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Dysregulated iron homeostasis underlies diverse pathologies, from ischemia-reperfusion injury to epithelial-mesenchymal transition and drug-tolerant "persister" cancer cell states. Here, we introduce ferrous iron-activatable luciferin-1 (FeAL-1), a small-molecule probe for bioluminescent imaging of the labile iron pool (LIP) in luciferase-expressing cells and animals. We find that FeAL-1 detects LIP fluctuations in cells after iron supplementation, depletion, or treatment with hepcidin, the master regulator of systemic iron in mammalian physiology. Utilizing FeAL-1 and a dual-luciferase reporter system, we quantify LIP in mouse liver and three different orthotopic pancreatic ductal adenocarcinoma tumors. We observed up to a 10-fold increase in FeAL-1 bioluminescent signal in xenograft tumors as compared to healthy liver, the major organ of iron storage in mammals. Treating mice with hepcidin further elevated hepatic LIP, as predicted. These studies reveal a therapeutic index between tumoral and hepatic LIP and suggest an approach to sensitize tumors toward LIP-activated therapeutics.
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Hierro , Neoplasias , Humanos , Ratones , Animales , Hepcidinas , Luciferinas , Xenoinjertos , Hígado , Luciferasas , MamíferosRESUMEN
The programmed cell death protein-1 (PD-1) is highly expressed on the surface of antigen-specific exhausted T cells and, upon interaction with its ligand PD-L1, can result in inhibition of the immune response. Anti-PD-1 treatment has been shown to extend survival and result in durable responses in several cancers, yet only a subset of patients benefit from this therapy. Despite the implication of metabolic alteration following cancer immunotherapy, mechanistic associations between antitumor responses and metabolic changes remain unclear. Here, we used desorption electrospray ionization mass spectrometry imaging to examine the lipid profiles of tumor tissue from three syngeneic murine models with varying treatment sensitivity at the baseline and at three time points post-anti-PD-1 therapy. These imaging experiments revealed specific alterations in the lipid profiles associated with the degree of response to treatment and allowed us to identify a significant increase of long-chain polyunsaturated lipids within responsive tumors following anti-PD-1 therapy. Immunofluorescence imaging of tumor tissues also demonstrated that the altered lipid profile associated with treatment response is localized to dense regions of tumor immune infiltrates. Overall, these results indicate that effective anti-PD-1 therapy modulates lipid metabolism in tumor immune infiltrates, and we thereby propose that further investigation of the related immune-metabolic pathways may be useful for better understanding success and failure of anti-PD-1 therapy.
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Anticuerpos Monoclonales , Antígeno B7-H1 , Neoplasias , Animales , Humanos , Ratones , Anticuerpos Monoclonales/uso terapéutico , Antígeno B7-H1/antagonistas & inhibidores , Inmunoterapia , Lípidos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Linfocitos T/metabolismo , Microambiente TumoralRESUMEN
Chemotherapeutic treatment with conventional drug formulations pose numerous challenges, such as poor solubility, high cytotoxicity and serious off-target side effects, low bioavailability, and ultimately subtherapeutic tumoral concentration leading to poor therapeutic outcomes. In the field of Nanomedicine, advances in nanotechnology have been applied with great success to design and develop novel nanoparticle-based formulations for the treatment of various types of cancer. The approval of the first nanomedicine, Doxil® (liposomal doxorubicin) in 1995, paved the path for further development for various types of novel delivery platforms. Several different types of nanoparticles, especially organic (soft) nanoparticles (liposomes, polymeric micelles, and albumin-bound nanoparticles), have been developed and approved for several anticancer drugs. Nanoparticulate drug delivery platform have facilitated to overcome of these challenges and offered key advantages of improved bioavailability, higher intra-tumoral concentration of the drug, reduced toxicity, and improved efficacy. This review introduces various commonly used nanoparticulate systems in biomedical research and their pharmacokinetic (PK) attributes, then focuses on the various physicochemical and physiological factors affecting the in vivo disposition of chemotherapeutic agents encapsulated in nanoparticles in recent years. Further, it provides a review of the current landscape of soft nanoparticulate formulations for the two most widely investigated anticancer drugs, paclitaxel, and doxorubicin, that are either approved or under investigation. Formulation details, PK profiles, and therapeutic outcomes of these novel strategies have been discussed individually and in comparison, to traditional formulations. This article is categorized under: Nanotechnology Approaches to Biology > Cells at the Nanoscale Diagnostic Tools > In Vivo Nanodiagnostics and Imaging Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease.
