RESUMO
We report the synthesis of biocompatible perfluorinated micelles designed to improve radiotherapeutic efficacy in a radioresistant tumor environment. In vitro and in vivo behaviors of perfluorinated micelles were assessed at both cellular and tissular levels. The micellar platform offers key advantages as theranostic tool: (i) small size, allowing deep tissue penetration; (ii) oxygen transport to hypoxic tissues; (iii) negligible toxicity in the absence of ionizing radiation; (iv) internalization into cancer cells; (v) potent radiosensitizing effect; and (vi) excellent tumor-targeting properties, as monitored by positron emission tomography. We have demonstrated strong in vitro radiosensitizing effects of the micelle and in vivo tumor targeting, making this nanometric carrier a promising tool for the potentiation of focused radiotherapy.
Assuntos
Micelas , Tomografia por Emissão de Pósitrons , Radiossensibilizantes , Nanomedicina Teranóstica , Animais , Humanos , Radiossensibilizantes/química , Radiossensibilizantes/farmacologia , Radiossensibilizantes/síntese química , Camundongos , Linhagem Celular Tumoral , Fluorocarbonos/química , Fluorocarbonos/farmacologia , Neoplasias/diagnóstico por imagem , Neoplasias/tratamento farmacológico , Neoplasias/patologiaRESUMO
Biomolecules labeled with positron-emitting radionuclides like fluorine-18 or radiometals like copper-64 and zirconium-89 are increasingly employed in nuclear medicine for diagnosis purposes. Given the fragility and complexity of these compounds, their labeling requires mild conditions. Besides, it is essential to develop methods inducing minimal modification of the tertiary structure, as it is fundamental for the biological activity of such complex entities. Given these requirements, disulfide rebridging represents a promising possibility since it allows protein modification as well as conservation of the tertiary structure. In this context, we have developed an original radiofluorinated dibromopyridazine dione prosthetic group for labeling of disulfide-containing biomolecules via rebridging. We employed it to radiolabel octreotide, a somatostatin analogue, and to radiolabel fragment antigen binding (Fab) targeting programmed death-ligand 1 (PD-L1), whose properties were then evaluated in vitro and in vivo by positron emission tomography (PET) imaging. We next extended our strategy to the radiolabeling of cetuximab, a monoclonal antibody, with various radiometals commonly used in PET imaging (zirconium-89, copper-64) by developing various rebridging molecules bearing the appropriate chelators. The stabilities of the radiolabeled antibody conjugates were assessed in biological conditions.
Assuntos
Radioisótopos de Cobre , Radioisótopos de Flúor , Radioisótopos , Zircônio , Radioisótopos de Cobre/química , Radioisótopos de Flúor/química , Tomografia por Emissão de Pósitrons/métodos , Compostos RadiofarmacêuticosRESUMO
Tumor-specific drug delivery is a major challenge for the pharmaceutical industry. Nanocarrier systems have been widely investigated to increase and control drug delivery to the heterogeneous tumor microenvironment. Classically, the uptake of nanocarriers by solid tumor tissues is mainly mediated by the enhanced permeability and retention effect (EPR). This EPR effect depends on the tumor type, its location, the physicochemical properties of the carriers, and the blood perfusion of the tumoral lesions. The main goal of this study was to evaluate in vivo tumor uptake of micelle carriers, assisted by microbubble/ultrasound sonoporation. Micelles were tracked using bi-modal imaging techniques to precisely localize both the nanocarrier and its payload. Micelles were loaded with a near infrared fluorophore and radiolabeled with zirconium-89. Their pharmacokinetics, biodistribution and passive tumor targeting properties were evaluated in a subcutaneous glioblastoma (U-87 MG) mouse model using optical and PET imaging. Finally, accumulation and diffusion into the tumor micro-environment was investigated under microbubble-assisted sonoporation, which helped homogenize the delivery of the micelles. The in vivo experiments showed a good correlation between optical and PET images and demonstrated the stability of the micelles in biological media, their high and long-term retention in the tumors and their clearance through the hepato-biliary pathway. This study demonstrates that bi-modal imaging techniques are powerful tools for the development of new nanocarriers and that sonoporation is a promising method to homogenize nanomedicine delivery to tumors.
