Comparative Oncology Program Laboratory

比较肿瘤学项目实验室

• 批准号: 10262403
• 负责人: Amy Leblanc
• 金额: $ 134.22万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10262403
• 关键词: Address; Adolescent; Adult; Adverse event; Age; Agonist; Aliquot; Amputation; Animal Testing; Antineoplastic Agents; Area; Back; Biological; Biological Assay; Biological Markers; Biological Models; Biological Specimen Banks; Biology; Brain Neoplasms; Cancer Biology; Cancer Model; Cancer Patient; Canis familiaris; Carboplatin; Cell Line; Characteristics; Child; Childhood; Childhood Osteosarcoma; Clinical; Clinical Data; Clinical Trials; Collaborations; Collection; Companions; Complex; Conduct Clinical Trials; Custom; DNA; DNA Resequencing; DNA Sequence Alteration; DNA copy number; DNA sequencing; Data; Databases; Decision Making; Development; Disease; Drug Combinations; Drug Targeting; Early treatment; Engraftment; Event; Formalin; Freezing; Future; Gene Expression; Genes; Genome; Genomics; Goals; Growth; Health; Human; Human Cell Line; In Vitro; Infrastructure; Inherited; Investigation; Knowledge; Laboratories; Libraries; Link; Malignant Bone Neoplasm; Malignant Neoplasms; Metastatic Neoplasm to the Lung; Mission; Modeling; Molecular; Multi-Institutional Clinical Trial; Mus; Natural History; Neoplasm Metastasis; Normal tissue morphology; Oncogenes; Outcome; Pathologist; Pathology; Pathway interactions; Patients; Pediatric Neoplasm; Pharmaceutical Preparations; Phase; Physicians; Positioning Attribute; Pre-Clinical Model; Preclinical Drug Development; Preclinical Testing; Primary Neoplasm; Progression-Free Survivals; Proteomics; Puma; RNA; Reagent; Recurrent disease; Regimen; Resource Development; SNP array; Sampling; Single Nucleotide Polymorphism; Specimen; Structure; Survival Rate; Techniques; Testing; Time; Tissue Preservation; Toxicity Tests; Toxicogenomics; Translating; Treatment Failure; Treatment Protocols; Tumor stage; Work; Xenograft procedure; assay development; base; cancer clinical trial; cancer type; central database; chemotherapy; clinical application; cohort; comparative; comparative genomic hybridization; demographics; design; dog genome; drug development; drug efficacy; exome; exome sequencing; experimental study; genome-wide; genomic profiles; improved; improved outcome; in vivo; in vivo Model; inhibitor/antagonist; innovation; laboratory experiment; mouse model; neglect; new therapeutic target; novel therapeutic intervention; novel therapeutics; oncology program; osteosarcoma; pre-clinical; programs; repository; sarcoma; screening; success; transcriptome sequencing; translation to humans; treatment response; tumor

Laboratory activities and projects carried out by the COP laboratory have the specific goal of improving the understanding of the impact of anti-cancer agents on comparative aspects of metastasis biology by virtue of parallel study of murine, canine and human cell lines in a variety of in vitro, ex vivo/in vivo (Pulmonary Metastasis Assay or PuMA) model systems. Data generated in this manner improves understanding of naturally-occurring canine osteosarcoma (OS) models and could be employed to answer unique in vivo questions regarding the anti-metastatic potential of agents, via