RESUMEN
In a Phase 2 clinical trial, BMS-986020, a lysophosphatidic acid receptor-1 (LPA1) antagonist, produced hepatobiliary toxicity (increased ALT, AST, and ALP; cholecystitis) and increases in plasma bile acids (BA). Nonclinical investigations conducted to identify a potential mechanism(s) for this toxicity examined BMS-986020 and two LPA1 antagonists structurally distinct from BMS-986020 (BMS-986234 and BMS-986278). BMS-986020 inhibited hepatic BA efflux transporters BSEP (IC50 1.8 µM), MRP3 (IC50 22 µM), and MRP4 (IC50 6.2 µM) and inhibited BA canalicular efflux in human hepatocytes (68% at 10 µM). BMS-986020 inhibited mitochondrial function (basal and maximal respiration, ATP production, and spare capacity) in human hepatocytes and cholangiocytes at ≥10 µM and inhibited phospholipid efflux in human hepatocytes (MDR3 IC50 7.5 µM). A quantitative systems toxicology analysis (DILIsym®), considering pharmacokinetics, BA homeostasis, mitochondrial function, oxidative phosphorylation, and reactive intermediates performed for BMS-986020 recapitulated clinical findings ascribing the effects to BA transporter and mitochondrial electron transport chain inhibition. BMS-986234 and BMS-986278 minimally inhibited hepatic BA transporters (IC50 ≥20 µM) and did not inhibit MDR3 activity (IC50 >100 µM), nor did BMS-986234 inhibit BA efflux (≤50 µM) or mitochondrial function (≤30 µM) (BMS-986278 not evaluated). Multiple mechanisms may be involved in the clinical toxicity observed with BMS-986020. The data indicate that this toxicity was unrelated to LPA1 antagonism since the mechanisms that likely influenced the adverse clinical outcome of BMS-986020 were not observed with equipotent LPA1 antagonists BMS-986234 and BMS-986278. This conclusion is consistent with the lack of hepatobiliary toxicity in nonclinical and clinical safety studies with BMS-986278.
Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Enfermedades del Sistema Digestivo/inducido químicamente , Fibrosis Pulmonar Idiopática/tratamiento farmacológico , Hígado/efectos de los fármacos , Receptores del Ácido Lisofosfatídico/antagonistas & inhibidores , Subfamilia B de Transportador de Casetes de Unión a ATP/metabolismo , Ácidos y Sales Biliares/metabolismo , Transporte Biológico/fisiología , Línea Celular , Línea Celular Tumoral , Transporte de Electrón/fisiología , Células HEK293 , Células Hep G2 , Hepatocitos/efectos de los fármacos , Humanos , Mitocondrias/efectos de los fármacosRESUMEN
Two new cassaine-type diterpenoids, namely erythrofordins D (1) and E (2), sourced from a Cameroon collection of Erythrophleum suaveolens were isolated and assessed for anti-tumor activity. In the NCI-60 cancer cell assay, erythrofordins D (1) and E (2) were found to be cytotoxic in the low micro molar ranges with a mean GI50 value of 2.45 and 0.71⯵M, mean TGI value of 9.77 and 2.29⯵M, and a mean LC50 of 26.92 and 11.48⯵M for 1 and 2 respectively. Using the COMPARE algorithm, the new compounds were found to have similar NCI-60 response profiles to the known cardiac glycosides hyrcanoside and strophanthin. In addition, in an assay examining the viability and contractile function in human cardiomyocytes derived from induced pluripotent stem-cells, erythrofordins showed cardiotoxicity effects at concentrations as low as 0.03⯵g/mL.