Assuntos
Glioma , Micelas , Camundongos , Animais , Distribuição Tecidual , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos/métodos , Glioma/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Portadores de Fármacos/química , Microambiente TumoralRESUMO
BACKGROUND: The resistance of glioblastoma stem cells (GSCs) to treatment is one of the causes of glioblastoma (GBM) recurrence. Endothelin A receptor (ETA) overexpression in GSCs constitutes an attractive biomarker for targeting this cell subpopulation, as illustrated by several clinical trials evaluating the therapeutic efficacy of endothelin receptor antagonists against GBM. In this context, we have designed an immunoPET radioligand combining the chimeric antibody targeting ETA, chimeric-Rendomab A63 (xiRA63), with 89Zr isotope and evaluated the abilities of xiRA63 and its Fab (ThioFab-xiRA63) to detect ETA+ tumors in a mouse model xenografted orthotopically with patient-derived Gli7 GSCs. RESULTS: Radioligands were intravenously injected and imaged over time by µPET-CT imaging. Tissue biodistribution and pharmacokinetic parameters were analyzed, highlighting the ability of [89Zr]Zr-xiRA63 to pass across the brain tumor barrier and achieve better tumor uptake than [89Zr]Zr-ThioFab-xiRA63. CONCLUSIONS: This study shows the high potential of [89Zr]Zr-xiRA63 in specifically targeting ETA+ tumors, thus raising the possibility of detecting and treating ETA+ GSCs, which could improve the management of GBM patients.
Assuntos
Glioblastoma , Animais , Camundongos , Humanos , Glioblastoma/diagnóstico por imagem , Receptor de Endotelina A , Tomografia por Emissão de Pósitrons/métodos , Distribuição Tecidual , Anticorpos , Células-Tronco , Linhagem Celular Tumoral , ZircônioRESUMO
Positron emission tomography (PET) imaging of the myelin sheath is a powerful tool to investigate multiple sclerosis, monitor its evolution, and support drug development. Radiotracers based on N,N-dimethylaminostilbene (MeDAS) fluorinated analogs have been designed for myelin PET imaging but were never translated to humans. We have synthesized three original fluorinated analogs of MeDAS with low metabolic rates for which binding to myelin in a healthy rat brain was demonstrated by fluorescence microscopy. A tosyl precursor was synthesized for the lead compound PEGMeDAS and automated fluorine-18 radiolabeling afforded [18F]PEGMeDAS in 25 ± 5% radiochemical yield and 102 ± 15 GBq/µmol molar activity. Biodistribution in healthy rats demonstrated the brain penetration with low penetration of radiometabolites. However, E to Z isomerization observed in plasma hampers further investigations of this family of molecules and requires complementary data on the in vivo behavior of the Z isomer.
Assuntos
Esclerose Múltipla , Bainha de Mielina , Ratos , Humanos , Animais , Bainha de Mielina/metabolismo , Distribuição Tecidual , Tomografia por Emissão de Pósitrons/métodos , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Esclerose Múltipla/metabolismo , Compostos Radiofarmacêuticos , Radioisótopos de Flúor/químicaRESUMO
Personalized medicine approach in radiotherapy requires the delivery of precise dose to the tumor. The concept is to increase the effectiveness of radiotherapy while sparing the surrounding heathy tissue. This can be achieved by the use of high-Z metal-based nanoparticles (NPs) as radio-enhancers and PET imaging for mapping NPs distribution to guide the irradiation. In the present study, radio-enhancing platinum NPs were radiolabeled and imaged to assess their pharmacokinetics over time. PET imaging of these NPs revealed high enhanced permeation and retention effect. The maximal tumor accumulation (4.8 ± 0.8 %ID/cc) was observed at 24 h post-injection along with persistent accumulation of the NPs, especially at the tumor ring, even after several days. These properties positively suggest the potential clinical use of these NPs.
