In vitro ADME phenotyping in drug discovery: current challenges and future solutions

Drug-metabolizing enzymes and drug transporters are key regulators of drug disposition and pharmacodynamics, which are closely linked to drug efficacy and safety. In this article, current challenges and future solutions to predicting their influence on pharmacokinetics and inter-organ distribution in humans, from data generated during the drug discovery decision-making process, are presented. In vitro phenotyping strategies for drug metabolizing enzymes (eg, CYP3A4, UGT1A1) and transporters (eg, OATP1B1) are offered, including perspectives on a selection of in vitro systems, novel in vitro phenotyping reagents and remaining technology gaps, challenges in extrapolating in vitro data to the in vivo situation, in silico models for the prediction of whether compounds are enzyme or transporter substrates, and the impact of pharmacogenomics.     

Preclinical models for pediatric solid tumor drug discovery: current trends,challenges and the scopes for improvement

Introduction: The enhancement in pediatric cancer survival achieved in the past few decades has been confined to low- and moderate-risk cancers, whereas no notable improvement in survival was observed in high-risk and advanced-stage childhood cancers. High attrition rate of candidate drugs in clinical trials is a major hurdle in the development of effective therapies for pediatric solid tumors. In order to reduce the failure rate of candidate drugs in clinical trials, more effective strategies are needed to enhance the predictability of preclinical testing.

Areas covered: The authors have described the current trends in preclinical drug development for treating pediatric solid tumors. Furthermore, the authors review their limitations and the available remedies, with regards to choice of models, pharmacokinetic considerations and the criteria for assessing the long-term efficacy of a candidate drug.

Expert opinion: In many solid tumors, common differences between pediatric and adult cancers have been observed, and therefore, clinical trials for pediatric solid tumors must be conducted on the basis of preclinical observations in pediatric solid tumor models. There is a need to invest in extensive preclinical testing on pediatric solid tumor models. None of the preclinical models can fully recapitulate the human cancers. Therefore, these limitations must be considered while conducting a preclinical trial. The dose and schedule of drugs used for preclinical testing must be clinically relevant. While testing the efficacy of drugs, the markers of apoptosis, drug resistance, hypoxia and tumor-initiating cells can inform us about the long-term therapeutic response of a cancer.     

Predictive models for nanotoxicology: current challenges and future opportunities

Characterizing the risks posed by nanomaterials is extraordinarily complex because these materials can have a wide range of sizes, shapes, chemical compositions and surface modifications, all of which may affect toxicity. There is an urgent need for a testing strategy that can rapidly and efficiently provide a screening approach for evaluating the potential hazard of nanomaterials and inform the prioritization of additional toxicological testing where necessary. Predictive toxicity models could form an integral component of such an approach by predicting which nanomaterials, as a result of their physico-chemical characteristics, have potentially hazardous properties. Strategies for directing research towards predictive models and the ancillary benefits of such research are presented here.     

Peripheral primitive neuroectodermal tumor presenting in the retroperitoneum: a case report with immunohistochemical study

It is now believed that malignant peripheral primitive neuroectodermal tumor (PNET) form a distinct tumor entity from other malignant small round cell tumors exhibiting neuroectodermal differentiation by morphologic, immunohistochemical or electron microscopic analyses. A 17-year-old Ethiopian boy was found to have a big upper extra osseous retroperitoneal tumor mass not associated with peripheral nerve that had infiltrated the body of the pancreas. Histologic sections from excised biopsy showed neoplastic cells with a high nuclear-cytoplasmic ratio and had an indistinct cytoplasm with numerous Homer-Wright rosettes. Immunohistochemically, isolated tumor cells and the centre of rosettes disclosed strong positivity to neural markers, synaptophysin and chromogranin. To the best of our knowledge, this case report represents the first patient described in Ethiopia.     

肾上腺原始神经外胚层肿瘤二例诊治分析并文献复习

目的 探讨国内外肾上腺原始神经外胚层肿瘤(PNET)的诊断和治疗.方法 回顾性分析桐乡市中医医院诊治的2例肾上腺PNET患者的临床资料,并结合相关国内外文献探讨其诊断和治疗方法.结果 B超、CT检查示两例患者肾上腺区边界不清的囊实性软组织肿块,最大直径分别约3 cm和15 cm.2例患者均行手术治疗.术后病理及免疫组化确诊为肾上腺PNET.患者1尚无肿瘤复发证据.患者2随访3个月后发现腹膜后广泛淋巴结转移.6个月后死亡.结论 肾上腺PNET临床罕见,恶性程度高且预后不良,确诊主要依靠病理和免疫组化,治疗提倡多手段综合治疗.     