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Antineoplásicos , Nanopartículas , Neoplasias , Humanos , Sistemas de Liberación de Medicamentos , Antineoplásicos/uso terapéutico , Liposomas/uso terapéutico , Doxorrubicina/uso terapéutico , Neoplasias/tratamiento farmacológicoRESUMEN
Palbociclib, ribociclib and abemaciclib were recently approved as chemotherapeutic agents and are currently in the post-marketing surveillance phase. They are used in combination with aromatase inhibitors anastrozole and letrozole or antiestrogen fulvestrant for HR+, HER2- breast cancer treatment. Here, a novel bioanalytical LC-ESI-MS/MS method was developed for the quantitation of these six drugs in human plasma. The samples were prepared by simple protein precipitation followed by solvent evaporation. A Kinetex biphenyl column (150 × 4.6 mm, 2.6 µm) used for chromatographic analysis adequately resolved even the closely eluting aromatase inhibitors' peaks. The mobile phase consisted of 0.1% formic acid in water and in ACN, in a linear gradient. An additional gradient step was added to eliminate the observed carry-over. The proposed method was fully validated in the relevant linear ranges covering the expected plasma concentrations of all six drugs (correlation coefficients between 0.9996 and 0.9931). The intra-day method precision (CV) ranged from 3.1% to 15%, while intra-day accuracy (%bias) was between -1.5% and 15.0%. The inter-day precision ranged from 1.6% to 14.9%, with accuracy between -14.3% and 14.6%, which is in accordance with the EMA and ICH guidelines on bioanalytical method validation. The method was successfully applied to samples from patients treated for HR+, HER2- breast cancer.
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A liquid chromatography-tandem mass spectrometry assay was developed and qualified for the multiplexed quantitation of a small molecule stimulator of soluble guanylate cyclase (sGC) and its target engagement biomarker, 3',5'-cyclic guanosine monophosphate (cGMP), in ocular tissues and plasma from a single surrogate matrix calibration curve. A surrogate matrix approach was used in this assay due to the limited quantities of blank ocular matrices in a discovery research setting. After optimization, the assay showed high accuracy, precision, and recovery as well as parallelism between the surrogate matrix and the biological matrices (rabbit plasma, vitreous, and retina-choroid). This assay provided pharmacokinetic and target engagement data after intravitreal administration of the sGC stimulator. The nitric oxide-sGC-cGMP pathway is a potential target to address glaucoma. Increasing sGC-mediated production of cGMP could improve aqueous humor outflow and ocular blood flow. The sGC stimulator showed dose-dependent exposure in rabbit vitreous, retina-choroid, and plasma. The cGMP exhibited a delayed yet sustained increase in vitreous humor but not retina-choroid. Multiplexed measurement of both pharmacokinetic and target engagement analytes reduced animal usage and provided improved context for interpreting PK and PD relationships.
