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Biocompatible thermoresponsive copolymers based on 2-(2-methoxyethoxy) ethyl methacrylate (MEO2MA) and oligo (ethylene glycol) methacrylate (OEGMA) were grown from the surface of ZnO quantum dots (QDs) by surface initiated atom transfer radical polymerization with activators regenerated by electron transfer (SI-ARGET ATRP) in order to design smart and fluorescent core/shell nanosystems to be used toward cancer cells. Tunable lower critical solution temperature (LCST) values were obtained and studied in water and in culture medium. The complete efficiency of the process was demonstrated by the combination of spectroscopic and microscopic studies. The colloidal behavior of the ZnO/copolymer core/shell QDs in water and in physiological media with temperature was assessed. Finally, the cytotoxicity toward human colon cancer HT29 cells of the core/shell QDs was tested. The results showed that the polymer-capped QDs exhibited almost no toxicity at concentrations up to 12.5 µg.mL-1, while when loaded with doxorubicin hydrochloride (DOX), a higher cytotoxicity and a decreased HT29 cancer cell viability in a short time were observed.
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Doxorrubicina/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Neoplasias/tratamento farmacológico , Pontos Quânticos/uso terapêutico , Sobrevivência Celular/efeitos dos fármacos , Coloides , Doxorrubicina/farmacologia , Células HT29 , Humanos , Metacrilatos/química , Polimerização , Pontos Quânticos/química , Pontos Quânticos/toxicidade , Óxido de ZincoRESUMO
The aim of this article is to give an insight into the future of photodynamic therapy (PDT) in head and neck squamous cell carcinoma (HNSCC). Through the combination of a photosensitizing agent with light and oxygen, PDT produces highly cytotoxic reactive oxygen species leading to selective tumor eradication. PDT is an attractive treatment for focal therapy of localized tumors, especially in the case of unresectable tumors. In HNSCC, over 1500 patients have been treated by PDT, and the majority of them responded quite favorably to this treatment. However, the non-negligible photosensitization of healthy tissue is a major limitation for the clinical application of PDT. Improvement in tumor selectivity is the main challenge that can be taken up by the use of a new generation of photosensitizing nanoparticles. Passive targeting, by using functionalised nanocarriers to target to overexpressed transmembrane receptors afford attractive solutions. To this day, epidermal growth factor receptor (EGFR) remains the only validated molecular target for HNSCC, and photosensitizer immunoconjugates to EGFR have been developed for the intracellular delivery of photosensitizing agents. Depending on coordinated research between biomarkers, specific ligands, and photosensitizers, similar approaches could be rapidly developed. In addition, some photosensitizers hold high fluorescence yield and therefore could emerge as theranostic agents.
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Carcinoma de Células Escamosas/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Terapia de Alvo Molecular/métodos , Fotoquimioterapia/métodos , Animais , Humanos , Pesquisa Translacional BiomédicaRESUMO
INTRODUCTION: Plasma biomarkers may be useful in diagnosing acute cerebral infarction requiring urgent reperfusion, but their performance remains to be confirmed. If confirmed, these molecules could be used to develop rapid and reliable decentralised measurement methods, making it possible to initiate reperfusion therapy before hospital admission. The FLAG-1 large prospective study will constitute a plasma bank to assess the diagnostic performance of two biomarkers: glutathione S-transferase-π and peroxiredoxin 1. These molecules are involved in the oxidative stress response and could identify cerebral infarction within a therapeutic window of less than 4.5 hours following the onset of symptoms. Secondary objectives include assessing performance of these biomarkers within 3-hour and 6-hour windows; identifying additional biomarkers diagnosing cerebral infarction and significant criteria guiding therapeutic decisions: ischaemic features of stroke, presence of diffusion/fluid-attenuated inversion recovery mismatch, volume of cerebral infarction and penumbra on cerebral MRI. METHODS AND ANALYSIS: The exploratory, prospective, multicentre FLAG-1 Study will include 945 patients with acute stroke symptoms (onset ≤12 hours, National Institute of Health Stroke Scale score ≥3). Each patient's 25 mL blood sample will be associated with cerebral MRI data. Two patient groups will be defined based on the time of blood collection (before and after 4.5 hours following onset). Receiver operating characteristic analysis will determine the diagnostic performance of each biomarker, alone or in combination, for the identification of cerebral infarction <4.5 hours. ETHICS AND DISSEMINATION: The protocol has been approved by an independent ethics committee. Biological samples are retained in line with best practices and procedures, in accordance with French legislation. Anonymised data and cerebral imaging records are stored using electronic case report forms and a secure server, respectively, registered with the French Data Protection Authority (Commission Nationale de l'Informatique et des Libertés (CNIL)). Results will be disseminated through scientific meetings and publication in peer-reviewed medical journals. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov Registry (NCT03364296).
