In vitro models of pancreatic cancer for translational oncology research

Background: Pancreatic cancer is a disease of near uniform fatality and the overwhelming majority of patients succumb to their advanced malignancy in a few months of diagnosis. Despite considerable advances in our understanding of molecular mechanisms underlying pancreatic carcinogenesis, this knowledge has not yet been fully translated into clinically available treatment strategies that yield significant improvements in disease free or overall survival. Objective: Cell line-based in vitro model systems provide powerful tools to identify potential molecular targets for therapeutic intervention as well as for initial preclinical evaluation of novel drug candidates. Here, we provide a brief overview of recent literature on cell line-based model systems of pancreatic cancer and their application in the search for novel therapeutics against this vicious disease. Conclusion: Although in vitro models of pancreatic cancer are of tremendous value for genetic studies and for initial functional screenings in drug discovery, they carry several immanent drawbacks and are often poor in predicting therapeutic response in humans. Therefore, in most instances, they are successfully exploited to generate hypothesis and identify molecular targets for novel therapeutics, which are subsequently subject to further in-depth characterization using more advanced in vivo model systems and clinical trials.     

MicroRNAs as therapeutic targets and their potential applications in cancer therapy

Introduction: The results of cancer-associated miRNA research have yielded surprising insights into the pathogenesis of a range of different cancers. Many of the dysregulated miRNAs are involved in the regulation of genes that are essential for carcinogenesis.

Areas covered: This review discusses the latest discovery of miRNAs acting as oncogenes and tumor suppressor genes, as well as the potential applications of miRNA regulations in cancer therapy. Several translational studies have demonstrated the feasibility of targeting oncogenic miRNAs and restoring tumor-suppressive miRNAs for cancer therapy using in vivo model systems.

Expert opinion: miRNAs are extensive regulators of cancer progression. With increasing understanding of the miRNA target genes and the cellular behaviors influenced by them, modulating the miRNA activities may provide exciting opportunities for cancer therapy. Despite the hurdles incurred in acquiring effective systemic drug delivery systems, in vivo delivery of miRNAs for therapeutic purposes in preclinical animal models is rapidly developing. Accumulating evidences indicate that using miRNA expression alterations to influence molecular pathways has the potential of being translated into clinical applications.     

Challenges with in vitro and in vivo experimental models of urinary bladder cancer for novel drug discovery

ABSTRACT

Introduction: Urinary bladder cancer (UBC) is the second most frequent malignancy of the urinary system and the ninth most common cancer worldwide, affecting individuals over the age of 65. Several investigations have embarked on advancing knowledge of the mechanisms underlying urothelial carcinogenesis, understanding the mechanisms of antineoplastic drugs resistance and discovering new antineoplastic drugs. In vitro and in vivo models are crucial for providing additional insights into the mechanisms of urothelial carcinogenesis. With these models, various molecular pathways involved in urothelial carcinogenesis have been discovered, allowing therapeutic manipulation.

Areas covered: This paper provides critical information on existing in vitro and in vivo models to screen the efficacy and toxicity of innovative UBC therapies and point out the challenges for new and improved models.

Expert opinion: In our opinion, results obtained with in vitro and in vivo models should be interpreted together, as a set of delicate biological tools that can be used at different stages in the drug discovery process, to address specific questions. With the development of new technologies, new assays and biomarkers are going to play an important role in the study of UBC. The molecular diagnostics and genomic revolution will not only help to develop new drug therapies, but also to achieve tailored therapies.     

Drug discovery in prostate cancer mouse models

Introduction: The mouse is an important, though imperfect, organism with which to model human disease and to discover and test novel drugs in a preclinical setting. Many experimental strategies have been used to discover new biological and molecular targets in the mouse, with the hopes of translating these discoveries into novel drugs to treat prostate cancer in humans. Modeling prostate cancer in the mouse, however, has been challenging, and often drugs that work in mice have failed in human trials.