Assuntos
Nanopartículas Metálicas , Nanopartículas , Neoplasias , Humanos , Platina , Tomografia por Emissão de Pósitrons/métodos , Distribuição TecidualRESUMO
Checkpoint inhibitors have revolutionized cancer treatment. However, only a minority of patients respond to these immunotherapies. Here, we report that blocking the inhibitory NKG2A receptor enhances tumor immunity by promoting both natural killer (NK) and CD8+ T cell effector functions in mice and humans. Monalizumab, a humanized anti-NKG2A antibody, enhanced NK cell activity against various tumor cells and rescued CD8+ T cell function in combination with PD-x axis blockade. Monalizumab also stimulated NK cell activity against antibody-coated target cells. Interim results of a phase II trial of monalizumab plus cetuximab in previously treated squamous cell carcinoma of the head and neck showed a 31% objective response rate. Most common adverse events were fatigue (17%), pyrexia (13%), and headache (10%). NKG2A targeting with monalizumab is thus a novel checkpoint inhibitory mechanism promoting anti-tumor immunity by enhancing the activity of both T and NK cells, which may complement first-generation immunotherapies against cancer.
Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Carcinoma de Células Escamosas , Cetuximab/uso terapêutico , Imunidade Celular/efeitos dos fármacos , Imunoterapia , Células Matadoras Naturais/imunologia , Subfamília C de Receptores Semelhantes a Lectina de Células NK , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Carcinoma de Células Escamosas/imunologia , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/terapia , Ensaios Clínicos Fase II como Assunto , Humanos , Células Matadoras Naturais/patologia , Camundongos , Subfamília C de Receptores Semelhantes a Lectina de Células NK/antagonistas & inibidores , Subfamília C de Receptores Semelhantes a Lectina de Células NK/imunologiaRESUMO
The objective of lung gene therapy is to reach the respiratory epithelial cells in order to deliver a functional nucleic acid sequence. To improve the synthetic carrier's efficacy, knowledge of their biodistribution and elimination pathways, as well as cellular barriers faced, depending on the administration route, is necessary. Indeed, the in vivo fate guides the adaptation of their chemical structure and formulation to increase their transfection capacity while maintaining their tolerance. With this goal, lipidic fluorescent probes were synthesized and formulated with cationic lipophosphoramidate KLN47 (KLN: Karine Le Ny). We found that such formulations present constant compaction properties and similar transfection results without inducing additional cytotoxicity. Next, biodistribution profiles of pegylated and unpegylated lipoplexes were compared after systemic injection in mice. Pegylation of complexes led to a prolonged circulation in the bloodstream, whereas their in vivo bioluminescent expression profiles were similar. Moreover, systemic administration of pegylated lipoplexes resulted in a transient liver toxicity. These results indicate that these new fluorescent compounds could be added into lipoplexes in small amounts without perturbing the transfection capacities of the formulations. Such additional properties allow exploration of the in vivo biodistribution profiles of synthetic carriers as well as the expression intensity of the reporter gene.
Assuntos
Amidas/administração & dosagem , Amidas/farmacocinética , Corantes Fluorescentes , Técnicas de Transferência de Genes , Ácidos Fosfóricos/administração & dosagem , Ácidos Fosfóricos/farmacocinética , Amidas/química , Amidas/toxicidade , Animais , Linhagem Celular , Sobrevivência Celular , DNA/química , Corantes Fluorescentes/química , Humanos , Lipossomos , Medições Luminescentes/métodos , Camundongos , Imagem Molecular , Estrutura Molecular , Ácidos Fosfóricos/química , Ácidos Fosfóricos/toxicidade , Plasmídeos/química , Distribuição Tecidual , TransfecçãoRESUMO
Cationic lipids constitute a family of synthetic vectors commonly used for nucleic acids delivery. We herein report the results of a systematic study that aimed to compare the transfection efficacies of cationic lipophosphoramidates possessing either two identical lipid chains (termed symmetric cationic lipids) or two different lipid chains (non-symmetric cationic lipids). In addition, we also compared the transfection results of such a 'molecular approach' (the two different lipid chains being included in the same molecule) with those of a 'supramolecular approach' in which two types of symmetrical cationic lipids were mixed in one liposomal formulation. Thus, the present work allowed us first to optimize the methods used to synthesize non-symmetric cationic lipophosphoramidates. In addition, we could also identify two non-symmetric cationic lipids exhibiting high transfection efficiencies with a series of mammalian cell lines, both vectors being characterized by a single phytanyl chain and either an oleyl or a lauryl lipid chain.