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Glutationa S-Transferase pi , Peroxirredoxinas , Infarto Cerebral/diagnóstico por imagem , Humanos , Estudos Multicêntricos como Assunto , Estudos Prospectivos , ReperfusãoRESUMO
Immunohistochemistry to active caspase-3, recently recommended for apoptosis detection, is inappropriate to detect apoptosis involving caspase-7. Cleavage of poly-ADP-ribose polymerase 1 (PARP-1), a major substrate of both caspases, is a valuable marker of apoptosis. Apoptosis evaluation induced in vitro either by paclitaxel or by photodynamic treatment (PDT) with Foscan in HT29 or KB monolayer cells and HT29 spheroids yielded a close percentage of labeled cells whatever the antibody used, whereas in control specimens, cleaved PARP (c-PARP) immunostaining failed to detect apoptosis as efficiently as active caspase-3 or -7 immunostaining. Studies in MDA-MB231 monolayer cells and HT29 xenografts either subjected or not subjected to Foscan-PDT resulted in a significant higher number of active caspase-3-labeled cells, although immunofluorescence analysis showed c-PARP and active caspase-3 perfectly colocalized in tumors. A restricted expression of c-PARP was obvious in the greater part of caspase-3 expressing cells from control tumor, whereas photosensitized tumors showed a higher number of cells expressing large fluorescent spots from both active caspase-3 and c-PARP. These results support the assumption that c-PARP expression was dependent on treatment-induced apoptosis. The absence of caspase-7 activation in some caspase-3-expressing cells undergoing Foscan-PDT shows the relevance of using antibodies that can discriminate caspase-dependent apoptotic pathways.
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Adenocarcinoma/patologia , Apoptose , Caspase 3/metabolismo , Caspase 7/metabolismo , Neoplasias do Colo/patologia , Poli(ADP-Ribose) Polimerases/metabolismo , Esferoides Celulares/metabolismo , Adenocarcinoma/metabolismo , Animais , Antineoplásicos/farmacologia , Neoplasias do Colo/metabolismo , Humanos , Imuno-Histoquímica , Mesoporfirinas/farmacologia , Camundongos , Camundongos Nus , Transplante de Neoplasias , Paclitaxel/farmacologia , Poli(ADP-Ribose) Polimerase-1 , Transplante Heterólogo , Triazenos/farmacologiaRESUMO
OBJECTIVE: To identify regulators of the cartilaginous phenotype, on the basis of their differential expression in human conventional chondrogenic tumors compared with articular cartilage. METHODS: Differential proteomics analysis revealed matrilin-3 (MATN3) as a candidate regulator of the cartilaginous phenotype. Its capacity to modulate gene expression was investigated in human HCS-2/8 chondrosarcoma cells and transfected chondrocytes, using cell culture fractionation, reverse transcription-polymerase chain reaction, and Western blot analyses. RESULTS: Increased expression of the cartilage-specific matrix protein MATN3 was specifically observed in enchondromas and conventional chondrosarcomas. A substantial fraction of MATN3 was found in cytoplasmic structures of tumor cells, as demonstrated by immunohistochemistry. Analyses of intracellular MATN3 revealed that it corresponded to an imperfectly maturated MATN3 polypeptide, both in HCS-2/8 human chondrosarcoma cells and in transfected human chondrocytes. Moderately increased expression of MATN3 resulted in its intracellular retention. Antibody-mediated blockade of soluble, extracellular MATN3 in HCS-2/8 cell cultures resulted in increased expression of MATN3 and the chondrogenic transcription factor SOX9. Conversely, increased ectopic expression of MATN3 resulted in decreased expression of MATN3 and SOX9 in primary chondrocytes, while a mutant MATN3 lacking its first epidermal growth factor (EGF)-like domain failed to down-regulate SOX9. CONCLUSION: Aberrant expression and processing of MATN3 are hallmarks of conventional cartilaginous neoplasms. A particular step in the maturation of MATN3 limits its processing through the secretion machinery, resulting in its intracellular accumulation upon increased expression. Soluble, secreted MATN3, however, down-regulates SOX9 at the messenger RNA and protein levels. The first EGF-like domain of MATN3 is a critical determinant of its regulatory activity toward SOX9. These activities of MATN3 suggest that its increased expression in osteoarthritis might contribute to the degeneration of articular cartilage.