Areas covered: The authors discuss the similarities and differences between mice and men; the types of mouse models that exist to model prostate cancer; practical questions one must ask when using a mouse as a model; and potential reasons that drugs do not often translate to humans. They also discuss the current value in using mouse models for drug discovery to treat prostate cancer and what needs are still unmet in field.

Expert opinion: With proper planning and following practical guidelines by the researcher, the mouse is a powerful experimental tool. The field lacks genetically engineered metastatic models, and xenograft models do not allow for the study of the immune system during the metastatic process. There remain several important limitations to discovering and testing novel drugs in mice for eventual human use, but these can often be overcome. Overall, mouse modeling is an essential part of prostate cancer research and drug discovery. Emerging technologies and better and ever-increasing forms of communication are moving the field in a hopeful direction.     

Three-Dimensional Cultures of Prostatic Cells: Tissue Models for the Development of Novel Anti-Cancer Therapies

This review addresses the application of three-dimensional cultures of prostatic cells to the development of novel anti-cancer therapies. A variety of therapeutic agents to combat prostate cancer are currently under development. These include cytotoxins, differentiation agents and, more recently, genetically modified tumor vaccines. Three-dimensional cultures of prostatic cells are increasingly used in preclinical research in the design of new therapies and in the development of delivery strategies for these treatments. These tissue-like structures more realistically model the structural architecture and differentiated function of the human prostate than a cellular monolayer. In doing so, three-dimensional cultures produce an in vivo-like response to therapeutic agents. Advances in tissue engineering have improved the variety, fidelity and quantity of these prostate models. To date, they have been applied to estimate the dose of new drug therapies, evaluate drug penetration into solid tumors, assess the effectiveness of drug combinations, and develop tumor vaccines.     

Protein kinase inhibitors in metastatic colorectal cancer. Let's pick patients,tumors, and kinase inhibitors to piece the puzzle together!

Introduction: Increased understanding in intracellular signaling pathways leading to carcinogenesis, proliferation, migration, invasion, angiogenesis, and anti-apoptosis of colorectal cancer cells has been critical for target identification and drug development. Specific protein kinase inhibitors (KIs) have been developed to block activated pathways associated with tumor growth and progression. Although showing promising activity in preclinical models, until now, the majority of KIs were not able to demonstrate clinically meaningful efficacy in Phase II/III trials.

Areas covered: The major pathways altered in colorectal cancer will be highlighted, and molecularly defined targets will be discussed. The mechanisms of action and the proof of principle demonstrated in preclinical models of KIs and the disappointing efficacy in clinical trials will be reviewed.

Expert opinion: Despite recent negative study results, KIs have the potential to be the next class of therapeutics in the treatment of metastatic colorectal cancer. Molecular classification of the individual tumors and identification of molecular escape mechanisms for primary (intrinsic) and secondary resistances to KI treatment is critical to select the patients' most likely to benefit. Appropriate drug combinations based on those mechanisms of resistance have to be tested in selected patient populations to ensure progress and efficacy with the goal to lead to a clinically meaningful prolongation of patients' lives.     

Colorectal cancer models for novel drug discovery

Introduction: Despite increased screening rates and advances in targeted therapy, colorectal cancer (CRC) remains the third leading cause of cancer-related mortality. CRC models that recapitulate key features of human disease are essential to the development of novel and effective therapeutics. Classic methods of modeling CRC such as human cell lines and xenograft mice, while useful for many applications, carry significant limitations. Recently developed in vitro and in vivo models overcome some of these deficiencies and thus can be utilized to better model CRC for mechanistic and translational research.

Areas covered: The authors review established models of in vitro cell culture and describe advances in organoid culture for studying normal and malignant intestine. They also discuss key features of classic xenograft models and describe other approaches for in vivo CRC research, including patient-derived xenograft, carcinogen-induced, orthotopic transplantation and transgenic mouse models. We also describe mouse models of metastatic CRC.

Expert opinion: No single model is optimal for drug discovery in CRC. Genetically engineered models overcome many limitations of xenograft models. Three-dimensional organoids can be efficiently derived from both normal and malignant tissue for large-scale in vitro and in vivo (transplantation) studies and are thus a significant advance in CRC drug discovery.     