Asunto(s)
Caesalpinia/química , Diterpenos/farmacología , Miocitos Cardíacos/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Diterpenos/química , Diterpenos/aislamiento & purificación , Relación Dosis-Respuesta a Droga , Humanos , Estructura Molecular , Relación Estructura-ActividadRESUMEN
Targeting the anti-apoptotic protein Mcl-1 holds a promise to improve therapy of multiple types of Mcl-1 dependent cancers but raises concerns of on-target cardiotoxicity due to the presence and reported role of Mcl-1 in heart. Herein, we investigated the importance of Mcl-1 in the survival and contractile function of human pluripotent stem cell-derived cardiomyocytes in culture. Effective knockdown of Mcl-1 with siRNAs reproducibly resulted in early (measured at Day 3) marginal alterations in caspase 3/7 activity, LDH leakage, ATP content and cellular impedance. After 14â¯days of Mcl-1 knockdown, loss of mitochondrial membrane potential, deteriorating effects on mitochondrial ultrastructure, and alterations in beat rate and amplitude were revealed. Inhibition of Bcl-xL by siRNA-mediated knockdown or selective inhibitors did not cause any overt cellular responses except for a minimal increase in caspase 3/7 activity; however, loss of Mcl-1 concomitant with down-regulated Bcl-xL activated apoptosis and caused extensive cell death as reflected by an 80% loss in cell index, activation of caspase-3 with associated PARP cleavage, and a decrease in beat amplitude and mitochondrial membrane potential after 3â¯days of Mcl-1/Bcl-xL knockdown., Together, these findings suggest that Mcl-1 and Bcl-xL provide duplicate safeguard measures in maintaining structural and functional integrity of cardiomyocytes. Hence, BH3-mimetic drugs targeting Mcl-1 may be well tolerated in the presence of intact Bcl-xL.
Asunto(s)
Muerte Celular/fisiología , Células Madre Pluripotentes Inducidas/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Miocitos Cardíacos/metabolismo , Adenosina Trifosfato/metabolismo , Apoptosis/fisiología , Proteínas Reguladoras de la Apoptosis/metabolismo , Caspasa 3/metabolismo , Caspasa 7/metabolismo , Humanos , Potencial de la Membrana Mitocondrial/fisiología , Mitocondrias/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Proteína bcl-X/metabolismoRESUMEN
Batracylin (NSC-320846) is a dual inhibitor of DNA topoisomerases I and II. Batracylin advanced as an anticancer agent to Phase I clinical trials where dose limiting hemorrhagic cystitis (bladder inflammation and bleeding) was observed. To further investigate batracylin's mechanism of toxicity, studies were conducted in Fischer 344 rats. Once daily oral administration of 16 or 32 mg/kg batracylin to rats for 4 days caused overt toxicity. Abnormal clinical observations and adverse effects on clinical pathology, urinalysis, and histology indicated acute renal damage and urothelial damage and bone marrow dysfunction. Scanning electron microscopy revealed sloughing of the superficial and intermediate urothelial layers. DNA damage was evident in kidney and bone marrow as indicated by histone γ-H2AX immunofluorescence. After a single oral administration of 16 or 32 mg/kg, the majority of batracylin was converted to N-acetylbatracylin (NAB) with a half-life of 4 hr to 11 hr. Mesna (Mesnex™), a drug known to reduce the incidence of hemorrhagic cystitis induced by ifosfamide or cyclophosphamide, was administered to rats prior to batracylin, but did not alleviate batracylin-induced bladder and renal toxicity. These findings suggest that batracylin results in DNA damage-based mechanisms of toxicity and not an acrolein-based mechanism of toxicity as occurs after ifosfamide or cyclophosphamide administration.
Asunto(s)
Neoplasias Renales/inducido químicamente , Quinazolinas/toxicidad , Neoplasias de la Vejiga Urinaria/metabolismo , Animales , Biomarcadores de Tumor/análisis , Peso Corporal/efectos de los fármacos , Femenino , Glucosuria/inducido químicamente , Histonas/metabolismo , Neoplasias Renales/metabolismo , Neoplasias Renales/patología , Masculino , Mesna/farmacología , Fosfoproteínas/metabolismo , Quinazolinas/farmacocinética , Distribución Aleatoria , Ratas , Neoplasias de la Vejiga Urinaria/inducido químicamente , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
NSC-743380 (1-[(3-chlorophenyl)-methyl]-1H-indole-3-carbinol) is in early stages of development as an anticancer agent. Two metabolites reflect sequential conversion of the carbinol functionality to a carboxaldehyde and the major metabolite, 1-[(3-chlorophenyl)-methyl]-1H-indole-3-carboxylic acid. In an exploratory toxicity study in rats, NSC-743380 induced elevations in liver-associated serum enzymes and biliary hyperplasia. Biliary hyperplasia was observed 2 days after dosing orally for 2 consecutive days at 100mg/kg/day. Notably, hepatotoxicity and biliary hyperplasia were observed after oral administration of the parent compound, but not when major metabolites were administered. The toxicities of a structurally similar but pharmacologically inactive molecule and a structurally diverse molecule with a similar efficacy profile in killing cancer cells in vitro were compared to NSC-743380 to explore scaffold versus target-mediated toxicity. Following two oral doses of 100mg/kg/day given once daily on two consecutive days, the structurally unrelated active compound produced hepatic toxicity similar to NSC-743380. The structurally similar inactive compound did not, but, lower exposures were achieved. The weight of evidence implies that the hepatotoxicity associated with NSC-743380 is related to the anticancer activity of the parent molecule. Furthermore, because biliary hyperplasia represents an unmanageable and non-monitorable adverse effect in clinical settings, this model may provide an opportunity for investigators to use a short-duration study design to explore biomarkers of biliary hyperplasia.