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Neoplasias Ósseas/metabolismo , Cartilagem Articular/metabolismo , Condroma/metabolismo , Condrossarcoma/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Osteoartrite/metabolismo , Fatores de Transcrição SOX9/metabolismo , Adolescente , Adulto , Idoso , Western Blotting , Neoplasias Ósseas/genética , Diferenciação Celular/genética , Células Cultivadas , Criança , Pré-Escolar , Condrócitos/citologia , Condrócitos/metabolismo , Condroma/genética , Condrossarcoma/genética , Regulação para Baixo , Fator de Crescimento Epidérmico/genética , Proteínas da Matriz Extracelular/genética , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Masculino , Proteínas Matrilinas , Pessoa de Meia-Idade , Osteoartrite/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição SOX9/genética , Transdução de Sinais/genéticaRESUMO
UNLABELLED: Apoptosis detection in histological section is very important, but usual methods (TUNEL and morphologic changes) are controversial. Immunohistochemical stains of activated proteins during apoptosis improve detection of cell death in tissue sections. Activated caspase-3 and cleaved cytokeratin-18 are more and more used. OBJECTIVE: This study compared immunohistochemical markers (activated caspase-3, cleaved cytokeratin-18 and two antibodies not yet evaluated: activated caspase-7 and cleaved PARP-1), cellular morphology and TUNEL. MATERIAL AND METHODS: Tumour xenografts of the human colon cancer cell line HT29 were used, treated by photodynamic therapy, to obtain large numbers of cells undergoing apoptosis. Apoptotic cells were quantified and apoptotic indices were determined for each marker. RESULTS: Comparison of apoptosis index indicated statistically best sensibility with activated-caspase-3 and cleaved-cytokeratin-19. CONCLUSION: Immunohistochemistry for activated caspase-3 and cleaved cytkeratin-18 appear to be an easy, sensitive, and reliable method for detecting and quantifying apoptosis in this model. There are therefore recommended for the detection and quantification of apoptosis in tissue sections. Other markers as cleaved PARP-1 and activated caspase-7 can be an interessant solution: advantages and inconvenience for each methods are exposed.
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Apoptose , Adenocarcinoma/patologia , Neoplasias do Colo/patologia , Células HT29 , Humanos , Patologia/métodosRESUMO
Complete surgical resection is the ideal cure for ovarian peritoneal carcinomatosis, but remains challenging. Fluorescent guided surgery can be a promising approach for precise cytoreduction when appropriate fluorophore is used. In the presence paper, we review already developed near- and short-wave infrared fluorescent nanoparticles, which are currently under investigation for peritoneal carcinomatosis fluorescence imaging. We also highlight the main ways to improve the safety of nanoparticles, for fulfilling prerequisites of clinical application.