Individualized risk for statin-induced myopathy: current knowledge, emerging challenges and potential solutions

Skeletal muscle toxicity is the primary adverse effect of statins. In this review, we summarize current knowledge regarding the genetic and nongenetic determinants of risk for statin induced myopathy. Many genetic factors were initially identified through candidate gene association studies limited to pharmacokinetic (PK) targets. Through genome-wide association studies, it has become clear that SLCO1B1 is among the strongest PK predictors of myopathy risk. Genome-wide association studies have also expanded our understanding of pharmacodynamic candidate genes, including RYR2. It is anticipated that deep resequencing efforts will define new loci with rare variants that also contribute, and sophisticated computational approaches will be needed to characterize gene-gene and gene-environment interactions. Beyond environment, race is a critical covariate, and its influence is only partly explained by geographic differences in the frequency of known pharmacodynamic and PK variants. As such, admixture analyses will be essential for a full understanding of statin-induced myopathy.     

In vivo tumor targeting of tumor necrosis factor-alpha-loaded stealth nanoparticles: effect of MePEG molecular weight and particle size.

The aim of this study is to reveal the influence of methoxypolyethyleneglycol (MePEG) molecular weight and particle size of stealth nanoparticles on their in vivo tumor targeting properties. Three sizes (80, 170 and 240nm) of poly methoxypolyethyleneglycol cyanoacrylate-co-n-hexadecyl cyanoacrylate (PEG-PHDCA) nanoparticles loading recombinant human tumor necrosis factor-alpha (rHuTNF-alpha) were prepared at different MePEG molecular weights (MW=2000, 5000 and 10,000) using double emulsion method. The opsonization in mouse serum was evaluated by Coomassie brilliant blue staining of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Phagocytosis was evaluated by incubating (125)I-rHuTNF-alpha-loaded nanoparticles with mouse macrophages (RAW264.7). The pharmacokinetics, biodistribution and tumor targeting studies were performed in S-180 tumor-bearing mice. Higher MePEG molecular weight provided thicker fixed aqueous layer thickness (FALT) and smaller particle size offered higher surface MePEG density. The serum protein adsorption and phagocytic uptake were markedly decreased for the nanoparticles with higher MePEG molecular weight or smaller size. The particles (80nm) made of PEG(5000)-PHDCA, possessing a thicker FALT (5.16nm) and a shortest distance (0.87nm) between two neighboring MePEG chains, showed the strongest capacity of decreasing protein adsorption and phagocytic uptake. These particles extended the half-life of rHuTNF-alpha in S-180 tumor-bearing mice by 24-fold (from 28.2 min to 11.33 h), elevated the rHuTNF-alpha peak concentration in S-180 tumors by 2.85-fold and increased the area under the intratumoral rHuTNF-alpha concentration curve by 7.44-fold. The results of the present study showed PEG-PHDCA nanoparticles with higher MePEG molecular weight and smaller particle size could achieve higher in vivo tumor targeting efficiency.     

In vivo antitumor activity of sparsomycin and its analogues in eight murine tumor models

Summary Sparsomycin (Sm) is a known inhibitor of ribosomal protein synthesis with an attractive anticancer potential. Recently, several analogues of Sm which are more active than the parent drug were selected for further study on the basis of in vitro investigations. Six analogues as well as the parent drug were tested for their antitumor activity in eight in vivo murine tumor models: P388 and L1210 leukemias, RC renal cell carcinoma, B16 melanoma, C38 colon carcinoma, LL Lewis lung carcinoma, C22LR osteosarcoma and M5076 sarcoma. Sm itself appeared to have only borderline activity on L1210 leukemia. The analogues that were most active in vitro showed also the highest in vivo activity. The most sensitive tumors were RC, L1210 and P388. Minimal activity was found on B16 and no activity on C22LR, M5076, C38 and LL. The most active compounds are deshydroxy-Sm, ethyl-deshydroxy-Sm and n-pentyl-Sm. There was a considerable loss of activity when L1210 leukemia was implanted sc while the drugs were administered iv. Only one drug, ethyl-deshydroxy-Sm appeared to be active in this assay. No single most effective compound could be found in this study. The overall activity of Sm and its analogues is moderate. The three analogues which show high activity in three ascitic tumors will be further investigated using human tumor xenograft models.Part of this work was presented in the 5th EORTC/NCI symposium on New Drugs in Cancer Therapy, Amsterdam, 22–24 October 1986.     