Asunto(s)
Enfermedad Aguda , Enfermedades de las Vías Biliares/inducido químicamente , Sistema Biliar/efectos de los fármacos , Indoles/efectos adversos , Administración Oral , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/efectos adversos , Antineoplásicos/metabolismo , Antineoplásicos/farmacocinética , Sistema Biliar/metabolismo , Sistema Biliar/patología , Enfermedades de las Vías Biliares/sangre , Enfermedades de las Vías Biliares/metabolismo , Enfermedades de las Vías Biliares/patología , Biomarcadores/sangre , Biotransformación , Enfermedad Hepática Inducida por Sustancias y Drogas/sangre , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Enfermedad Hepática Inducida por Sustancias y Drogas/fisiopatología , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos/métodos , Drogas en Investigación/administración & dosificación , Drogas en Investigación/efectos adversos , Drogas en Investigación/metabolismo , Drogas en Investigación/farmacocinética , Hiperplasia , Indoles/administración & dosificación , Indoles/sangre , Indoles/metabolismo , Indoles/farmacocinética , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Hígado/fisiopatología , Masculino , Distribución Aleatoria , Ratas Endogámicas F344 , Relación Estructura-ActividadRESUMEN
Endoxifen (4-hydroxy-N-desmethyl-tamoxifen), one of the major active metabolites of tamoxifen, has substantially greater estrogen antagonist properties and antiproliferative effects in breast tumor cells than tamoxifen, a mixed estrogen agonist/antagonist. An associated risk of endometrial cancer and hyperplasia has been linked to the estrogen agonist properties of tamoxifen. We evaluated endoxifen using a classic uterotrophic effects method. Rats were given endoxifen or tamoxifen orally for 3 days. Estradiol was the positive control. Endoxifen and tamoxifen plasma levels exceeded those previously observed clinically. Uterine weight was 3-fold higher in the estradiol group than in the tamoxifen or endoxifen groups, which did not differ from vehicle controls. Tamoxifen and endoxifen caused a greater increase in luminal epithelial cell height than estradiol. Both tamoxifen and endoxifen produced an increase in the stromal BrdU labeling index (LI) that was ≤ estradiol and inversely related to dose, but did not affect luminal epithelial cell BrdU LI. As expected, estradiol increased luminal epithelial cell proliferation. These results indicate that endoxifen induces uterotrophic effects, but is less potent than estradiol in eliciting these effects. Given prior preclinical observations that endoxifen has superior antitumor activity than tamoxifen, the observations of similar uterine effects suggest that the endoxifen risk/benefit ratio may be superior to tamoxifen.
Asunto(s)
Ovariectomía , Tamoxifeno/análogos & derivados , Tamoxifeno/toxicidad , Útero/efectos de los fármacos , Animales , Proliferación Celular/efectos de los fármacos , Endometrio/química , Endometrio/efectos de los fármacos , Femenino , Histocitoquímica , Hipertrofia/inducido químicamente , Tamaño de los Órganos/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Útero/químicaRESUMEN
Chronic heart failure is one of the most common reasons for hospitalization. Current risk stratification is based on ejection fraction, whereas many arrhythmic events occur in patients with relatively preserved ejection fraction. We aim to investigate the mechanistic link between proarrhythmic abnormalities, reduced contractility and diastolic dysfunction in heart failure, using electromechanical modelling and simulations of human failing cardiomyocytes. We constructed, calibrated and validated populations of human electromechanical models of failing cardiomyocytes, that were able to reproduce the prolonged action potential, reduced contractility and diastolic dysfunction as observed in human data, as well as increased propensity to proarrhythmic incidents such as early afterdepolarization and beat-to-beat alternans. Our simulation data reveal that proarrhythmic incidents tend to occur in failing myocytes with lower diastolic tension, rather than with lower contractility, due to the relative preserved SERCA and sodium calcium exchanger current. These results support the inclusion of end-diastolic volume to be potentially beneficial in the risk stratifications of heart failure patients.