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The increasing number of publications on the subject shows that nanomedicine is an attractive field for investigations aiming to considerably improve anticancer chemotherapy. Based on selective tumor targeting while sparing healthy tissue, carrier-mediated drug delivery has been expected to provide significant benefits to patients. However, despite reduced systemic toxicity, most nanodrugs approved for clinical use have been less effective than previously anticipated. The gap between experimental results and clinical outcomes demonstrates the necessity to perform comprehensive drug screening by using powerful preclinical models. In this context, in vitro three-dimensional models can provide key information on drug behavior inside the tumor tissue. The multicellular tumor spheroid (MCTS) model closely mimics a small avascular tumor with the presence of proliferative cells surrounding quiescent cells and a necrotic core. Oxygen, pH and nutrient gradients are similar to those of solid tumor. Furthermore, extracellular matrix (ECM) components and stromal cells can be embedded in the most sophisticated spheroid design. All these elements together with the physicochemical properties of nanoparticles (NPs) play a key role in drug transport, and therefore, the MCTS model is appropriate to assess the ability of NP to penetrate the tumor tissue. This review presents recent developments in MCTS models for a better comprehension of the interactions between NPs and tumor components that affect tumor drug delivery. MCTS is particularly suitable for the high-throughput screening of new nanodrugs.
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Sistemas de Liberação de Medicamentos/métodos , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Nanomedicina/métodos , Neoplasias/tratamento farmacológico , Esferoides Celulares , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacocinética , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/uso terapêutico , Matriz Extracelular/efeitos dos fármacos , Humanos , Nanopartículas/administração & dosagem , Esferoides Celulares/química , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/patologiaRESUMO
BACKGROUND: Telestroke is recognized as a safe and time-efficient way of treating stroke patients. However, admission centers (spokes) are subject to financial charges which can make them reluctant to join the system. We implemented and assessed an economic model supporting our telestroke system, Virtuall, France, which includes one expert center (hub) and six spokes. METHODS: The model is based on payment for the expertise provided by the hub, distribution of charges related to telemedicine according to the fees perceived by the spokes, and transfer of patients between the spokes and the hub. We performed a cost-benefit analysis for all patients included in Virtuall from January 2014 to December 2015 to assess the economic balance in each center. RESULTS: 321 patients were prospectively included in the study. Application of the economic model resulted in overall financial balance with funding of a dedicated medical service in the hub, and reduced costs directly related to telestroke by an average of 10% in the spokes. The conditions generating the highest costs for the spokes were: a patient returning from the hub for re-hospitalization (mean cost of $1,995/patient); management of patients treated by intravenous thrombolysis without transfer to the hub (mean cost of $2,075/patient). The most favorable financial condition for the spokes remained simple transfer of patients to the hub and no return (mean cost of $329/patient). CONCLUSION: We describe an economic model which can be applied to any telestroke system to ensure the optimal balance between hub and spoke centers.
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The unique physical properties of the superparamagnetic nanoparticles (SPIONs) have made them candidates of choice in nanomedicine especially for diagnostic imaging, therapeutic applications and drug delivery based systems. In this study, superparamagnetic Fe3O4 NPs were synthesized and functionalized with a biocompatible thermoresponsive copolymer to obtain temperature responsive core/shell NPs. The ultimate goal of this work is to build a drug delivery system able to release anticancer drugs in the physiological temperatures range. The core/shell NPs were first synthesized and their chemical, physical, magnetic and thermo-responsive properties where fully characterized in a second step. The lower critical solution temperature (LCST) of the core/shell NPs was tuned in physiological media in order to release the cancer drug at a controlled temperature slightly above the body temperature to avoid any premature release of the drug. The core/shell NPs exhibiting the targeted LCST were then loaded with Doxurubicin (DOX) and the drug release properties were then studied with the temperature. Moreover the cytotoxicity tests have shown that the core/shell NPs had a very limited cytotoxicity up to concentration of 25µg/mL. This investigation showed that the significant release occurred at the targeted temperature in the physiological media making those nano-systems very promising for further use in drug delivery platform.
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Antineoplásicos/administração & dosagem , Doxorrubicina/administração & dosagem , Sistemas de Liberação de Medicamentos , Nanopartículas de Magnetita/administração & dosagem , Polietilenoglicóis/administração & dosagem , Ácidos Polimetacrílicos/administração & dosagem , Antineoplásicos/química , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/química , Liberação Controlada de Fármacos , Células HT29 , Humanos , Nanopartículas de Magnetita/química , Polietilenoglicóis/química , Ácidos Polimetacrílicos/química , TemperaturaRESUMO
Human scent identification is based on a matching-to-sample task in which trained dogs are required to compare a scent sample collected from an object found at a crime scene to that of a suspect. Based on dogs' greater olfactory ability to detect and process odours, this method has been used in forensic investigations to identify the odour of a suspect at a crime scene. The excellent reliability and reproducibility of the method largely depend on rigor in dog training. The present study describes the various steps of training that lead to high sensitivity scores, with dogs matching samples with 90% efficiency when the complexity of the scents presented during the task in the sample is similar to that presented in the in lineups, and specificity reaching a ceiling, with no false alarms in human scent matching-to-sample tasks. This high level of accuracy ensures reliable results in judicial human scent identification tests. Also, our data should convince law enforcement authorities to use these results as official forensic evidence when dogs are trained appropriately.
