RESUMEN
Members of the solute carrier (SLC) family of transporters are responsible for the cellular influx of a broad range of endogenous compounds and xenobiotics. These proteins are highly expressed in the gastrointestinal tract and eliminating organs such as the liver and kidney, and are considered to be of particular importance in governing drug absorption and elimination. Many of the same transporters are also expressed in a wide variety of organs targeted by clinically important anticancer drugs, directly affect cellular sensitivity to these agents, and indirectly influence treatment-related side effects. Furthermore, targeted intervention strategies involving the use of transport inhibitors have been recently developed, and have provided promising lead candidates for combinatorial therapies associated with decreased toxicity. Gaining a better understanding of the complex interplay between transporter-mediated on-target and off-target drug disposition will help guide the further development of these novel treatment strategies to prevent drug accumulation in toxicity-associated organs, and improve the safety of currently available treatment modalities. In this report, we provide an update on this rapidly emerging field with particular emphasis on anticancer drugs belonging to the classes of taxanes, platinum derivatives, nucleoside analogs, and anthracyclines.
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Antineoplásicos , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Antineoplásicos/efectos adversos , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/tratamiento farmacológico , Humanos , Proteínas de Transporte de Membrana/metabolismo , Taxoides , XenobióticosRESUMEN
Doxorubicin is a commonly used anticancer agent that can cause debilitating and irreversible cardiac injury. The initiating mechanisms contributing to this side effect remain unknown, and current preventative strategies offer only modest protection. Using stem-cell-derived cardiomyocytes from patients receiving doxorubicin, we probed the transcriptomic landscape of solute carriers and identified organic cation transporter 3 (OCT3) (SLC22A3) as a critical transporter regulating the cardiac accumulation of doxorubicin. Functional validation studies in heterologous overexpression models confirmed that doxorubicin is transported into cardiomyocytes by OCT3 and that deficiency of OCT3 protected mice from acute and chronic doxorubicin-related changes in cardiovascular function and genetic pathways associated with cardiac damage. To provide proof-of-principle and demonstrate translational relevance of this transport mechanism, we identified several pharmacological inhibitors of OCT3, including nilotinib, and found that pharmacological targeting of OCT3 can also preserve cardiovascular function following treatment with doxorubicin without affecting its plasma levels or antitumor effects in multiple models of leukemia and breast cancer. Finally, we identified a previously unrecognized, OCT3-dependent pathway of doxorubicin-induced cardiotoxicity that results in a downstream signaling cascade involving the calcium-binding proteins S100A8 and S100A9. These collective findings not only shed light on the etiology of doxorubicin-induced cardiotoxicity, but also are of potential translational relevance and provide a rationale for the implementation of a targeted intervention strategy to prevent this debilitating side effect.
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Doxorrubicina/efectos adversos , Lesiones Cardíacas/inducido químicamente , Lesiones Cardíacas/tratamiento farmacológico , Terapia Molecular Dirigida , Transportadores de Anión Orgánico Sodio-Independiente/metabolismo , Animales , Niño , Regulación de la Expresión Génica , Lesiones Cardíacas/fisiopatología , Humanos , Ratones , Miocitos Cardíacos/metabolismo , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Transportadores de Anión Orgánico Sodio-Independiente/deficiencia , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Análisis de Secuencia de ARNRESUMEN
Dofetilide is a rapid delayed rectifier potassium current inhibitor widely used to prevent the recurrence of atrial fibrillation and flutter. The clinical use of this drug is associated with increases in QTc interval, which predispose patients to ventricular cardiac arrhythmias. The mechanisms involved in the disposition of dofetilide, including its movement in and out of cardiomyocytes, remain unknown. Using a xenobiotic transporter screen, we identified MATE1 (SLC47A1) as a transporter of dofetilide and found that genetic knockout or pharmacological inhibition of MATE1 in mice was associated with enhanced retention of dofetilide in cardiomyocytes and increased QTc prolongation. The urinary excretion of dofetilide was also dependent on the MATE1 genotype, and we found that this transport mechanism provides a mechanistic basis for previously recorded drug-drug interactions of dofetilide with various contraindicated drugs, including bictegravir, cimetidine, ketoconazole, and verapamil. The translational significance of these observations was examined with a physiologically-based pharmacokinetic model that adequately predicted the drug-drug interaction liabilities in humans. These findings support the thesis that MATE1 serves a conserved cardioprotective role by restricting excessive cellular accumulation and warrant caution against the concurrent administration of potent MATE1 inhibitors and cardiotoxic substrates with a narrow therapeutic window.