the COTC clinical trial mechanism. To extend our investigations into the comparative aspects of naturally-occurring OS in dogs to humans, we have recently initiated the DOG2 project: Decoding the Osteosarcoma Genome of the Dog. The DOG2 project fulfills the main mission of the NCI-COP, which is to strategically position the canine cancer patient in studies of cancer biology and drug development, in order to improve outcome for both dogs and humans. The COP has a longstanding focus in osteosarcoma (OS) biology and clinical trials. OS is an aggressive pediatric/AYA malignancy and the most common malignant bone tumor in children and adolescents. OS is also a common naturally-occurring canine cancer with strikingly similar clinical presentation and natural history; preliminary studies suggest a shared molecular landscape. While dogs largely develop OS in adulthood, the similar genomic features and clinical disease characteristics underscore the notion that age does not distinguish canine OS from the disease in children. Both human and canine patients with metastatic or relapsed disease have dismal outcomes, with survival rates less than 20% despite aggressive salvage regimens. We recognize that osteosarcoma is a complex disease and success is unlikely with a single approach. Therefore, to expand this work it is necessary to identify additional targets and drugs. Collectively this work is designed to address the following strategic priorities: 1. Improved knowledge of comparative OS cancer biology to enhance dog to human translation 2. Discovery of new targets and companion biomarkers in support of drug development 3. Assessment and prioritization of new therapeutic strategies in preclinical models 4. Harmonization of comparative canine and human oncology clinical trials to advance new therapeutic concepts Biospecimens. We will utilize an existing high-quality clinically-annotated biospecimen repository of 400 canine OS patient sample sets, unique to the NCI, gathered from ongoing and completed canine OS clinical trials. This repository contains multiple aliquots of both tumor and matched normal tissues, preserved as frozen, RNAlater, and formalin-fixed specimens. Samples have been derived from treatment-naive dogs that all underwent the same treatment protocol (amputation + 4 cycles of carboplatin chemotherapy). We have full clinical data on every patient including demographics, treatment details/adverse events, and event-free and overall survival. We propose beginning with a subset of n = 100 patient samples selected from the ends of a Kaplan-Meier curve that represent early treatment failures (progression-free survival less than 90 days) vs. elite responders (progression-free survival greater than 360 days). We will also query n = 25 matched pairs of tumors with their metastases that developed despite therapy. We have not attempted to perform viability testing for PDX engraftment in NSG mice; rather we use 4 unique canine OS cell lines developed in our lab for dog-in-mouse preclinical testing. All specimens will be subjected to pathology review by an expert board of veterinary and physician sarcoma pathologists to verify adequate sample quality for further genomic study. Genomic