Asunto(s)
Potenciales de Acción , Arritmias Cardíacas , Insuficiencia Cardíaca , Miocitos Cardíacos , Humanos , Miocitos Cardíacos/metabolismo , Insuficiencia Cardíaca/fisiopatología , Arritmias Cardíacas/fisiopatología , Arritmias Cardíacas/etiología , Diástole/fisiología , Modelos Cardiovasculares , Contracción Miocárdica/fisiología , Simulación por Computador , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico/metabolismoRESUMEN
Seizure liability remains a significant cause of attrition throughout drug development. Advances in stem cell biology coupled with an increased understanding of the role of ion channels in seizure offer an opportunity for a new paradigm in screening. We assessed the activity of 15 pro-seizurogenic compounds (7 CNS active therapies, 4 GABA receptor antagonists, and 4 other reported seizurogenic compounds) using automated electrophysiology against a panel of 14 ion channels (Nav1.1, Nav1.2, Nav1.6, Kv7.2/7.3, Kv7.3/7.5, Kv1.1, Kv4.2, KCa4.1, Kv2.1, Kv3.1, KCa1.1, GABA α1ß2γ2, nicotinic α4ß2, NMDA 1/2A). These were selected based on linkage to seizure in genetic/pharmacological studies. Fourteen compounds demonstrated at least one "hit" against the seizure panel and 11 compounds inhibited 2 or more ion channels. Next, we assessed the impact of the 15 compounds on electrical signaling using human-induced pluripotent stem cell neurons in microelectrode array (MEA). The CNS active therapies (amoxapine, bupropion, chlorpromazine, clozapine, diphenhydramine, paroxetine, quetiapine) all caused characteristic changes to electrical activity in key parameters indicative of seizure such as network burst frequency and duration. The GABA antagonist picrotoxin increased all parameters, but the antibiotics amoxicillin and enoxacin only showed minimal changes. Acetaminophen, included as a negative control, caused no changes in any of the parameters assessed. Overall, pro-seizurogenic compounds showed a distinct fingerprint in the ion channel/MEA panel. These studies highlight the potential utility of an integrated in vitro approach for early seizure prediction to provide mechanistic information and to support optimal drug design in early development, saving time and resources.
Asunto(s)
Células Madre Pluripotentes Inducidas , Humanos , Células Madre Pluripotentes Inducidas/fisiología , Neuronas/fisiología , Convulsiones/inducido químicamente , Microelectrodos , Canales IónicosRESUMEN
Much has been written and said about the promise and excitement of microphysiological systems, miniature devices that aim to recreate aspects of human physiology on a chip. The rapid explosion of the offerings and persistent publicity placed high expectations on both product manufacturers and regulatory agencies to adopt the data. Inevitably, discussions of where this technology fits in chemical testing paradigms are ongoing. Some end-users became early adopters, whereas others have taken a more cautious approach because of the high cost and uncertainties of their utility. Here, we detail the experience of a public-private collaboration established for testing of diverse microphysiological systems. Collectively, we present a number of considerations on practical aspects of using microphysiological systems in the context of their applications in decision-making. Specifically, future end-users need to be prepared for extensive on-site optimization and have access to a wide range of imaging and other equipment. We reason that cells, related reagents, and the technical skills of the research staff, not the devices themselves, are the most critical determinants of success. Extrapolation from concentration-response effects in microphysiological systems to human blood or oral exposures, difficulties with replicating the whole organ, and long-term functionality remain as critical challenges. Overall, we conclude that it is unlikely that a rodent- or human-equivalent model is achievable through a finite number of microphysiological systems in the near future; therefore, building consensus and promoting the gradual incorporation of these models into tiered approaches for safety assessment and decision-making is the sensible path to wide adoption.