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Cães , Ciências Forenses/educação , Odorantes , Animais , Crime , Feminino , Humanos , Masculino , Fatores de TempoRESUMO
Photodynamic therapy (PDT) is an approved anticancer treatment modality that eliminates unwanted cells by the photochemical generation of reactive oxygen species following absorption of visible light by a photosensitizer, which is selectively taken up by tumor cells. Present study reports the modalities of cell death after photosensitization of human adenocarcinoma HT29 monolayer and spheroid cells with a second generation photosensitizer Foscan. Kinetics of apoptosis and necrosis after Foscan-PDT in monolayer cells determined by flow cytometry using labeling of cleaved poly(ADP-ribose) polymerase (PARP) and staining with propidium iodide (PI) demonstrated that Foscan was not a strong inducer of apoptosis and necrosis was a prevailing mode of cell death. Cytochrome c release (cyt c) and mitochondrial membrane potential (Deltapsim) addressed by flow cytometry technique at different time points post-Foscan-PDT demonstrated that cell photoinactivation was governed by these mitochondrial events. Foscan-loaded HT29 multicell spheroids, subjected to irradiation with different fluence rates and equivalent light doses, displayed much better tumoricidal activity at the lowest fluence rate used. Apoptosis, measured by caspase-3 activation was evidenced only in spheroids irradiated with the lowest fluence rate and moderate fluence inducing 65% of cell death. Application of higher fluence rates for the same level of photocytotoxicity did not result in caspase-3 activation. The observation of the fluence rate-dependent modulation of caspase-3 activity in spheroids offers the possibility of regulating the mechanism of direct cell photodamage and could be of great potential in the clinical context.
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Apoptose/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Mesoporfirinas/farmacologia , Fármacos Fotossensibilizantes/farmacologia , Esferoides Celulares/efeitos dos fármacos , Western Blotting , Caspase 3 , Caspases/análise , Caspases/metabolismo , Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Ativação Enzimática , Citometria de Fluxo , Células HT29 , Humanos , Cinética , Potenciais da Membrana/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Necrose/patologia , FotoquimioterapiaRESUMO
Non-steroidal anti-inflammatory drugs (NSAIDs) have shown chemopreventive properties in colorectal cancer, involving both cyclooxygenase (COX)-dependent and -independent mechanisms. Apart from their selectivity for COX isoenzymes, NSAIDs differ in their acidic character which supports ability to uncouple oxidative phosphorylation. To assess the possible contribution of uncoupling to their antineoplastic properties, we compared the effect of sulindac sulfide (SS), an acidic NSAID and NS-398, a non-acidic tricyclic, on mitochondrial function and apoptosis in colorectal cancer cell lines (HT29, Caco-2, HCT15 and HCT116). Although cell lines displayed a different COX status, SS and NS-398 caused growth arrest in a dose-related manner. High dose (10(-4)M) of SS but not of NS-398, increased the percentage of subG1 cell population while reducing mitochondrial transmembrane potential (DeltaPsim). Cyclosporin A (CsA, 1 microM) prevented collapse of DeltaPsim induced by 10(-4)M SS but not by 7.5 microM FCCP used as a protonophoric control. SS and FCCP increased the cytosolic release of Smac/DIABLO which was differently affected by CsA pretreatment depending on the uncoupler. Finally, 7.5 microM FCCP failed to induce apoptosis whereas CsA prevented apoptosis induced by SS from 16% in HCT15 to 41% in HCT116. The present study shows that despite the ability of sulindac sulfide to behave as a protonophoric uncoupler, CsA-sensitive opening of mitochondrial permeability transition pore contributes little to its pro-apoptotic effect in colorectal cancer cells.