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.     

我国临床前药物致癌试验转基因动物模型研究进展

目的：综述我国临床前药物致癌试验转基因动物模型研究进展。方法：介绍致癌试验常用转基因动物模型的构建、利用转基因动物模型开展致癌试验的优势、国际认可及国际监管机构新药申报中的应用、国内外致癌试验相关指导原则、我国开展基于转基因动物致癌试验面临的困境以及转基因动物模型构建的最新进展。结果与结论：基于转基因动物模型的致癌试验具有诸多优势也是目前国际趋势,在我国建立临床前药物致癌试验转基因动物模型非常必要。     

Precision-cut intestinal slices: alternative model for drug transport,metabolism, and toxicology research

Introduction: The absorption, distribution, metabolism, excretion and toxicity (ADME-tox) processes of drugs are of importance and require preclinical investigation intestine in addition to the liver. Various models have been developed for prediction of ADME-tox in the intestine. In this review, precision-cut intestinal slices (PCIS) are discussed and highlighted as model for ADME-tox studies.Areas covered: This review provides an overview of the applications and an update of the most recent research on PCIS as an ex vivo model to study the transport, metabolism and toxicology of drugs and other xenobiotics. The unique features of PCIS and the differences with other models as well as the translational aspects are also discussed.Expert opinion: PCIS are a simple, fast, and reliable ex vivo model for drug ADME-tox research. Therefore, PCIS are expected to become an indispensable link in the in vitro–ex vivo–in vivo extrapolation, and a bridge in translation of animal data to the human situation. In the future, this model may be helpful to study the effects of interorgan interactions, intestinal bacteria, excipients and drug formulations on the ADME-tox properties of drugs. The optimization of culture medium and the development of a (cryo)preservation technique require more research.     

适配体C2min介导的可靶向2种前列腺癌基因的递送系统

目的 通过合成可靶向两种前列腺癌的基因载体PAMAM-PEG-C2min,以提高基因的转染效率和肿瘤靶向性。方法 将双功能聚乙二醇的一端与聚酰胺-胺（PAMAM）相连,另一端与适配体（C2min）连接,并利用¹H NMR技术对合成的PAMAM-PEG-C2min基因载体进行结构鉴定。通过两种前列腺癌PC3和LNCaP细胞的体外摄取和基因转染实验（包载siR-M基因）,考察纳米复合物的生物学特性。并利用动物活体成像技术考察合成的纳米复合物的体内分布特征。结果 核磁共振结果表明,本研究成功合成了PAMAM-PEG-C2min。PC3和LNCaP细胞对PAMAM-PEG-C2min的摄取结果体现出浓度依赖性。且与不经C2min修饰的PAMA-PEGM相比,PAMAM-PEG-C2min递药系统的基因转染效率和肿瘤细胞靶向性明显提高。体内靶向性结果表明,PAMAM-PEG-C2min可实现同时靶向2种前列腺癌组织的作用。结论 本研究合成的PAMAM-PEG-C2min递送载体具有良好的肿瘤靶向性,为前列腺癌的综合治疗和靶向治疗提供了新的技术平台。     

Animal and cellular models for hypolipidemic drugs

Background: The development of effective and safe lipid-lowering agents should set out from and rely on robust preclinical investigation. Objective: To accomplish this aim, the selection of proper cellular and animal models is crucial. Results: Because lipid-lowering agents are ultimately supposed to reduce the atherosclerotic burden in the arterial wall, they need to tackle directly or indirectly the multifactorial nature of atherosclerotic disease. Hence, these drugs may essentially prevent triglyceride-rich lipoprotein assembly or enhance low-density lipoprotein (LDL) clearance through the LDL or related receptors in the liver. Established animal models such as the apolipoprotein E- and the LDL-receptor knockout mice are widely used to test drug actions on these pathways. A different approach is testing the ability of candidate drugs to increase plasma high-density lipoprotein (HDL) levels. More recently, the focus has shifted to drugs enhancing HDL function rather than just plasma HDL levels. This in turn requires in vitro and particularly in vivo models of reverse cholesterol transport, which have become available by now. Conclusion: A positive outcome of preclinical studies is necessary but not sufficient for an investigational new drug to be eventually approved for clinical use.     