Asunto(s)
Dispositivos Laboratorio en un Chip , HumanosRESUMEN
The integrative responses of the cardiovascular (CV) system are essential for maintaining blood flow to provide oxygenation, nutrients, and waste removal for the entire body. Progress has been made in independently developing simple in vitro models of two primary components of the CV system, namely the heart (using induced pluripotent stem-cell derived cardiomyocytes) and the vasculature (using endothelial cells and smooth muscle cells). These two in vitro biomimics are often described as immature and simplistic, and typically lack the structural complexity of native tissues. Despite these limitations, they have proven useful for specific "fit for purpose" applications, including early safety screening. More complex in vitro models offer the tantalizing prospect of greater refinement in risk assessments. To this end, efforts to physically link cardiac and vascular components to mimic a true CV microphysiological system (CVMPS) are ongoing, with the goal of providing a more holistic and integrated CV response model. The challenges of building and implementing CVMPS in future pharmacological safety studies are many, and include a) the need for more complex (and hence mature) cell types and tissues, b) the need for more realistic vasculature (within and across co-modeled tissues), and c) the need to meaningfully couple these two components to allow for integrated CV responses. Initial success will likely come with simple, bioengineered tissue models coupled with fluidics intended to mirror a vascular component. While the development of more complex integrated CVMPS models that are capable of differentiating safe compounds and providing mechanistic evaluations of CV liabilities may be feasible, adoption by pharma will ultimately hinge on model efficiency, experimental reproducibility, and added value above current strategies.
Asunto(s)
Células Endoteliales , Células Madre Pluripotentes Inducidas , Modelos Cardiovasculares , Miocitos Cardíacos , Reproducibilidad de los ResultadosRESUMEN
Many in vitro gastrointestinal models have been developed with the hope that they will continue to improve in their similarity to the organs from which they were isolated. Intestinal organoids isolated from various species are now being used to investigate physiology and pathophysiology. In this study, intestinal stem cells were isolated from adult rat duodenum and culture conditions were optimized to promote the growth, differentiation and development of 3D organoids. We optimized and characterized rat duodenal organoids with light and electron microscopy, immunofluorescence and notably, global mRNA expression. The metabolic capacity of these cultures was investigated using probe substrates for multiple phase I and phase II drug metabolizing enzymes and found to be in line with previous results from intestinal primary cultures and a significant improvement over immortalized cell lines. Over the course of differentiation, the gene expression profiles of the rat duodenal organoids were consistent with expected trends in differentiation to various cell lineages reflecting the duodenum in vivo. Further, incubations of these cultures with naproxen and celecoxib resulted in cytotoxicity consistent with the direct cytotoxic effects of these drugs to duodenum in vivo. Based on these characteristics, the rat duodenal organoids described herein will provide a novel platform for investigating a wide variety of mechanistic questions.
Asunto(s)
Antiinflamatorios no Esteroideos/toxicidad , Diferenciación Celular/efectos de los fármacos , Duodeno/efectos de los fármacos , Mucosa Intestinal/efectos de los fármacos , Organoides/efectos de los fármacos , Animales , Antiinflamatorios no Esteroideos/metabolismo , Diferenciación Celular/fisiología , Células Cultivadas , Duodeno/citología , Duodeno/metabolismo , Femenino , Mucosa Intestinal/citología , Mucosa Intestinal/metabolismo , Organoides/metabolismo , Ratas , Ratas Sprague-Dawley , Células Madre/efectos de los fármacos , Células Madre/metabolismoRESUMEN
The human kidney contains approximately one million nephrons. As the functional unit of the kidney, the nephron affords an opportunity to approximate the kidney at a microphysiological scale. Recent emergence of physiologically accurate human tissue models has radically advanced the possibilities of mimicking organ biology and multi-organ combinations in vitro. Anatomically, the nephron is one of the most complex, sequentially integrated microfluidic units in the body making the miniaturized microfluidic systems excellent candidates for capturing the kidney biology in vitro. While these models are promising, there are a number of considerations for practical implementation into a drug development paradigm. Opportunities for pharmaceutical industry applications of new MPS models often start with drug safety testing. As such, the intent of this article is to focus on safety and ADME applications. This article reviews biological functions of the kidney and options for characterizing known roles in nephrotoxicity. The concept of "context-of-use" is introduced as a framework for describing and verifying the specific features of an MPS platform for use in drug development. Overall, we present a perspective on key attributes of microphysiological kidney models, which the pharmaceutical industry could leverage to improve confident safety and ADME evaluations of experimental therapies.