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Anti-Inflamatórios não Esteroides/farmacologia , Apoptose , Proteínas de Transporte/metabolismo , Neoplasias Colorretais/patologia , Proteínas Mitocondriais/metabolismo , Sulindaco/análogos & derivados , Sulindaco/farmacologia , Proteínas Reguladoras de Apoptose , Quimioprevenção , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Nitrobenzenos/farmacologia , Oxirredução , Fosforilação , Sulfonamidas/farmacologia , Células Tumorais Cultivadas , Desacopladores/farmacologiaRESUMO
The liver is the most frequent and fatal site of distant spreading of colorectal cancer. Most liver metastases animal models involve nude mice and an injection of tumour cells through the spleen or portal vein, or orthotopic implantation of tumour cells in the colon wall. The aim of this study was to develop a reliable rat model of liver metastases with human colorectal HT29 cells. Seventy male athymic Rowett nude rats weighing 300+/-30 g were separated into three groups. The first group (n=20) consisted of untreated rats, rats in the second group (n=20) were immunosuppressed by cyclosporin A, and those in the third group (n=30) were irradiated the day before cell grafting. Tumour cells (2 x 10(7)) were subcapsulary injected into the liver, and rats were sacrificed after 60 days. The livers were excised, and tumours were serially sectioned to determine size and volume, then fixed for histological studies. The take-rate was 55% in the first group, 35% in the second and 74% in the third group. The mean volume of tumours in the first group was 537+/-162 mm(3), 613+/-232 mm(3) in the second group and 2949+/-629 mm(3) in the third group. In conclusion, subcapsular injection of the human colonic HT29 cancer cells into the liver of preoperatively irradiated nude rats is a reliable, reproducible and easily obtained model, which should be useful for preclinical studies.
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Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Neoplasias Hepáticas Experimentais/secundário , Animais , Humanos , Masculino , Ratos , Ratos Nus , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
The development of chemotherapy using conventional anticancer drugs has been hindered due to several drawbacks related to their poor water solubility and poor pharmacokinetics, leading to severe adverse side effects and multidrug resistance in patients. Nanocarriers were developed to palliate these problems by improving drug delivery, opening the era of nanomedicine in oncology. Liposomes have been by far the most used nanovectors for drug delivery, with liposomal doxorubicin receiving US FDA approval as early as 1995. Antibody drug conjugates and promising drug delivery systems based on a natural polymer, such as albumin, or a synthetic polymer, are currently undergoing advanced clinical trials or have received approval for clinical applications. However, despite attractive results being obtained in preclinical studies, many well-designed nanodrugs fell short of expectations when tested in patients, evidencing the gap between nanoparticle design and their clinical translation. The aim of this review is to evaluate the extent of nanotherapeutics used in oncology by providing an insight into the most successful concepts. The reasons that prevent nanodrugs from expanding to clinic are discussed, and the efforts that must be taken to take full advantage of the great potential of nanomedicine are highlighted.
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Antineoplásicos/administração & dosagem , Antineoplásicos/uso terapêutico , Portadores de Fármacos/química , Nanomedicina , Nanopartículas/administração & dosagem , Nanopartículas/uso terapêutico , Neoplasias/tratamento farmacológico , Antineoplásicos/química , Antineoplásicos/farmacologia , Doxorrubicina/administração & dosagem , Doxorrubicina/química , Doxorrubicina/uso terapêutico , Portadores de Fármacos/administração & dosagem , Humanos , Lipossomos , Nanopartículas/químicaRESUMO
BACKGROUND: Inflammatory bowel diseases are incurable illnesses of the gastrointestinal tract, which substantially enhance the risk of developing colorectal carcinogenesis. Conventional photodynamic therapy is a clinically approved therapeutic modality used in the treatment of neoplastic diseases. Recent preclinical and clinical studies have shown that photodynamic therapy with low doses of photosensitizer and/or light improves inflammatory conditions, including colitis. This study aims therefore at investigating the therapeutic potential of low-dose photodynamic therapy (LDPDT) with a liposomal formulation of meta-tetra(hydroxyphenyl)chlorin (namely Foslip) in the prevention of colitis-associated cancer in mice. METHODS: LDPDT efficacy was evaluated by endoscopic, macroscopic, and histological analysis. Myeloperoxidase levels were quantified by enzyme linked immunosorbent assay and cytokines expression by quantitative RT-PCR analysis. The integrity of the intestinal barrier was evaluated by immunostaining, and bacterial composition of the fecal microbiota was determined by 454 pyrosequencing of V3-V4 region of bacterial 16S rRNA genes. RESULTS: LDPDT reduced intestinal tumor growth by decreasing the expression of a wide range of inflammatory mediators and by lowering neutrophil influx. LDPDT treatment prevents onset of a dysbiotic microbiota in the colitis-associated cancer model. CONCLUSIONS: LDPDT with Foslip could be considered as a novel treatment modality to prevent colorectal carcinogenesis in patients with inflammatory bowel disease.