Animal models of urinary bladder cancer and their application to novel drug discovery

Introduction: Urinary bladder cancer is a major human malignancy that afflicts millions of people worldwide every year. Urinary bladder cancer is usually superficial at presentation in 70 – 80% of patients. In these cases, a simple transurethral resection is adequate for removing the tumor. However, some patients experience recurrence or even tumor progression. In another 20 – 30% of patients, muscle-invasive carcinoma is diagnosed. Despite all the developments in this area, even today, the options for treatment of urinary bladder cancer remain inadequate. The search for the mechanisms involved in human urinary bladder cancer and for new and improved treatment methods has led to the development of many experimental models using laboratory animals over the past 40 years.

Areas covered: In this review, the authors provide a concise overview of the animal models used to study urinary bladder cancer. Furthermore, the authors discuss their advantages and disadvantages with regard to the search for new therapeutic approaches.

Expert opinion: The use of urinary bladder cancer models for understanding the mechanisms involved in tumors' response to new treatments is an important step in the drug discovery process. However, the authors believe that it will be necessary to develop our knowledge and understanding of the molecular processes underlying urothelial chemical carcinogenesis for us to better evaluate the efficacy of novel therapeutics.     

Orthotopic mouse models expressing fluorescent proteins for cancer drug discovery

Importance of the field: Currently used rodent tumor models, including transgenic tumor models, or subcutaneously growing human tumors in immunodeficient mice, do not sufficiently represent clinical cancer, especially with regard to metastasis and drug sensitivity.

Areas covered in this review: To obtain clinically accurate models, we have developed the technique of surgical orthotopic implantation (SOI) to transplant histologically intact fragments of human cancer, including tumors taken directly from the patient, to the corresponding organ of immunodeficient rodents. SOI allows the growth and metastatic potential of the transplanted tumors to be expressed and reflects clinical cancer of all types. Effective drugs can be discovered and evaluated in the SOI models utilizing human tumor cell lines and patient tumors. Visualization of many aspects of cancer initiation and progression in vivo has been achieved with fluorescent proteins. Tumors and metastases in the SOI models that express fluorescent proteins can be visualized noninvasively in intact animals, greatly facilitating drug discovery.

What the reader will gain: This review will provide information on the imageable mouse models of cancer that are clinically relevant, especially regarding metastasis and their use for drug discovery and evaluation.

Take home message: SOI mouse models of cancer reproduce the features of clinical cancer.     

Use of behavioural and long-term potentiation models in the development of memory-improving drugs

Background: There is a great need to develop memory-enhancing drugs for the treatment of memory dysfunctions. Although many targets have been identified in preclinical studies, the number of clinically effective drugs is limited. Objective: In this overview, the relation between drug effects on hippocampal long-term potentiation (LTP) and memory-enhancing effects is explored for drugs that modulate cholinergic or glutamatergic neurotransmission or inhibit cyclic nucleotide metabolism. Methods: We limited our analysis to drug targets that are in clinical use or in development for the treatment of cognitive deficits and that have been tested in LTP. Results/conclusion: Although these drugs have been shown to improve in vitro or in vivo LTP and memory performance in many different models, no clear correlation between positive effects in LTP and behavioural assays is possible. The effectiveness in behavioural models is based on various models assessing different aspects of cognition. More uniformity in the use of behavioural tests is encouraged to better understand drug effects on different memory processes (i.e., acquisition, consolidation and retrieval). The systematic use of in vitro models such as LTP improves our understanding of the molecular mechanism of drug efficacy. This may lead to a better selection of models relevant to the specific cognitive processes disrupted in different pathological conditions and a more differentiated development of memory-enhancing drugs.     

Preclinical drug development for childhood cancer

Importance of the field: More effective drugs are needed to treat poor prognosis paediatric malignancies. Development of anticancer agents for childhood cancers faces several unique challenges compared with their adult counterparts.