Asunto(s)
Riñón/efectos de los fármacos , Preparaciones Farmacéuticas/metabolismo , Desarrollo de Medicamentos , Evaluación Preclínica de Medicamentos/efectos adversos , Industria Farmacéutica , Humanos , Dispositivos Laboratorio en un Chip , Técnicas Analíticas Microfluídicas/instrumentación , Modelos Biológicos , Preparaciones Farmacéuticas/químicaRESUMEN
Cardiac troponin I is a useful biomarker of myocardial injury, but its use in mice and application to early drug discovery are not well described. The authors investigated the relationship between cTnI concentration in serum and histologic lesions in heart tissue from mice treated with isoproterenol (ISO). Cardiac TnI concentrations in serum increased in a dose-dependant manner and remained increased twenty-four to forty-eight hours after a single administration of isoproterenol. Increased cTnI concentration was of greater magnitude and longer duration than increased fatty acid binding protein 3 concentration, aspartate aminotransferase activity, and creatine kinase activity in serum. Isoproterenol-induced increases in cTnI concentrations were both greater and more sustained in BALB/c than in CD1 mice and correlated with incidence and severity of lesions observed in heart sections from both strains. In drug development studies in BALB/c mice with novel kinase inhibitors, cTnI concentration was a reliable stand-alone biomarker of cardiac injury and was used in combination with measurements of in vivo target inhibition to demonstrate an off-target contribution to cardiotoxicity. Additional attributes, including low cost and rapid turnaround time, made cTnI concentration in serum invaluable for detecting cardiotoxicity, exploring structure-activity relationships, and prioritizing development of compounds with improved safety profiles early in drug discovery.
Asunto(s)
Descubrimiento de Drogas/métodos , Cardiopatías/sangre , Cardiopatías/inducido químicamente , Isoproterenol/toxicidad , Inhibidores de Proteínas Quinasas/toxicidad , Troponina I/sangre , Animales , Aspartato Aminotransferasas/sangre , Biomarcadores/sangre , Cardiotónicos/toxicidad , Creatina Quinasa/antagonistas & inhibidores , Creatina Quinasa/sangre , Relación Dosis-Respuesta a Droga , Proteína 3 de Unión a Ácidos Grasos , Proteínas de Unión a Ácidos Grasos/sangre , Femenino , Ventrículos Cardíacos/efectos de los fármacos , Histocitoquímica , Inflamación/metabolismo , Ratones , Ratones Endogámicos BALB C , Miocardio/metabolismo , Miocardio/patología , NecrosisRESUMEN
To achieve therapeutic goals, many cancer chemotherapeutics are used at doses close to their maximally tolerated doses. Thus, it may be expected that when therapies are combined at therapeutic doses, toxicity profiles may change. In many ways, prediction of synergistic toxicities for drug combinations is similar to predicting synergistic efficacy, and is dependent upon building hypotheses from molecular mechanisms of drug toxicity. The key objective of this initiative was to generate and make publicly available key high-content data sets for mechanistic hypothesis generation as it pertains to a unique toxicity profile of a drug pair for several anticancer drug combinations. The expectation is that tissue-based genomic information that are derived from target tissues will also facilitate the generation and testing of mechanistic hypotheses. The view is that availability of these data sets for bioinformaticians and other scientists will contribute to analysis of these data and evaluation of the approach.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Perfilación de la Expresión Génica , Animales , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/genética , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/metabolismo , Especificidad de Órganos , Ratas , Análisis de Matrices TisularesRESUMEN
The lung is a complex organ; it is both the initial barrier for inhaled agents and the site of metabolism and therapeutic effect for a subset of systemically administered drugs. Comprised of more than 40 cell types that are responsible for various important functions, the lung's complexity contributes to the subsequent challenges in developing complex in vitro co-culture models (also called microphysiological systems (MPS), complex in vitro models or organs-on-a-chip). Although there are multiple considerations and limitations in the development and qualification of such in vitro systems, MPS exhibit great promise in the fields of pharmacology and toxicology. Successful development and implementation of MPS models may enable mechanistic bridging between non-clinical species and humans, and increase clinical relevance of safety endpoints, while decreasing overall animal use. This article summarizes, from a biopharmaceutical industry perspective, essential elements for the development and qualification of lung MPS models. Its purpose is to guide MPS developers and manufacturers to expedite MPS utilization for safety assessment in the biopharmaceutical industry.