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Colite/complicações , Neoplasias do Colo/prevenção & controle , Mesoporfirinas/uso terapêutico , Fotoquimioterapia , Animais , Colite/induzido quimicamente , Colite/patologia , Neoplasias do Colo/etiologia , Neoplasias do Colo/patologia , Colonoscopia , Citocinas/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Perfilação da Expressão Gênica , Humanos , Técnicas Imunoenzimáticas , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/metabolismo , Linfoma Difuso de Grandes Células B/patologia , Camundongos , Camundongos Endogâmicos C57BL , Fármacos Fotossensibilizantes/uso terapêuticoRESUMO
To determine the initial photodamage sites of Foscan-mediated photodynamic treatment, we evaluated the enzymatic activities in selected organelles immediately after light exposure of MCF-7 cells. The measurements indicated that the enzymes located in the Golgi apparatus (uridine 5'-diphosphate galactosyl transferase) and in the endoplasmic reticulum (ER) (nicotinamide adenine dinucleotide [reduced] [NADH] cytochrome c [cyt c] reductase) are inactivated by the treatment, whereas mitochondrial marker enzymes (cyt c oxidase and dehydrogenases) were unaffected. This indicates that the ER and the Golgi apparatus are the primary intracellular sites damaged by Foscan-mediated PDT in MCF-7 cells. We further investigated whether the specific mitochondria events could be associated with Foscan photoinduced cell death. The dose response profiles of mitochondrial depolarization and cytochrome c release immediately after Foscan-based PDT were very different from that of overall cell death. By 24 h post-PDT the fluence dependency was strikingly similar for both mitochondrial alterations and cell death. Therefore, although mitochondria are not directly affected by the treatment, they can be strongly implicated in Foscan-mediated MCF-7 cell death by late and indirect mechanism.
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Neoplasias da Mama/tratamento farmacológico , Mesoporfirinas/farmacologia , Mitocôndrias/efeitos da radiação , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/metabolismo , Apoptose , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Sobrevivência Celular , Relação Dose-Resposta a Droga , Retículo Endoplasmático/enzimologia , Feminino , Complexo de Golgi/enzimologia , Humanos , Potenciais da Membrana/efeitos dos fármacos , Mesoporfirinas/uso terapêutico , Mitocôndrias/enzimologia , Fármacos Fotossensibilizantes/uso terapêutico , Fatores de TempoRESUMO
Nucleic acid aptamers are molecules that are being used in a large number of biomedical applications. Aptamers have the properties to bind to a wide range of molecules with high specificity and affinity for their target. These properties together with their small size and their ease of synthesis make them very attractive and promising for targeting diseases and therapeutic applications. Aptamers can serve as cancer diagnostic tools by detecting specific biomarkers, circulating cancer cells or imaging diseased tissue. On the other hand, aptamers can be used as therapeutic agents due to their potential antagonist activity, or as targeting agents. Therefore, they can be designed to deliver antitumor molecules such as chemotherapeutic drugs, siRNA or photodynamic therapy sensitizers to diseased tissues. Attempts are also made to synthesize aptamers-targeted nanoplatforms capable to ferry cargo and load onto them both imaging and therapeutic functions creating so called nanotheragnostics agents. In the future, its seems likely that aptamers will play an important role in diagnosis and treatment of several pathologies including cancer.