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Antiarrítmicos , Fibrilación Atrial , Animales , Antiarrítmicos/farmacología , Humanos , Ratones , Fenetilaminas/farmacología , Sulfonamidas/uso terapéuticoRESUMEN
A multitude of disease and therapy related factors drive the frequent development of kidney disorders in cancer patients. Along with chemotherapy, the newer targeted therapeutics can also cause kidney dysfunction through on and off-target mechanisms. Interestingly, among the small molecule inhibitors approved for the treatment of cancers that harbor BRAF-kinase activating mutations, vemurafenib can trigger tubular damage and acute kidney injury. BRAF is a proto-oncogene involved in cell growth. To investigate the underlying mechanisms, we developed cell culture and mouse models of vemurafenib kidney toxicity. At clinically relevant concentrations vemurafenib induces cell-death in transformed and primary mouse and human kidney tubular epithelial cells. In mice, two weeks of daily vemurafenib treatment causes moderate acute kidney injury with histopathological characteristics of kidney tubular epithelial cells injury. Importantly, kidney tubular epithelial cell-specific BRAF gene deletion did not influence kidney function under normal conditions or alter the severity of vemurafenib-associated kidney impairment. Instead, we found that inhibition of ferrochelatase, an enzyme involved in heme biosynthesis contributes to vemurafenib kidney toxicity. Ferrochelatase overexpression protected kidney tubular epithelial cells and conversely ferrochelatase knockdown increased the sensitivity to vemurafenib-induced kidney toxicity. Thus, our studies suggest that vemurafenib-associated kidney tubular epithelial cell dysfunction and kidney toxicity is BRAF-independent and caused, in part, by off-target ferrochelatase inhibition.
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Ferroquelatasa , Proteínas Proto-Oncogénicas B-raf , Animales , Línea Celular Tumoral , Resistencia a Antineoplásicos , Humanos , Indoles/toxicidad , Riñón/metabolismo , Ratones , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas B-raf/metabolismo , Sulfonamidas/toxicidad , VemurafenibRESUMEN
Recent studies suggest that the ß-blocker drug carvedilol prevents skin carcinogenesis but the mechanism is unknown. Carvedilol is one of a few ß-blockers identified as biased agonist based on an ability to promote ß-arrestin-mediated processes such as ERK phosphorylation. To understand the role of phosphoproteomic signaling in carvedilol's anticancer activity, the mouse epidermal JB6 P+ cells treated with EGF, carvedilol, or their combination were analyzed using the Phospho Explorer Antibody Array containing 1318 site-specific and phospho-specific antibodies of over 30 signaling pathways. The array data indicated that both EGF and carvedilol increased phosphorylation of ERK's cytosolic target P70S6 K while its nuclear target ELK-1 were activated only by EGF; Furthermore, EGF-induced phosphorylation of ELK-1 and c-Jun was attenuated by carvedilol. Subcellular fractionation analysis indicated that ERK nuclear translocation induced by EGF was blocked by co-treatment with carvedilol. Western blot and luciferase reporter assays confirmed that the biased ß-blockers carvedilol and alprenolol blocked EGF-induced phosphorylation and activation of c-Jun/AP-1 and ELK-1. Consistently, both carvedilol and alprenolol strongly prevented EGF-induced neoplastic transformation of JB6 P+ cells. Remarkably, oral carvedilol treatment significantly inhibited the growth of A375 melanoma xenograft in SCID mice. As nuclear translocation of ERK is a key step in carcinogenesis, inhibition of this event is proposed as a novel anticancer mechanism for biased ß-blockers such as carvedilol.