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GMP Cíclico , Guanilato Ciclasa , Animales , Guanilato Ciclasa/metabolismo , Óxido Nítrico , Conejos , Transducción de Señal , Guanilil Ciclasa Soluble/metabolismoRESUMEN
BACKGROUND: GMC1 (2-(1H-benzimidazol-2-ylsulfanyl)-N-[(Z)-(4-methoxyphenyl) methylideneamino] acetamide) effectively inhibits androgen receptor function by binding directly to FKBP52. This is a novel mechanism for the treatment of castration resistant prostate cancer (CRPC). METHODS: an LC-MS/MS method was developed and validated to quantify GMC1 in plasma and urine from pharmacokinetics studies in rats. An ultra-high-performance liquid chromatography (UHPLC) system equipped with a Waters XTerra MS C18 column was used for chromatographic separation by gradient elution with 0.1% (v/v) formic acid in water and methanol. A Sciex 4000 QTRAP® mass spectrometer was used for analysis by multiple reaction monitoring (MRM) in positive mode; the specific ions [M+H]+m/z 340.995 â m/z 191.000 and [M+H]+ m/z 266.013 â m/z 234.000 were monitored for GMC1 and internal standard (albendazole), respectively. RESULTS: GMC1 and albendazole had retention times of 1.68 and 1.66 min, respectively. The calibration curves for the determination of GMC1 in rat plasma and urine were linear from 1-1000 ng/mL. The LC-MS/MS method was validated with intra- and inter-day accuracy and precision within the 15% acceptance limit. The extraction recovery values of GMC1 from rat plasma and urine were greater than 95.0 ± 2.1% and 97.6 ± 4.6%, respectively, with no significant interfering matrix effect. GMC1 is stable under expected sample handling, storage, preparation and LC-MS/MS analysis conditions. CONCLUSIONS: Pharmacokinetic evaluation of GMC1 revealed that the molecule has a biexponential disposition in rats, is distributed rapidly and extensively, has a long elimination half-life, and appears to be eliminated primarily by first order kinetics.
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INTRODUCTION: CLBQ14, a derivative of 8-hydroxyquinoline, exerts its chemotherapeutic effect by inhibiting methionine aminopeptidase (MetAP), the enzyme responsible for the post-translational modification of several proteins and polypeptides. MetAP is a novel target for infectious diseases. CLBQ14 is selective and highly potent against replicating and latent Mycobacterium tuberculosis making it an appealing lead for further development. METHODS: The physicochemical properties (solubility, pH stability and lipophilicity), in vitro plasma stability and metabolism, pre-clinical pharmacokinetics, plasma protein binding and tissue distribution of CLBQ14 in adult male Sprague-Dawley rats were characterized. RESULTS: At room temperature, CLBQ14 is practically insoluble in water (<0.07 mg/mL) but freely soluble in dimethyl acetamide (>80 mg/mL); it has a log P value of 3.03 ± 0.04. CLBQ14 exhibits an inverse Z-shaped pH decomposition profile; it is stable at acidic pH but is degraded at a faster rate at basic pH. It is highly bound to plasma proteins (>91%), does not partition to red blood cells (B/P ratio: 0.83 ± 0.03), and is stable in mouse, rat, monkey and human plasma. CLBQ14 exhibited a bi-exponential pharmacokinetics after intravenous administration in rats, bioavailability of 39.4 and 90.0%, respectively from oral and subcutaneous route. We observed a good correlation between predicted and observed rat clearance, 1.90 ± 0.17 L/kg/h and 1.67 ± 0.08 L/kg/h, respectively. Human hepatic clearance predicted from microsomal stability data and from the single species scaling were 0.80 L/hr/kg and 0.69 L/h/kg, respectively. CLBQ14 is extensively distributed in rats; following a 5 mg/kg intravenous administration, lowest and highest concentrations of 15.6 ± 4.20 ng/g of heart and 405.9 ± 77.11 ng/g of kidneys, respectively, were observed. In vitro CYP reaction phenotyping demonstrates that CLBQ14 is metabolized primarily by CYP 1A2. CONCLUSION: CLBQ14 possess appealing qualities of a drug candidate. The studies reported herein are imperative to the development of CLBQ14 as a new chemical entity for infectious diseases.