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Aptâmeros de Nucleotídeos/uso terapêutico , Neoplasias/tratamento farmacológico , Animais , Aptâmeros de Nucleotídeos/farmacocinética , Humanos , Neoplasias/diagnóstico , Técnica de Seleção de Aptâmeros , Distribuição TecidualRESUMO
Photodynamic therapy (PDT), an approved anticancer treatment, is reported as a potent inducer of programmed cell death (PCD) by both apoptosis and autophagy. The present study investigated the kinetics of both autophagy and caspase activation in MCF-7 cells submitted to mTHPC-PDT upon condition of treatment promoting ER accumulation of mTHPC. Fluence-dependent immediate cytochrome c (cyt C) release followed by caspase-9 and -7 activation at 1 h post-PDT evidenced a mitochondrial oxidative stress triggered by high light doses leading to >90% of cell death. ER oxidative stress was monitored by the induction of the glucose-related protein chaperone GRP78. From 6 h post-PDT, GRP78 induction was accompanied by the conversion of LC3-I into LC3-II, the hallmark of autophagosome formation. The formation of acid vesicles evidenced by fluorescence microscopy was obvious from 22 h post-PDT. Twenty-four hours post-PDT, cyt C release decreased and caspase-9 cleavage disappeared, while the expression of cleaved caspase-7 remained significant. At the same time, the profiles of GRP78, cleaved caspase-7 and LC3-II expression were similar irrespective of light doses. In contrast to an inhibitor of caspase activation Z-VAD-FMK, the use of autophagy inhibitor, Wortmannin, impaired cytotoxicity along with an increase in caspase-7 activation. These results demonstrate a valuable contribution of autophagy to cell death in mTHPC-photosensitized MCF-7 cells.
Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Estresse do Retículo Endoplasmático , Mesoporfirinas/farmacologia , Mitocôndrias/efeitos dos fármacos , Fotoquimioterapia , Fármacos Fotossensibilizantes/farmacologia , Clorometilcetonas de Aminoácidos/farmacologia , Androstadienos/farmacologia , Inibidores de Caspase , Caspases/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Retículo Endoplasmático/efeitos dos fármacos , Chaperona BiP do Retículo Endoplasmático , Humanos , Cinética , Proteínas Associadas aos Microtúbulos/metabolismo , Estresse Oxidativo , Poli(ADP-Ribose) Polimerases/metabolismo , Inibidores de Proteínas Quinases/farmacologia , WortmaninaRESUMO
PURPOSE: The present study investigates the efficacy of compartmental targeting in xenografted tumors treated by meta-tetra(hydroxyphenyl)chlorin (mTHPC)-mediated photodynamic therapy (PDT). The therapeutic efficacy was, furthermore, related to a regional photoinduced distribution of apoptosis and an mTHPC biodistribution profile. METHODS AND MATERIALS: Mice bearing EMT6 tumors were subjected to a single irradiation (10 J/cm(2)) of red laser light (652 nm) at different intervals after a single- (0.3 mg/kg or 0.15 mg/kg) or double-intravenous (2 × 0.15 mg/kg) injection(s) of mTHPC. Efficiency of the treatment was evaluated by monitoring tumor regrowth. mTHPC pharmacokinetics were assessed by high-performance liquid chromatography analysis of excised organs. The regional distribution of apoptosis in tumor sections was investigated with a newly developed colabelling immunohistochemistry technique. RESULTS: A fractionated double-injection protocol of mTHPC with 24-h and 3-h drug-light intervals (DLI) yielded 100% tumor cure, with tumors presenting a massive apoptosis of neoplastic cells along with a distortion of vessels. The best efficiency for a single injection (0.3 mg/kg) was about 54% tumor cure and corresponded to a DLI of 3 h. At this DLI, tumors showed apoptosis of endothelial cells in residual vessels. Concentrations of mTHPC observed in plasma and tumor for the fractionated injection were not statistically different and were less than the total drug dose in each compartment. CONCLUSIONS: The present work suggests that clinical PDT protocols with mTHPC could be greatly improved by fractionation of the drug administration. Time points should be chosen based on the intratumoral spatiotemporal drug distribution.