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Antagonistas Adrenérgicos beta/uso terapéutico , Anticarcinógenos/uso terapéutico , Carcinogénesis/efectos de los fármacos , Carvedilol/uso terapéutico , Melanoma/prevención & control , Antagonistas Adrenérgicos beta/farmacología , Animales , Anticarcinógenos/farmacología , Carcinogénesis/metabolismo , Carcinogénesis/patología , Carvedilol/farmacología , Factor de Crecimiento Epidérmico/metabolismo , Células HEK293 , Humanos , Masculino , Melanoma/metabolismo , Melanoma/patología , Ratones Endogámicos NOD , Ratones SCID , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Fosforilación/efectos de los fármacos , Proteoma/metabolismo , Proteínas Proto-Oncogénicas c-jun/metabolismoRESUMEN
RATIONALE: In order to determine the degree of cross-linking on the surface and its variations in a nanometer-scale depth of organic materials, we developed an approach based on time-of-flight secondary ion mass spectrometry (TOF-SIMS), which provides rich chemical information in the form of fragment ions. TOF-SIMS is extremely surface-sensitive and capable of depth profiling with the use of a sputter ion beam to remove controllable amounts of substance. METHODS: Poly(methyl methacrylate) (PMMA) films spin-coated on a Si substrate were cross-linked using a recently developed, surface sensitive, hyperthermal hydrogen projectile bombardment technique. The ion intensity ratio between two ubiquitous hydrocarbon ions, C6 H- and C4 H- , detected in TOF-SIMS, denoted as ρ, was used to assess the degree of cross-linking of the PMMA films. The cross-linking depth of the PMMA films was revealed by depth profiling ρ into the polymer films using a C60+ sputter beam. RESULTS: The control PMMA film spin-coated on a Si substrate was characterized by ρ = 32% on its surface when using a 25 keV Bi3+ primary ion beam. This parameter on the PMMA films subjected to HHIC treatment for 10, 100 and 500 s increased to 45%, 56% and 65%, respectively. The depth profiles of ρ obtained using a 10 keV C60+ ion beam resembled an exponential decay, from which the cross-linking depth was estimated to be 3, 15 and 39 nm, respectively, for the three cross-linked PMMA films. CONCLUSIONS: We demonstrated that the ion intensity ratio of C6 H- to C4 H- detected in TOF-SIMS provides a unique and simple means to assess the degree of cross-linking of the surface of PMMA films cross-linked by the surface sensitive hyperthermal hydrogen projectile bombardment technique. With a C60+ sputter beam, we were able to depth profile the PMMA films and determine cross-linking depths of the cross-linked polymer films at nanometer resolutions. Copyright © 2017 John Wiley & Sons, Ltd.
RESUMEN
Although the primary elimination pathway for most tyrosine kinase inhibitors (TKIs) involves CYP3A4-mediated metabolism, the mechanism by which these agents are brought into hepatocytes remains unclear. Here, we optimized and validated a competitive counterflow (CCF) assay to examine TKIs as substrates of the hepatic uptake transporter OATP1B1. The CCF method was based on the stimulated efflux of radiolabeled estradiol-17ß-glucuronide under steady-state conditions in HEK293 cells engineered to overexpress OATP1B1. Out of 62 approved TKIs examined, 13 agents were identified as putative substrates of OATP1B1, and pazopanib was selected as a representative hit for further validation studies. The transport of pazopanib by OATP1B1 was confirmed by decreased activity of its target VEGFR2 in OATP1B1-overexpressing cells, but not cells lacking OATP1B1, consistent with molecular docking analyses indicating an overlapping binding orientation on OATP1B1 with the known substrate, estrone-3-sulfate. In addition, the liver-to-plasma ratio of pazopanib in vivo was decreased in mice with a deficiency of the orthologous transporters, and this was accompanied by diminished pazopanib-induced hepatotoxicity, as determined by changes in the levels of liver transaminases. Our study supports the utility of CCF assays to assess substrate affinity for OATP1B1 within a large set of agents in the class of TKIs, and sheds light on the mechanism by which these agents are taken up into hepatocytes in advance of metabolism.
RESUMEN
Vincristine is a widely used chemotherapeutic drug for the treatment of multiple malignant diseases that causes a dose-limiting peripheral neurotoxicity. There is no clinically effective preventative treatment for vincristine-induced sensory peripheral neurotoxicity (VIPN), and mechanistic details of this side effect remain poorly understood. We hypothesized that VIPN is dependent on transporter-mediated vincristine accumulation in dorsal root ganglion neurons. Using a xenobiotic transporter screen, we identified OATP1B3 as a neuronal transporter regulating the uptake of vincristine. In addition, genetic or pharmacological inhibition of the murine orthologue transporter OATP1B2 protected mice from various hallmarks of VIPN - including mechanical allodynia, thermal hyperalgesia, and changes in digital maximal action potential amplitudes and neuronal morphology - without negatively affecting plasma levels or antitumor effects of vincristine. Finally, we identified α-tocopherol from an untargeted metabolomics analysis as a circulating endogenous biomarker of neuronal OATP1B2 function, and it could serve as a companion diagnostic to guide dose selection of OATP1B-type transport modulators given in combination with vincristine to prevent VIPN. Collectively, our findings shed light on the fundamental basis of VIPN and provide a rationale for the clinical development of transporter inhibitors to prevent this debilitating side effect.