Counterfeit drugs: defining the problem and finding solutions

The problem of counterfeit drugs is increasingly becoming a top priority of drug regulatory agencies, licit pharmaceutical manufacturers and healthcare professionals, and is of rising concern among consumers. A review of the current literature reveals that counterfeiting is no longer isolated in developing nations, but is a worldwide pandemic. Although international organisations such as the World Health Organization, have devised specific measures to combat the counterfeit trade, the problem still remains quite daunting. Although the practice of drug counterfeiting is unlikely to ever completely disappear, it may be controlled if governments and all relevant parties combine forces to identify and disrupt the counterfeit chain.     

Counterfeit drugs: defining the problem and finding solutions

The problem of counterfeit drugs is increasingly becoming a top priority of drug regulatory agencies, licit pharmaceutical manufacturers and healthcare professionals, and is of rising concern among consumers. A review of the current literature reveals that counterfeiting is no longer isolated in developing nations, but is a worldwide pandemic. Although international organisations such as the World Health Organization, have devised specific measures to combat the counterfeit trade, the problem still remains quite daunting. Although the practice of drug counterfeiting is unlikely to ever completely disappear, it may be controlled if governments and all relevant parties combine forces to identify and disrupt the counterfeit chain.     

In vitro blood-brain barrier models: current and perspective technologies

Even in the 21st century, studies aimed at characterizing the pathological paradigms associated with the development and progression of central nervous system diseases are primarily performed in laboratory animals. However, limited translational significance, high cost, and labor to develop the appropriate model (e.g., transgenic or inbred strains) have favored parallel in vitro approaches. In vitro models are of particular interest for cerebrovascular studies of the blood-brain barrier (BBB), which plays a critical role in maintaining the brain homeostasis and neuronal functions. Because the BBB dynamically responds to many events associated with rheological and systemic impairments (e.g., hypoperfusion), including the exposure of potentially harmful xenobiotics, the development of more sophisticated artificial systems capable of replicating the vascular properties of the brain microcapillaries are becoming a major focus in basic, translational, and pharmaceutical research. In vitro BBB models are valuable and easy to use supporting tools that can precede and complement animal and human studies. In this article, we provide a detailed review and analysis of currently available in vitro BBB models ranging from static culture systems to the most advanced flow-based and three-dimensional coculture apparatus. We also discuss recent and perspective developments in this ever expanding research field.     

In vivo models of childhood leukemia for preclinical drug testing

The number of new anti-cancer drugs emerging for clinical trials in humans far exceeds the availability of pediatric acute leukemia patients to be entered into clinical trials. Therefore, preclinical testing of new agents for the treatment of childhood acute leukemia is essential to ensure that the most promising drugs are prioritized to enter clinical trials. Historically, the murine system has been central to modeling human leukemia in vivo. A greater knowledge of the molecular lesions underlying particular subtypes of leukemia has led to the generation of genetically engineered murine models, generally involving the knockin or knockout of certain genes and fusion genes at their normal genetic locus. However, the most predominant in vivo models for preclinical drug testing have been human leukemia xenografts. Successful engraftment of all subtypes of acute lymphoblastic leukemia, most subtypes of acute myeloid leukemia as well as juvenile myelomonocytic leukemia, chronic myeloid leukemia and chronic lymphocytic leukemia have been described in various immune-deficient murine hosts. Preclinical testing of novel therapeutics in vivo will likely identify the most promising new agents to enter clinical trials, and will allow their future use to be optimized in combination with other novel and conventional chemotherapeutics.     