profiling. We will employ the following platforms to execute a combination of experiments to understand relationships between genomic complexity and integrity (CGH and WES to identify structural rearrangements, single nucleotide variants, copy number gain/loss), and gene expression (through RNAseq). The specific breed of dog will be verified through use of a germline SNP array recently made available through MARS/Wisdom Health. Samples for genomic profiling will be sent to our collaborator at NCSU (Matthew Breen) for DNA and RNA isolation, library prep, CGH for genome-wide DNA copy number profiling (Agilent CGH array), and for DNA whole exome sequencing (150b paired end reads at 100x for tumors/30x for normal, NovaSeq6000 platform with Roche dog exome kit), methyl-DNA resequencing, as well as RNA sequencing (125b paired end reads, Illumina HiSeq2500 platform). Computational approach. This project is an extension of an existing collaboration with NCSU and the NCATS Division or Preclinical Innovation (DPI) and Therapy for Rare and Neglected Diseases (TRND) program. Through experiments we have already conducted in n = 12 canine OS sample sets similar to those described above as a discovery cohort, the TRND bioinformaticians have developed a strategy that identifies distinct gene co-expression models between and specific to a set of 5 selected canine cancer types, of which OS is one, from which they derive cancer-specific gene panels. A similar analysis is performed on existing RNAseq data publicly available from human tumor samples from the same tumor types to produce cancer-specific human gene panels. These cancer-specific shared gene panels are compared, and genes common to both are retained to facilitate dog-to-human translation of genomic alterations. The RNAseq data, after alignment to reference, is then compared back to CGH and whole-exome DNA sequencing carried out in the same tumor sample to identify genomic changes (CNV, SNV) that are linked to expression changes and that may represent new druggable targets. These oncogenes or other drug targets in given pathway(s) were used to identify inhibitors, suppressors or agonists utilizing Pharos, which interfaces to a central database containing information about the targets collected by the Illuminating the Druggable Genome (IDG) program and the Comparative Toxicogenomic Database (CTD). This combined experimental/computational approach has already been tested using both proteomic and RNAseq data generated from the first n = 12 OS tumor/normal pairs. The data has been analyzed at NCATS to match changes in gene expression to drugs known to modulate the pathways associated with the associated genes. Canine OS cell line screening is already underway through NCATS to determine the in vitro impact of 40 drugs, including 30 synergistic combinations, based on the respective drugs' pathway modulation profile. These data serve as a proving ground for additional work carried out in a larger, outcome-linked, collection of tumor samples proposed herein. Single agents or combinations of drugs identified in Phase I will be tested in our existing panel of human and canine OS cell lines and screened rapidly through available xenograft