Asunto(s)
Enfermedades del Sistema Nervioso Periférico , Xenobióticos , Ratones , Animales , Vincristina/toxicidad , Enfermedades del Sistema Nervioso Periférico/inducido químicamente , Enfermedades del Sistema Nervioso Periférico/tratamiento farmacológico , Enfermedades del Sistema Nervioso Periférico/prevención & control , Hiperalgesia/inducido químicamente , Ganglios Espinales , Proteínas de Transporte de MembranaRESUMEN
Acute myeloid leukemia (AML) with mutations in the tumor suppressor gene TP53 confers a dismal prognosis with 3-year overall survival of <5%. While inhibition of kinases involved in cell cycle regulation induces synthetic lethality in a variety of TP53 mutant cancers, this strategy has not been evaluated in mutant TP53 AML. Previously, we demonstrated that TP-0903 is a novel multikinase inhibitor with low nM activity against AURKA/B, Chk1/2, and other cell cycle regulators. Here, we evaluated the preclinical activity of TP-0903 in TP53 mutant AML cell lines, including a single-cell clone of MV4-11 containing a TP53 mutation (R248W), Kasumi-1 (R248Q), and HL-60 (TP 53 null). TP-0903 inhibited cell viability (IC50, 12−32 nM) and induced apoptosis at 50 nM. By immunoblot, 50 nM TP-0903 upregulated pChk1/2 and pH2AX, suggesting induction of DNA damage. The combination of TP-0903 and decitabine was additive in vitro, and in vivo significantly prolonged median survival compared to single-agent treatments in mice xenografted with HL-60 (vehicle, 46 days; decitabine, 55 days; TP-0903, 63 days; combination, 75 days) or MV4-11 (R248W) (51 days; 62 days; 81 days; 89 days) (p < 0.001). Together, these results provide scientific premise for the clinical evaluation of TP-0903 in combination with decitabine in TP53 mutant AML.
RESUMEN
Chemotherapy-induced peripheral neuropathy is a debilitating toxicity that adversely affects patient quality and course of treatment. Recent findings have demonstrated that the etiology of peripheral neuropathy is dependent on transporter-mediated accumulation in dorsal root ganglia, and targeting this mechanism can afford neurological protection without compromising therapeutic efficacy.
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Organic cation transporter 1 (OCT1) is a transporter that regulates the hepatic uptake and subsequent elimination of diverse cationic compounds. Although OCT1 has been involved in drug-drug interactions and causes pharmacokinetic variability of many prescription drugs, details of the molecular mechanisms that regulate the activity of OCT1 remain incompletely understood. Based on an unbiased phospho-proteomics screen, we identified OCT1 as a tyrosine-phosphorylated transporter, and functional validation studies using genetic and pharmacological approaches revealed that OCT1 is highly sensitive to small molecules that target the protein kinase YES1, such as dasatinib. In addition, we found that dasatinib can inhibit hepatic OCT1 function in mice as evidenced from its ability to modulate levels of isobutyryl L-carnitine, a hepatic OCT1 biomarker identified from a targeted metabolomics analysis. These findings provide novel insight into the post-translational regulation of OCT1 and suggest that caution is warranted with polypharmacy regimes involving the combined use of OCT1 substrates and kinase inhibitors that target YES1.