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Enfermedades Transmisibles/tratamiento farmacológico , Inhibidores Enzimáticos/farmacocinética , Metionil Aminopeptidasas/antagonistas & inhibidores , Oxiquinolina/análogos & derivados , Animales , Química Física , Enfermedades Transmisibles/metabolismo , Inhibidores Enzimáticos/sangre , Inhibidores Enzimáticos/química , Corazón , Humanos , Riñón , Macaca fascicularis , Masculino , Metionil Aminopeptidasas/metabolismo , Ratones , Estructura Molecular , Oxiquinolina/sangre , Oxiquinolina/química , Oxiquinolina/farmacocinética , Ratas , Ratas Sprague-Dawley , Termodinámica , Distribución TisularRESUMEN
CLBQ14 is an 8-hydroxyquinoline analogue that inhibits methionine aminopeptidase (MetAP), an enzyme responsible for the post-translational modification of several proteins and polypeptides. MetAP has been validated as druggable target for some infectious diseases, and its inhibitors have been investigated as potential therapeutic agents. In this study, we developed and validated a liquid chromatography tandem-mass spectrometry (LC-MS/MS) method for the quantification of CLBQ14 in solution, and in rat plasma and urine. This method was applied to the pharmacokinetic evaluation of CLBQ14 in adult male Sprague Dawley (SD) rats. Chromatographic separation was achieved using an ultra-high-performance liquid chromatography (UHPLC) system equipped with Waters XTerra MS C18 column (3.5⯵m, 125â¯Å, 2.1â¯×â¯50â¯mm) using 0.1% formic acid in acetonitrile/water gradient system as mobile phase. Chromatographic analysis was performed with a 4000 QTRAP® mass spectrometer using MRM in positive mode for CLBQ14 transition [Mâ¯+â¯H]+m/z 257.919â¯ââ¯m/z 151.005, and IS (clioquinol) transition [Mâ¯+â¯H]+m/z 305.783â¯ââ¯m/z 178.917. CLBQ14 was extracted from plasma and urine samples by protein precipitation. The retention times for CLBQ14 and IS were 1.31 and 1.40â¯min respectively. The standard curves were linear for CLBQ14 concentration ranging from 1 to 1000â¯ng/mL. The intra-day and inter-day accuracy and precision were found to be within 15% of the nominal concentration. Extraction recoveries were >96.3% and 96.6% from rat plasma and urine respectively, and there was no significant matrix effect from the biological matrices. CLBQ14 is stable in samples subjected to expected storage, preparation, and handling conditions. Pharmacokinetic studies revealed that CLBQ14 has a bi-exponential disposition in SD rats, is extensively distributed with a long plasma half-life and is eliminated primarily by liver metabolism.
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Cloroquinolinoles/farmacocinética , Cromatografía Liquida/métodos , Espectrometría de Masas en Tándem/métodos , Animales , Cloroquinolinoles/sangre , Cloroquinolinoles/química , Cloroquinolinoles/orina , Estabilidad de Medicamentos , Modelos Lineales , Masculino , Ratas , Ratas Sprague-Dawley , Reproducibilidad de los ResultadosRESUMEN
Tumor hypoxia is a well-recognized driver of resistance to traditional cancer therapies such as chemotherapy and radiation therapy. We describe development of a new nanoconstruct composed of gold nanorods (GNRs) conjugated to carbonic anhydrase IX (CAIX) antibody that specifically binds to CAIX, a biomarker of hypoxia, to facilitate targeting tumor hypoxic areas for focused photothermal ablation. Physicochemical characterization studies confirmed the size, shape, monodispersity, surface charge, and serum stability of the GNRs. Enzyme-linked immunosorbent assays and cellular binding and uptake studies confirmed successful conjugation of antibody to the GNRs and specificity for CAIX. Near-infrared irradiation of CAIX-overexpressing cells treated with GNR/anti-CAIX resulted in significantly higher cell death than cells treated with control GNRs. In vivo biodistribution studies using hyperspectral imaging and inductively coupled plasma mass spectrometry confirmed intravenous administration results not only in greater accumulation of GNR/anti-CAIX in tumors than control GNRs but also greater penetration into hypoxic areas of tumors. Near-infrared ablation of these tumors showed no tumor regression in the sham-treated group, regression but recurrence in the non-targeted-GNR group, and complete tumor regression in the targeted-GNR group. GNR/anti-CAIX nanoconstructs show promise as hypoxia targeting and photothermal ablation agents for cancer treatment.