Asunto(s)
Técnicas de Cocultivo , Dispositivos Laboratorio en un Chip , Pulmón/metabolismo , Técnicas Analíticas Microfluídicas , Modelos Biológicos , Técnicas de Cocultivo/instrumentación , Humanos , Pulmón/efectos de los fármacos , Pulmón/patología , Técnicas Analíticas Microfluídicas/instrumentaciónRESUMEN
Cardiovascular (CV) toxicity from cancer therapy is a significant and growing concern. Conventional oncology clinical trial designs focused singularly on cancer treatment efficacy have not provided sufficient information on both CV risk factors and outcomes. Similarly, traditional CV trials evaluating standard interventions typically exclude cancer patients, particularly those actively receiving cancer therapy. Neither trial type simultaneously evaluates the balance between CV toxicity and cancer outcomes. However, there is increasing collaboration among oncologists and cardiologists to design new cardio-oncology trials that address this important need. In this review, we detail five ongoing, oncology-based trials with integrated CV endpoints. Key design features include: 1) a careful assessment of baseline risk factors for CV disease; 2) an introduction of cardioprotective interventions at various timepoints in cancer therapy; 3) a balance of the risk of subclinical CV injury with the need for ongoing cancer treatment; and 4) an understanding of the time profile for development of clinically apparent CV toxicity. Additional critical priorities in cardio-oncology clinical research include harmonization of data collection and definitions for all physician- and patient-reported exposures and outcomes.
RESUMEN
Homozygous p53 deficient knockout mice were used to assess the role of p53 in tumor promotion by the protein phosphatase inhibitor and hepatic tumor promoter microcystin-LR (MCLR). More than 50% of human cancers bear mutations in the p53 gene, and in particular, p53 tumor suppressor gene mutations have been shown to play a major role in hepatocarcinogenesis. Trp53 homozygous (inactivated p53) and age-matched wild-type control mice were assigned to vehicle or MCLR-treated groups. MCLR or saline was administered daily for up to 28 days. RNA from the 28-day study was hybridized onto Mouse Genome GeneChip arrays. Selected RNA from 28 days and earlier time points was also processed for quantitative polymerase chain reaction (PCR). Livers from the 28-day, Trp53-deficient, MCLR group displayed greater hyperplastic and dysplastic changes morphologically and increases in Ki-67 and phosphohistone H3 (mitotic marker) immunoreactivity. Gene-expression analysis revealed significant increases in expression of cell-cycle regulation and cellular proliferation genes in the MCLR-treated, p53-deficient mutant mice compared to controls. These data suggest that regulation of the cell cycle by p53 is important in preventing the proliferative response associated with chronic, sublethal microcystin exposure, and therefore, conclude that p53 plays an important role in MCLR-induced tumor promotion.
Asunto(s)
Toxinas Bacterianas/toxicidad , Ciclo Celular/genética , Expresión Génica/efectos de los fármacos , Genes p53/genética , Hepatocitos/efectos de los fármacos , Microcistinas/toxicidad , Mitosis/genética , Animales , Carcinógenos/toxicidad , Proliferación Celular/efectos de los fármacos , Perfilación de la Expresión Génica , Silenciador del Gen , Hepatocitos/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Toxinas Marinas , Ratones , Ratones Noqueados , Índice Mitótico , Fosfoproteínas Fosfatasas/antagonistas & inhibidores , ARN Mensajero/metabolismo , Proteína p53 Supresora de Tumor/deficiencia , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Approved and marketed drugs are frequently studied in nonclinical models to evaluate the potential application to additional disease indications or to gain insight about molecular mechanisms of action. A survey of the literature reveals that nonclinical experimental designs (in vitro or in vivo) often include evaluation of drug concentrations or doses that are much higher than what can be achieved in patients (i.e., above the maximally tolerated dose or much higher than the clinically relevant exposures). The results obtained with these high concentrations may be particularly helpful in elucidating off-target effects and toxicities, but it is critical to have a dose-response curve that includes the minimally effective or clinically effective concentration for comparison. We have reviewed the clinical literature and drug product labels for all small molecules and biological agents approved by the FDA for use in oncology to identify and compile the available pharmacokinetic parameters. The data summarized here can serve as a guide for selection of in vitro concentrations and in vivo plasma exposures for evaluation of drug effects in nonclinical studies. Inclusion of drug concentrations or exposures that are relevant to those observed in clinical practice can improve translation of nonclinical mechanism of action findings into potentially relevant clinical effects. Clin Cancer Res; 23(14); 3489-98. ©2017 AACR.