RESUMEN
PURPOSE: OATP1B1 (SLCO1B1) is the most abundant and pharmacologically relevant uptake transporter in the liver and a key mediator of xenobiotic clearance. However, the regulatory mechanisms that determine OATP1B1 activity remain uncertain, and as a result, unexpected drug-drug interactions involving OATP1B1 substrates continue to be reported, including several involving tyrosine kinase inhibitors (TKI). EXPERIMENTAL DESIGN: OATP1B1-mediated activity in overexpressing HEK293 cells and hepatocytes was assessed in the presence of FDA-approved TKIs, while rosuvastatin pharmacokinetics in the presence of an OATP1B1 inhibiting TKI were measured in vivo. Tyrosine phosphorylation of OATP1B1 was determined by LC/MS-MS-based proteomics and transport function was measured following exposure to siRNAs targeting 779 different kinases. RESULTS: Twenty-nine of 46 FDA-approved TKIs studied significantly inhibit OATP1B1 function. Inhibition of OATP1B1 by TKIs, such as nilotinib, is predominantly noncompetitive, can increase systemic concentrations of rosuvastatin in vivo, and is associated with reduced phosphorylation of OATP1B1 at tyrosine residue 645. Using genetic screens and functional validation studies, the Src kinase LYN was identified as a potential regulator of OATP1B1 activity that is highly sensitive to inhibition by various TKIs at clinically relevant concentrations. CONCLUSIONS: A novel kinase-dependent posttranslational mechanism of OATP1B1 activation was identified and interference with this process by TKIs can influence the elimination of a broad range of xenobiotic substrates.
Asunto(s)
Células HEK293/metabolismo , Hepatocitos/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/fisiología , Proteínas Tirosina Quinasas/fisiología , Animales , Humanos , Ratones , FosforilaciónRESUMEN
INTRODUCTION: . Membrane transporters are integral to the maintenance of cellular integrity of all tissue and cell types. While transporters play an established role in the systemic pharmacokinetics of therapeutic drugs, tissue specific expression of uptake transporters can serve as an initiating mechanism that governs the accumulation and impact of cytotoxic drugs. AREAS COVERED: . This review provides an overview of organic cation transporters as determinants of chemotherapy-induced toxicities. We also provide insights into the recently updated FDA guidelines for in vitro drug interaction studies, with a particular focus on the class of tyrosine kinase inhibitors as perpetrators of transporter-mediated drug interactions. EXPERT OPINION: . Studies performed over the last few decades have highlighted the important role of basolateral uptake and apical efflux transporters in the pathophysiology of drug-induced organ damage. Increased understanding of the mechanisms that govern the accumulation of cytotoxic drugs has provided insights into the development of novel strategies to prevent debilitating toxicities. Furthermore, we argue that current regulatory guidelines provide inadequate recommendations for in vitro studies to identify substrates or inhibitors of drug transporters. Therefore, the translational and predictive power of FDA-approved drugs as modulators of transport function remains ambiguous and warrants further revision of the current guidelines.
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Antineoplásicos/administración & dosificación , Proteínas de Transporte de Catión Orgánico/metabolismo , Proteínas Transportadoras de Solutos/metabolismo , Animales , Antineoplásicos/efectos adversos , Antineoplásicos/farmacocinética , Interacciones Farmacológicas , Humanos , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/farmacocinética , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
Effective treatment for AML is challenging due to the presence of clonal heterogeneity and the evolution of polyclonal drug resistance. Here, we report that TP-0903 has potent activity against protein kinases related to STAT, AKT, and ERK signaling, as well as cell cycle regulators in biochemical and cellular assays. In vitro and in vivo, TP-0903 was active in multiple models of drug-resistant FLT3 mutant AML, including those involving the F691L gatekeeper mutation and bone marrow microenvironment-mediated factors. Furthermore, TP-0903 demonstrated preclinical activity in AML models with FLT3-ITD and common co-occurring mutations in IDH2 and NRAS genes. We also showed that TP-0903 had ex vivo activity in primary AML cells with recurrent mutations including MLL-PTD, ASXL1, SRSF2, and WT1, which are associated with poor prognosis or promote clinical resistance to AML-directed therapies. Our preclinical studies demonstrate that TP-0903 is a multikinase inhibitor with potent activity against multiple drug-resistant models of AML that will have an immediate clinical impact in a heterogeneous disease like AML.