human-in-mouse and canine-in-mouse models within the COP laboratory. This preclinical animal testing is critical to ranking and advancing appropriate candidates for further study and to understand the comparative features of how drugs behave in our in vivo models. We have expertise in both primary tumor and metastasis modelling to evaluate drug impact at several stages of tumor development and progression. These results will be the basis (go/no go decision making) for canine comparative oncology clinical trials to test the toxicity and efficacy of the drug combinations in vivo. Drug combination candidates that perform well in canine clinical trials would then be considered for human trials.

COP实验室开展的实验室活动和项目的具体目标是通过对鼠类，肺动物（肺动物转移分析或Puma）模型的鼠类，犬和人类细胞系的平行研究来提高对抗癌剂对转移生物学比较方面的影响的理解。以这种方式生成的数据改善了对自然存在的犬骨肉瘤（OS）模型的理解，并且可以通过COTC临床试验机制来回答有关药物抗转移潜力的独特问题。为了扩展我们对狗对人类自然疾病OS的比较方面的研究，我们最近启动了Dog2项目：解码狗的骨肉瘤基因组。 DOG2项目履行了NCI-COP的主要任务，该任务是在策略性地将犬类癌症患者定位在癌症生物学和药物开发研究中，以改善狗和人类的预后。 COP长期关注骨肉瘤（OS）生物学和临床试验。 OS是一种侵略性的儿科/AYA恶性肿瘤，是儿童和青少年最常见的恶性骨肿瘤。 OS也是一种常见的天然犬类癌，具有非常相似的临床表现和自然历史。初步研究表明共享分子景观。虽然狗在成年后的大部分开发OS，但类似的基因组特征和临床疾病特征强调了年龄不会区分儿童疾病的犬OS的观念。人类和犬类转移性疾病或复发性疾病的患者均具有令人沮丧的结果，尽管有积极的救助方案，但生存率却低于20％。我们认识到骨肉瘤是一种复杂的疾病，单一方法不太可能成功。因此，要扩大这项工作，有必要确定其他靶标和药物。这项工作旨在解决以下战略优先级：1。对比较OS癌症生物学的知识提高了对人类翻译的增强狗的知识。2。发现新的靶标和伴侣生物标志物以支持药物开发3.围栏模型中新的治疗策略的评估和优先级别4.比较罐头和人类Oncomological oncomologic incopic cline Clinic consecececececececic conspect conspectic conceptics概念化。我们将利用现有的高质量的临床临床注释的生物镜存储库，该存储库是400个犬OS患者样品集（NCI独有的），该样本是由正在进行的和完成的犬OS临床试验中收集的。该存储库包含肿瘤和匹配的正常组织的多个等分试样，这些组织被保存为冷冻，rnalater和福尔马林固定标本。样品是从均接受相同治疗方案的治疗犬（截肢 + 4个循环卡铂化学疗法）中得出的。我们拥有有关每个患者的完整临床数据，包括人口统计数据，治疗细节/不良事件以及无事件和整体生存。我们提出的是，从n = 100个患者样本的子集中从Kaplan-Meier曲线的末端选择，该样品代表早期治疗失败（无进展生存率小于90天）与精英响应者（无进展生存率大于360天）。我们还将查询n = 25对肿瘤对与它们的转移酶，尽管治疗了。我们尚未尝试对NSG小鼠的PDX植入进行生存测试。相反，我们使用在实验室中开发的4种独特的犬OS细胞系来进行幼时临床前测试。所有标本将受到兽医和医师肉瘤病理学家专家委员会的病理审查，以验证足够的样本质量以进行进一步的基因组研究。基因组分析。我们将采用以下平台来执行实验的组合，以了解基因组复杂性与完整性之间的关系（CGH和WES（CGH和WES）以识别结构重排，单核苷酸变体，拷贝数增益/损失）和基因表达（通过RNASEQ）。通过使用MARS/Wisdom Health提供的种系SNP阵列，将通过使用种系SNP阵列来验证狗的特定品种。用于基因组分析的样本将发送给我们在NCSU的合作者（Matthew Breen），用于DNA和RNA隔离，图书馆准备，CGH，CGH用于全基因组DNA副本拷贝数分析（Agilent CGH阵列），DNA全部外部序列（150B exts）（150B配对的tumore dog for tamase/30x）dog exase-kit exse dog exse dog exse not kit）重新陈述，以及RNA测序（125B配对端读，Illumina Hiseq2500平台）。计算方法。该项目是与NCSU和NCATS部门或临床前创新（DPI）的现有合作的扩展，以及针对罕见和被忽视的疾病（TRND）计划的治疗。通过实验，我们已经在N = 12犬OS样品集中进行了类似于上述发现队列的样品集，TRND生物信息学家已经制定了一种策略，该策略识别了与5种选定的犬类癌类型之间的不同基因共表达模型，其中一种OS是其中一种OS，它们从中得出了一种癌症基因基因。对现有的RNASEQ数据进行了类似的分析，可从相同肿瘤类型的人类肿瘤样品公开获得，以产生癌症特异性的人类基因板。比较了这些特定于癌症的共享基因面板，并保留了两者共有的基因，以促进基因组改变的狗至人类翻译。然后，将RNASEQ数据与参考后的对齐后，然后将其回到同一肿瘤样品中进行的CGH和全外活体DNA测序，以鉴定与表达变化相关的基因组变化（CNV，SNV），这些变化可能代表新的药物靶标。在给定途径中，这些癌症基因或其他药物靶标用于鉴定利用Pharos的抑制剂，抑制剂或激动剂，这些抑制剂，抑制剂或激动剂将其连接到中央数据库，这些数据库包含有关启发性药物基因组（IDG）程序和比较毒素毒素数据库（比较毒素数据库（CTD）收集的目标的信息）。这种合并的实验/计算方法已经使用了第一个N = 12 OS肿瘤/正常对产生的蛋白质组学和RNASEQ数据已经测试了。已经在NCAT上分析了数据，以使基因表达的变化与已知调节与相关基因相关的途径的药物匹配。犬类OS细胞系筛查已经在NCAT中进行，以确定40种药物的体外影响，包括30种协同组合，基于各自的药物的途径调节曲线。这些数据是在本文提出的较大，结合的肿瘤样本集合中进行的额外工作的探索基础。在我们现有的人类和犬OS细胞系列面板中，将对I期鉴定的药物的单一药物或组合进行测试，并通过COP实验室内的可用异种移植物和鼠标迅速筛选。这种临床前动物测试对于对进一步研究的适当候选者进行排名和推进至关重要，并了解药物在我们的体内模型中的表现的比较特征。我们在原发性肿瘤和转移模型中都有专业知识，可以评估肿瘤发育和进展的几个阶段的药物影响。这些结果将是犬比较肿瘤学临床试验的基础（GO/NO GO决策），以测试体内药物组合的毒性和功效。在犬临床试验中表现良好的候选药物组合将被考虑进行人体试验。