Asunto(s)
Antineoplásicos/uso terapéutico , Relación Dosis-Respuesta a Droga , Oncología Médica/tendencias , Neoplasias/tratamiento farmacológico , Antineoplásicos/efectos adversos , Humanos , Estados Unidos , United States Food and Drug AdministrationRESUMEN
Chemotherapy-induced peripheral neuropathy (CIPN) is a major, dose-limiting adverse effect experienced by cancer patients. Advancements in mechanism-based risk mitigation and effective treatments for CIPN can be aided by suitable in vitro assays. To this end, we developed a multiparametric morphology-centered rat dorsal root ganglion (DRG) assay. Morphologic alterations in subcellular structures of neurons and non-neurons were analyzed with an automated microscopy system. Stains for NeuN (a neuron-specific nuclear protein) and Tuj-1 (ß-III tubulin) were used to identify neuronal cell nuclei and neuronal cell bodies/neurites, respectively. Vimentin staining (a component of Schwann cell intermediate filaments) was used to label non-neuronal supporting cells. Nuclei that stained with DAPI, but lacked NeuN represented non-neuronal cells. Images were analyzed following 24 h of continuous exposure to CIPN-inducing agents and 72 h after drug removal to provide a dynamic measure of recovery from initial drug effects. Treatment with bortezomib, cisplatin, eribulin, paclitaxel or vincristine induced a dose-dependent loss of neurite/process areas, mimicking the 'dying back' degeneration of axons, a histopathological hallmark of clinical CIPN in vivo. The IC50 for neurite loss was within 3-fold of the maximal clinical exposure (Cmax) for all five CIPN-inducing drugs, but was >4- or ≥ 28-fold of the Cmax for 2 non-CIPN-inducing agents. Compound-specific effects, eg, neurite fragmentation by cisplatin or bortezomib and enlarged neuronal cell bodies by paclitaxel, were also observed. Collectively, these results support the use of a quantitative, morphologic evaluation and a DRG cell culture model to inform risk and examine mechanisms of CIPN.
Asunto(s)
Antineoplásicos/efectos adversos , Ganglios Espinales/efectos de los fármacos , Neuronas/efectos de los fármacos , Animales , Biomarcadores/metabolismo , Cuerpo Celular/efectos de los fármacos , Cuerpo Celular/metabolismo , Cuerpo Celular/patología , Forma del Núcleo Celular/efectos de los fármacos , Forma de la Célula/efectos de los fármacos , Células Cultivadas , Evaluación Preclínica de Medicamentos/métodos , Electroforesis Capilar , Técnica del Anticuerpo Fluorescente , Ganglios Espinales/metabolismo , Ganglios Espinales/patología , Procesamiento de Imagen Asistido por Computador , Cinética , Peso Molecular , Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/metabolismo , Neuritas/efectos de los fármacos , Neuritas/metabolismo , Neuritas/patología , Neuronas/metabolismo , Neuronas/patología , Síndromes de Neurotoxicidad/etiología , Síndromes de Neurotoxicidad/metabolismo , Síndromes de Neurotoxicidad/patología , Forma de los Orgánulos/efectos de los fármacos , Tamaño de los Orgánulos/efectos de los fármacos , Enfermedades del Sistema Nervioso Periférico/etiología , Enfermedades del Sistema Nervioso Periférico/metabolismo , Enfermedades del Sistema Nervioso Periférico/patología , RatasRESUMEN
The vascular endothelial growth factor receptor (VEGFR) is an important angiogenic target for cancer gene therapy. In this study, we designed an mRNA-cleaving oligodeoxynucleotide that targets the VEGF receptor 2 (VEGFR2) transcript (VEGFR2 DNAzyme). This DNAzyme was found to digest efficiently mRNA substrates of VEGFR2 in a concentration- and time-dependent manner. We also showed that the DNAzyme induces apoptosis and markedly inhibits endothelial cell growth compared with a disabled DNAzyme and untreated controls. In contrast, the DNAzyme did not inhibit the growth of MDA-MB-435 cells in vitro. The DNAzyme in complex with a nonviral carrier also significantly inhibited tumor growth in vivo. After the fourth injection, there was nearly a 75% reduction of tumor size in the DNAzyme-treated group compared with the saline-injected control group (P = 0.024). Marked cell death in the peripheral regions of the tumor accompanied by a reduction in blood vessel density is consistent with the antiangiogenic mechanism of the DNAzyme. This study indicates that DNAzymes, targeting angiogenic growth factors of tumors, show promise as antitumor agents.