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Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Pirimidinas/farmacología , Sulfonamidas/farmacología , Animales , Línea Celular Tumoral , Resistencia a Antineoplásicos/efectos de los fármacos , Femenino , Duplicación de Gen/efectos de los fármacos , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Masculino , Ratones , Ratones Desnudos , Mutación/efectos de los fármacos , Inhibidores de Proteínas Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas/metabolismo , Pirimidinas/metabolismo , Sulfonamidas/metabolismo , Microambiente Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Peripheral neurotoxicity is a debilitating condition that afflicts up to 90% of patients with colorectal cancer receiving oxaliplatin-containing therapy. Although emerging evidence has highlighted the importance of various solute carriers to the toxicity of anticancer drugs, the contribution of these proteins to oxaliplatin-induced peripheral neurotoxicity remains controversial. Among candidate transporters investigated in genetically engineered mouse models, we provide evidence for a critical role of the organic cation transporter 2 (OCT2) in satellite glial cells in oxaliplatin-induced neurotoxicity, and demonstrate that targeting OCT2 using genetic and pharmacological approaches ameliorates acute and chronic forms of neurotoxicity. The relevance of this transport system was verified in transporter-deficient rats as a secondary model organism, and translational significance of preventive strategies was demonstrated in preclinical models of colorectal cancer. These studies suggest that pharmacological targeting of OCT2 could be exploited to afford neuroprotection in cancer patients requiring treatment with oxaliplatin.
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Neuroglía/metabolismo , Neuronas/metabolismo , Síndromes de Neurotoxicidad/metabolismo , Transportador 2 de Cátion Orgánico/metabolismo , Oxaliplatino , Animales , Femenino , Masculino , Ratones , Ratones Noqueados , Neuroglía/patología , Neuronas/patología , Síndromes de Neurotoxicidad/genética , Síndromes de Neurotoxicidad/patología , Transportador 2 de Cátion Orgánico/genética , Oxaliplatino/efectos adversos , Oxaliplatino/farmacocinética , Oxaliplatino/farmacología , RatasRESUMEN
Chemotherapy-induced peripheral neuropathy (CIPN) is a common and dose-limiting toxicity, negatively affecting both quality of life and disease outcomes. To date, there is no proven preventative strategy for CIPN. Although multiple randomized trials have evaluated a variety of pharmacologic interventions for the treatment of CIPN, only duloxetine has shown clear efficacy in a phase III study. The National Cancer Institute's Symptom Management and Health-Related Quality of Life Steering Committee has identified CIPN as a priority for translational research in cancer care. Promising advances in preclinical research have identified several novel preventative and therapeutic targets, which have the potential to transform the care of patients with this debilitating neurotoxicity. Here, we provide an overarching view of emerging strategies and therapeutic targets that are currently being evaluated in CIPN.
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Antineoplásicos/efectos adversos , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/tratamiento farmacológico , Clorhidrato de Duloxetina/uso terapéutico , Enfermedades del Sistema Nervioso Periférico/tratamiento farmacológico , Antineoplásicos/uso terapéutico , Ensayos Clínicos Fase III como Asunto , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/patología , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/prevención & control , Humanos , Enfermedades del Sistema Nervioso Periférico/patología , Enfermedades del Sistema Nervioso Periférico/prevención & control , Calidad de Vida , Ensayos Clínicos Controlados Aleatorios como AsuntoRESUMEN
A novel method using UPLC with tandem mass-spectrometric detection (UPLC-MS/MS) with positive electrospray ionization was developed for the detection of the antiarrhythmic drug, dofetilide, in mouse plasma and urine. Protein precipitation was performed on 10 µL of plasma and 2 µL of urine samples using dofetilide-D4 as an internal standard, and separation of the analyte was accomplished on a C18 analytical column with the flow of 0.40 mL/min. Subsequently, the method was successfully applied to determine the pharmacokinetic parameters of dofetilide following oral and intravenous administration. The calibration curve was linear over the selected concentration range (R2 ≥ 0.99), with a lower limit of quantitation of 5 ng/mL. The intra-day and inter-day precisions, and accuracies obtained from a 5-day validation ranged from 3.00 to 7.10%, 3.80-7.20%, and 93.0-106% for plasma, and 3.50-9.00%, 3.70-10.0%, 87.0-106% for urine, while the recovery of dofetilide was 93.7% and 97.4% in plasma and urine, respectively. The observed pharmacokinetic profiles revealed that absorption is the rate-limiting step in dofetilide distribution and elimination. Pharmacokinetic studies illustrate that the absolute bioavailability of dofetilide in the FVB strain mice is 34.5%. The current developed method allows for accurate and precise quantification of dofetilide in micro-volumes of plasma and urine, and was found to be suitable for supporting in vivo pharmacokinetic studies.