The challenges facing block copolymer micelles for cancer therapy: In vivo barriers and clinical translation

The application of block copolymer micelles (BCMs) in oncology has benefitted from advances in polymer chemistry, drug formulation and delivery as well as in vitro and in vivo biological models. While great strides have been made in each of these individual areas, there remains some disappointment overall, citing, in particular, the absence of more BCM formulations in clinical evaluation and practice. In this review, we aim to provide an overview of the challenges presented by in vivo systems to the effective design and development of BCMs. In particular, the barriers posed by systemic administration and tumor properties are examined. The impact of critical features, such as the size, stability and functionalization of BCMs is discussed, while key pre-clinical endpoints and models are critiqued. Given clinical considerations, we present this work as a means to stimulate a renewed focus on the unique chemical versatility bestowed by BCMs and a measured grasp of representative in vitro and in vivo models.     

In vitro and in vivo models for assessing the host response to biomaterials

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In vitro models to evaluate the permeability of poorly soluble drug entities: challenges and perspectives

The application of in vitro models in drug permeability studies represents a useful screening tool for assessing the biopharmaceutical appropriateness of new chemical entities (NCEs). Of note, there remains an ever-increasing number of NCEs which exhibit poor aqueous solubility. However, in their classical configuration, both cellular and non-cellular in vitro models show numerous deficiencies in their ability to accurately model the absorption of such compounds. As a consequence, investigators continue to explore the possibility of modifying different experimental parameters in an attempt to yield a more bio-relevant model system which offers good compatibility with poorly soluble compounds. Moreover, in many instances poorly soluble drugs necessitate the inclusion of excipients to facilitate efficient delivery and to enhance their bioavailability. Thus, there exists an increasing demand for in vitro models which can effectively appraise the effects of excipients on a drug's permeability. Herein, we provide an overview of those models currently in use and discuss their associated benefits and drawbacks. Furthermore, we review the challenges encountered in assaying the permeability of poorly soluble drugs and critically assess those experimental modifications and solutions employed thus far in terms of their capacity to generate results with improved accuracy and precision.     

In vitro and in vivo models of celiac disease

ABSTRACT

Introduction: Human in vitro blood-brain barrier (BBB) models could be important tools for studying BBB development, maintenance and regulation. However, our capacity to obtain information from these models is still limited in part because only in recent years have (i) these models been derived from non-brain cell sources (e.g. stem cells), (ii) microfluidic systems been developed to recapitulate aspects of BBB physiology and (iii) new insights into the molecular and cellular mechanisms of BBB diseases (e.g. Huntington´s, Allan-Herndon-Dudley Syndrome) been described.

Area covered: This article reviews the technological advances in the derivation of human cells from the neurovascular unit using stem cells and the creation of personalized BBB models generated from patients with neurodegenerative diseases. It also reviews the scientific advances generated from in vitro BBB models.

Expert opinion: The recent technological advances in the derivation of human cells from the neurovascular unit from stem cells as well as in the generation of BBB-on-a-chip that recapitulate in vitro part of the BBB physiology are significant to generate more robust BBB models; however, a considerable effort is still needed to validate the potential of these models to recapitulate the in vivo cellular and molecular mechanisms, in particular regarding BBB function in health and disease.     

Review: In vivo models for measuring placental glutatione-S-transferase (GST-P 7-7) levels: a suitable biomarker for understanding cancer pathogenesis

The Glutatione-S-transferases (GSTs) comprise a family of enzymes closely associated with the cell detoxification of xenobiotics. GSTs exist as homo- or heterodimers and have been grouped into at least seven distinct classes. The main function of GSTs is to catalyze the conjugation of reduced glutathione (GSH) to an electrophilic site of a broad range of potentially toxic and carcinogenic compounds, thereby making such compounds less dangerous and enabling their ready-excretion. Placental GST, known as GST-P 7-7, is the main isoform found in normal placental tissue and comprises 67% of the total GST concentration in this tissue. During development, GST-P 7-7 decreases in concentration and is absent in adult tissues. Interestingly, GST-P 7-7 expression has been detected in adult tissues after exposure to carcinogenic agents in several experimental test systems, being considered a reliable biomarker of exposure and susceptibility in early phases of carcinogenesis. In this article, we review a series of studies involving GST-P 7-7 expression as a suitable tool for understanding cancer pathogenesis, especially cancer risk.