performed using the Rf-1 catalyst-substrate pair and indicated that cell death is associated with the efficient light-induced formation of cisplatin. Accordingly, Western blot analysis revealed signs of DNA damage and activation of cell death pathways through Rf-mediated photochemical activation. Phosphorylation of H2AX as indicator for DNA damage, was detected for Rf-1 in a strictly light-dependent fashion while in case of free cisplatin also in the dark. Photochemical induction of nuclear pH2AX foci by Rf-1 was confirmed in fluorescence microscopy again proving efficient light-induced cisplatin release from the prodrug system.
Asunto(s)
Antineoplásicos/metabolismo , Luz , Compuestos Organoplatinos/metabolismo , Procesos Fotoquímicos , Profármacos/metabolismo , Riboflavina/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , HumanosRESUMEN
Due to their high biological activity, thiosemicarbazones have been developed for treatment of diverse diseases, including cancer, resulting in multiple clinical trials especially of the lead compound Triapine. During the last years, a novel subclass of anticancer thiosemicarbazones has attracted substantial interest based on their enhanced cytotoxic activity. Increasing evidence suggests that the double-dimethylated Triapine derivative Me2NNMe2 differs from Triapine not only in its efficacy but also in its mode of action. Here we show that Me2NNMe2- (but not Triapine)-treated cancer cells exhibit all hallmarks of paraptotic cell death including, besides the appearance of endoplasmic reticulum (ER)-derived vesicles, also mitochondrial swelling and caspase-independent cell death via the MAPK signaling pathway. Subsequently, we uncover that the copper complex of Me2NNMe2 (a supposed intracellular metabolite) inhibits the ER-resident protein disulfide isomerase, resulting in a specific form of ER stress based on disruption of the Ca2+ and ER thiol redox homeostasis. Our findings indicate that compounds like Me2NNMe2 are of interest especially for the treatment of apoptosis-resistant cancer and provide new insights into mechanisms underlying drug-induced paraptosis.
Asunto(s)
Antineoplásicos/farmacología , Retículo Endoplásmico/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Proteína Disulfuro Isomerasas/antagonistas & inhibidores , Tiosemicarbazonas/farmacología , Antineoplásicos/síntesis química , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Complejos de Coordinación/química , Complejos de Coordinación/farmacología , Cobre/química , Cobre/metabolismo , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/patología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Expresión Génica , Células HCT116 , Humanos , Sistema de Señalización de MAP Quinasas/genética , Mitocondrias/metabolismo , Mitocondrias/patología , Dilatación Mitocondrial/efectos de los fármacos , Oxidación-Reducción/efectos de los fármacos , Proteína Disulfuro Isomerasas/genética , Proteína Disulfuro Isomerasas/metabolismo , Piridinas/farmacología , Compuestos de Sulfhidrilo/antagonistas & inhibidores , Compuestos de Sulfhidrilo/metabolismo , Tiosemicarbazonas/síntesis químicaRESUMEN
One of the most promising classes of iron chelators are α-N-heterocyclic thiosemicarbazones with Triapine as the most prominent representative. In several clinical trials Triapine showed anticancer activity against hematological diseases, however, studies on solid tumors failed due to widely unknown reasons. Some years ago, it was recognized that "terminal dimethylation" of thiosemicarbazones can lead to a more than 100-fold increased activity, probably due to interactions with cellular copper depots. To better understand the structural requirements for the switch to nanomolar cytotoxicity, we systematically synthesized all eight possible N-methylated derivatives of Triapine and investigated their potential against Triapine-sensitive as well as -resistant cell lines. While only the "completely" methylated compound exerted nanomolar activity, the data revealed that all compounds with at least one N-dimethylation were not affected by acquired Triapine resistance. In addition, these compounds were highly synergistic with copper treatment accompanied by induction of reactive oxygen species and massive necrotic cell death.