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Cromatografía Líquida de Alta Presión/métodos , Fenetilaminas/sangre , Fenetilaminas/orina , Plasma/química , Sulfonamidas/sangre , Sulfonamidas/orina , Espectrometría de Masas en Tándem/métodos , Animales , Disponibilidad Biológica , Líquidos Corporales/química , Calibración , Límite de Detección , Masculino , Ratones , Fenetilaminas/farmacocinética , Sulfonamidas/farmacocinéticaRESUMEN
Carvedilol is reported to prevent cancers in humans and animal models. However, a molecular mechanism has yet to be established, and the extent to which other ß-blockers are chemopreventive remains relatively unknown. A comparative pharmacological approach was utilized with the expectation that a mechanism of action could be devised. JB6 Cl 41-5a (JB6 P+) murine epidermal cells were used to elucidate the chemopreventative properties of ß-blockers, as JB6 P+ cells recapitulate in vivo tumor promotion and chemoprevention. The initial hypothesis was that ß-blockers that are GRK/ß-arrestin biased agonists, like carvedilol, are chemopreventive. Sixteen ß-blockers of different classes, isoproterenol, and HEAT HCl were individually co-administered with epidermal growth factor (EGF) to JB6 P+ cells to examine the chemopreventative properties of each ligand. Cytotoxicity was examined to ensure that the anti-transformation effects of each ligand were not due to cellular growth inhibition. Many of the examined ß-blockers suppressed EGF-induced JB6 P+ cell transformation in a non-cytotoxic and concentration-dependent manner. However, the IC50 values are high for the most potent inhibitors (243, 326, and 431 nM for carvedilol, labetalol, and alprenolol, respectively) and there is no correlation between pharmacological properties and inhibition of transformation. Therefore, the role of α1- and ß2-adrenergic receptors (AR) was examined by standard competition assays and shRNA targeting ß2-ARs, the only ß-AR expressed in JB6 P+ cells. The results reveal that pharmacological inhibition of α1- and ß2-ARs and genetic knockdown of ß2-ARs did not abrogate carvedilol-mediated inhibition of EGF-induced JB6 P+ cell transformation. Furthermore, topical administration of carvedilol protected mice from UV-induced skin damage, while genetic ablation of ß2-ARs increased carvedilol-mediated effects. Therefore, the prevailing hypothesis that the chemopreventive property of carvedilol is mediated through ß-ARs is not supported by this data.
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Antagonistas Adrenérgicos beta/farmacología , Carvedilol/farmacología , Factor de Crecimiento Epidérmico/metabolismo , Receptores Adrenérgicos/metabolismo , Alprenolol/farmacología , Animales , Línea Celular , Proliferación Celular , Transformación Celular Neoplásica/efectos de los fármacos , Concentración 50 Inhibidora , Labetalol/farmacología , Ligandos , Ratones , Ratones Endogámicos C57BL , ARN Interferente Pequeño/metabolismo , Receptores Adrenérgicos beta/metabolismo , Transducción de Señal/efectos de los fármacos , Piel/efectos de los fármacos , Piel/efectos de la radiación , Neoplasias Cutáneas/tratamiento farmacológico , Rayos UltravioletaRESUMEN
The solute carrier superfamily comprises of uptake transporters that can contribute to the absorption and elimination of a broad array of clinically important drugs. Recent studies have suggested that the tissue-specific expression of these transporters may have important consequences for an individual's susceptibility to drug-induced organ damage or to drug-drug interactions. Polymorphic variants have been identified in genes encoded by this family, and some of these have been associated with functional changes in transport function and response to anthracycline-induced toxicity and efficacy. Here, we review recent advances in the role solute carrier transporters play in anthracycline-induced cardiotoxicity, highlight potential implications of genetic variants that may contribute to drug response and discuss novel technologies to study mechanisms of anthracycline transport.
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Antraciclinas/efectos adversos , Transporte Biológico/genética , Cardiotoxicidad/genética , Proteínas de Transporte de Membrana/genética , Interacciones Farmacológicas/genética , Variación Genética/genética , HumanosRESUMEN
Murine pharmacokinetics (PK) represents the absorption, distribution, metabolism, and elimination of drugs from the body, which helps to guide clinical studies, ultimately resulting in more effective drug treatment. The purpose of this protocol is to describe a serial bleeding protocol, obtaining blood samples at six time points from single mouse to yield a complete PK profile. This protocol has proved to be rapid, highly repeatable, and relatively easy to acquire. Comparing with the conventional PK studies, this method not only dramatically reduces animal usage, but also decreases sample variation obtained from different animals.