Areas covered in this review: We demonstrate how recent advances in preclinical drug development may overcome these difficulties and challenges. We explain the role of academia, regulators and industry in this field, address issues with preclinical models and illustrate several examples of biology-driven drug development in childhood cancers.

What the reader will gain: Increased knowledge about preclinical drug development in paediatric oncology including different preclinical models, established preclinical research networks, and relationships among academia, industry and regulators, as illustrated by several examples of targeted agents in childhood solid malignancies.

Take home message: It is anticipated that emerging advanced preclinical models and testing platforms will provide a more efficient, biologically-driven rationale to support the use of targeted therapies in several malignancies such as neuroblastoma, medulloblastoma or high grade glioma which account for the majority of deaths related to childhood cancer.     

Finding the place of histone deacetylase inhibitors in prostate cancer therapy

Histone deacetylase inhibitors (HDACIs) are showing promise as therapeutic agents for hematological malignancies and solid tumors. In the case of prostate cancer, HDACIs are effective at inhibiting proliferation and inducing apoptosis in a range of in vitro and in vivo experimental models. Recent studies have revealed that the actions of HDACIs in prostate cancer cells extend beyond regulation of histone acetylation and affect proteins involved in maintaining cellular homeostasis and tumor progression, including the androgen receptor, p21^WAF1 and VEGF. The broad spectrum of HDACI targets has allowed rational design of combinations with other therapeutic agents to target multiple pathways involved in prostate cancer progression, including angiogenesis and androgen signaling. In particular, synergistic inhibition of prostate cancer cell growth has been demonstrated using HDACIs in combination with radio- and chemo-therapy, Apo2L/TRAIL, angiogenesis inhibitors, heat-shock protein 90 inhibitors and androgen receptor antagonists. This review examines the current understanding of the actions of HDACIs in prostate cancer cells, both in a laboratory and a clinical context and discusses the potential utility of combination strategies for the treatment of prostate cancer.     

纳米药物制剂体内分析方法及药动学研究进展和问题策略分析

近年来随着纳米技术的不断发展,纳米药物制剂在改善药物递送、提高生物利用度方面显示出独特优势,已成为临床新药开发研究的热点,为诸多疾病尤其是恶性肿瘤的治疗提供了新思路。然而,由于对纳米药物制剂的体内过程了解不够全面,导致纳米药物制剂的临床转化率极低,严重制约了纳米药物制剂的发展。基于纳米药物制剂良好的应用前景及目前药动学研究中存在的关键问题,调研了国内外的相关文献,首先介绍了常见的具有不同纳米载体类型的纳米药物制剂的种类,对纳米药物制剂体内药物浓度分析测定的方法进行归纳总结,最后分析纳米载体的理化性质对纳米药物制剂体内药动学行为的影响,旨在为纳米药物制剂的体内过程研究提供参考,以获取更为全面的体内药动学数据,提高药物的临床转化率。同时针对目前纳米药物制剂的载体研究、体内浓度定量分析方法以及药动学研究中存在的问题进行讨论,以期为纳米药物制剂的研究与开发利用提供方向。     

The role of early in vivo toxicity testing in drug discovery toxicology

The cost impact of late-stage failures of drug candidates has motivated the pharmaceutical industry to develop, validate, and implement a more proactive testing paradigm, including an emphasis on conducting predictive in vitro and in vivo studies earlier. The goal of drug discovery toxicology is not to reduce or eliminate attrition, as is often mis-stated as such, but rather to reprioritize efforts to shift attrition of future failing molecules upstream in discovery. This shift in attrition requires additional studies and investment earlier in the candidate evaluation process in order to avoid spending resources on molecules with soon-to-be-discovered development-limiting liabilities. While in silico and in vitro models will continually be developed and refined, in vivo preclinical safety models remain the gold standard for assessing human risk. For in vivo testing to influence early discovery effectively, it must: i) require low amounts of compound; ii) provide rapid results to drive decision-making and medicinal chemistry efforts; and iii) be flexible and provide results relevant to the development plan tailored to each target, drug class, and/or indication.