Assuntos
Amidas/química , Técnicas de Transferência de Genes , Lipídeos/química , Ácidos Fosfóricos/química , Cátions/química , Linhagem Celular , Humanos , Estrutura MolecularRESUMO
Lipophosphoramidates have previously been identified as efficient vectors for gene delivery. The incorporation of functional groups that respond to a physiological stimulus is hypothesised to further improve the efficacy of this type of vector and eventually reduce its cytotoxicity. In the present work, we report the effects of the incorporation of two disulfide motifs into the hydrophobic domain, close to the phosphoramidate group. Three cationic vectors possessing such a red/ox sensitive function were synthesised. The capability of one of them (5b) to compact DNA is reported jointly with its ability to release that DNA in the presence of a reducing agent. Finally, compound 5b was tested as a vector for gene delivery into human cells in vitro and its cytotoxicity was also evaluated.
Assuntos
Amidas/química , Dissulfetos/química , Técnicas de Transferência de Genes , Vetores Genéticos/química , Ácidos Fosfóricos/química , Amidas/farmacologia , Cátions/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , DNA/química , DNA/genética , Dissulfetos/farmacologia , Vetores Genéticos/genética , Vetores Genéticos/farmacologia , Células HeLa , Humanos , Oxirredução , Ácidos Fosfóricos/farmacologia , Relação Estrutura-AtividadeRESUMO
The biodistribution of intravenously injected DNA lipid nanocapsules (DNA LNCs), encapsulating pHSV-tk, was analysed by in vivo imaging on an orthotopic melanoma mouse model and by a subsequent treatment with ganciclovir (GCV), using the gene-directed enzyme prodrug therapy (GDEPT) approach. Luminescent melanoma cells, implanted subcutaneously in the right flank of the mice, allowed us to follow tumour growth and tumour localisation with in vivo bioluminescence imaging (BLI). In parallel, DNA LNCs or PEG DNA LNCs (DNA LNCs recovered with PEG(2000)) encapsulating a fluorescent probe, DiD, allowed us to follow their biodistribution with in vivo biofluorescence imaging (BFI). The BF-images confirmed a prolonged circulation-time for PEG DNA LNCs as was previously observed on an ectotopic model of glioma; comparison with BL-images evidenced the colocalisation of PEG DNA LNCs and melanoma cells. After these promising results, treatment with PEG DNA LNCs and GCV on a few animals was performed and the treatment efficacy measured by BLI. The first results showed tumour growth reduction tendency and, once optimised, this therapy strategy could become a new option for melanoma treatment.
Assuntos
DNA/administração & dosagem , Técnicas de Transferência de Genes , Genes Transgênicos Suicidas/genética , Lipídeos/química , Melanoma Experimental/terapia , Imagem Molecular/métodos , Nanocápsulas/química , Animais , Benzotiazóis/administração & dosagem , Benzotiazóis/metabolismo , Carbocianinas/administração & dosagem , Carbocianinas/química , Carbocianinas/metabolismo , Carbocianinas/farmacocinética , Eletroforese em Gel de Ágar , Ácidos Graxos Monoinsaturados/química , Feminino , Corantes Fluorescentes/administração & dosagem , Corantes Fluorescentes/química , Corantes Fluorescentes/metabolismo , Corantes Fluorescentes/farmacocinética , Ganciclovir/uso terapêutico , Glicerol/análogos & derivados , Glicerol/química , Herpes Simples/enzimologia , Herpes Simples/genética , Humanos , Luciferases/genética , Luciferases/metabolismo , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Antígenos Específicos de Melanoma/metabolismo , Camundongos , Camundongos Nus , Octoxinol/química , Ácidos Oleicos/química , Tamanho da Partícula , Fosfatidiletanolaminas/química , Plasmídeos/administração & dosagem , Plasmídeos/genética , Polietilenoglicóis/química , Compostos de Amônio Quaternário/química , Eletricidade Estática , Ácidos Esteáricos/química , Propriedades de Superfície , Timidina Quinase/genética , Distribuição Tecidual , Resultado do Tratamento , Triglicerídeos/química , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The diagnosis of acute hypoxic hepatitis remains problematic. We describe a series of 14 patients who were initially hospitalized in an hepatic care unit with a diagnosis of fulminant hepatitis, and were subsequently found to have acute hypoxic hepatitis ('liver shock') secondary to heart failure. A diagnostic algorithm is proposed.