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Canine Glioma and Embryonic Neural Stem Cell Project

犬神经胶质瘤和胚胎神经干细胞项目

基本信息

批准号：
8552977
负责人：
Howard Fine
金额：
$ 63.3万
依托单位：
DIVISION OF BASIC SCIENCES - NCI
依托单位国家：
美国
项目类别：
财政年份：
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/8552977
关键词：
10q 10q21-q22 10q21.1 9p21-p22 Adult Animals Apoptosis Apoptosis Regulator Area Automobile Driving BMPR1A gene Belief Biology Boston Brain Neoplasms Breast Cancer Cell Breeding CDKN2A gene Canis familiaris Cell Cycle Cell Cycle Regulation Cell Proliferation Cells Chromosomal Duplication Chromosome Deletion Chromosomes Colon Carcinoma Cues Cyclic GMP Databases Embryo Employee Strikes Epidermal Growth Factor Receptor Event Exhibits Failure Gene Targeting Genes Genetic Genome Genomics Glial Differentiation Glioblastoma Glioma Gliomagenesis Growth Growth Factor Histopathologic Grade Human Human Genome IFNAR1 gene Incidence Individual Induction of Apoptosis Interferons Investigation Laboratories Link MDM2 gene Malignant - descriptor Malignant Glioma Malignant Neoplasms Maps Modeling Molecular Biology Neoplasms Neoplastic Cell Transformation Neuroglia Neuronal Differentiation Neurons Normal Cell Oncogenes PRKG2 gene PTEN gene Pathologic Pathway Analysis Pathway interactions Patients Phenotype Phosphotransferases Physiological Population Predisposition Pregnancy Receptor Inhibition Receptor Signaling Refractory Regulatory Pathway Reporting Role Sampling Secondary to Series Signal Transduction Syntenic Conservation The Cancer Genome Atlas Trisomy Tumor Cell Invasion Tumor Stem Cells Tumor Suppressor Genes Tumor Suppressor Proteins Tumor-Derived Tumorigenicity Xenograft procedure angiogenesis embryonic stem cell genetic regulatory protein improved insight interest neoplastic cell nerve stem cell novel oncology predictive modeling prevent progenitor smoothened signaling pathway transcription factor tumor tumor xenograft tumorigenesis tumorigenic vasodilator-stimulated phosphoprotein

项目摘要

The domestic dog represents the only model which allows all of these facets to be studied within the same species, and in respect to glioma TICs and adult NSCs, syngeneic samples from the same individual. As the ability to interrogate the canine genome increases, the clearly defined breed predispositions in regard to canine gliomagenesis may elucidate biologically relevant genomic alterations found in human glioma, enhancing our ability to classify this heterogeneous neoplasm by molecular biology, which may greatly improve our understanding over traditional pathologic grading schema.In order to more completely define minimal regions of alteration within large chromosomal amplifications or deletions often found in human glioblastoma, we identified regions of shared CNAs (between human GBMs and canine GSCs) and mapped these regions to the corresponding canine chromosomes. PTEN (located on HSA 10q.23.3) is usually deleted along with large segments of HSA 10q in human glioblastom. A long-standing question in glioma genetics is whether the target of HSA 10q deletion is PTEN alone or whether there are additional target genes that are co-deleted with PTEN that contribute to the tumorigenic phenotype.In the dog, PTEN is located at the telomeric end of CFA 26, which was deleted in the GSCs derived from the secondary and tertiary xenografts. Genes co-deleted in our canine GSCs alongside PTEN in CFA 26 included Dkk1, which functions to inhibit Wnt signaling and may function to inhibit differentiation potential or clonogenic growth potential. Additionally, cyclic guanosine monophosphate (cGMP)-dependent kinase (PRKG1) was also co-deleted with PTEN in the canine tertiary tumors and, alongside PRKG2, has been suggested to be potential tumor suppressor genes in colon carcinoma and glioma. PRKG1 also functions to inhibit angiogenesis through vasodilator-stimulated phosphoprotein (VASP). Thus, the deletion PRKG1 in both canine and human GSCs may have a mechanistic role in the aberrant cell cycle control and angiogenic phenotype of malignant gliomas.In addition to the deletions in CFA 26 containing PTEN (HSA 10q 89.2-90.9Mb), the canine tertiary xenograft tumors also exhibit an expanded deletion in CFA 4 that is syntenic to areas of HSA 10 that flank the PTEN loci and are often deleted in human GBMs (HSA 10q, 59.6-88.7 and 91-91.16Mb). Therefore, while the secondary and tertiary canine GSCs exhibit loss at CFA 26 corresponding to PTEN, progression of xenograft malignancy in our canine GSC population is associated with an additional, separate chromosomal deletion of CFA 4 that contains a syntenic region in immediate proximity to PTEN on the human genome. We examined genes present within this co-deleted segment of CFA 4 corresponding to HSA 10 and identified several genes within this region that have suspected roles in gliomagenesis. ANXA7 (located on HSA 10q21.1-q21.2), which is frequently deleted in human GBMs, has been previously suggested as a tumor suppressor gene independent of its human chromosomal proximity to PTEN. It has been hypothesized that loss of ANXA7 stabilizes and thus augments EGFR signaling and is negatively associated with patient survival. The loss of ANXA7 secondary to deletion of CFA 4, and separate from that containing PTEN (CFA 26), in canine gliomas supports an independent and important role for both genes as potential tumor suppressors in GBMs and demonstrates the power of the dog as a predictive model for interrogating the glioma genome. Additional genes co-deleted within CFA 4 and corresponding to the syntenic regions of HSA 10q involve other tumor suppressor gene candidates including BMPR1A, and CCAR1. We, and others have shown that disruptions in BMP signaling may impact the tumorigenic potential and capacity for differentiation in GSCs, and BMPR1A deletions are known to predispose individuals to colon cancer formation. CCAR1, or cell cycle and apoptosis regulator 1 (also referred to as cell cycle and apoptosis regulatory protein 1 or CARP-1) is located on HSA 10q21-q22 and has been reported to suppress the clonogenic growth, tumorigenicity, and invasion of human breast cancer cells and is integral to the induction of apoptosis following EGFR inhibition. Loss of CCAR1 may explain in part the failure of some patients to respond to EGFR inhibition therapeutically or enable tumor cells to survive in conditions of low growth factor concentration.We performed similar analyses on the other commonly altered genomic foci shared between our canine GSCs and human GBMs. The canine GSC genomic deletion containing CDK2NA is evolutionarily related to a very small region of HSA 9p21-p22, highlighting the importance of CDKN2A in glioma biology and diminishing the potential importance of a large number of other genes (passenger genes) that are co-deleted in the large CNAs found in HSA 9. Indeed, the canine chromosomal regions flanking the small locus containing CDKN2A are amplified, suggesting a specific role of CDKN2A deletion in glioma biology while potentially excluding a number of linked co-deleted genes in HSA 9. Within that minimally deleted region containing the p16/Ink4a locus in our canine GSCs are several interferon genes (IFNB1, IFNA5, IFNA13, and IFNA7) that are also deleted. This may be of interest for the IFNAR1 gene was recently identified by Cerami et al. as a linker gene in the PIK3R1 module through an automated network analysis (Human Interaction Network). This module, linking several IFNA genes and IFNB1 to IFNAR1, was found altered in 25% of GBMs in the TCGA database, but as the authors express, was of unknown significance due to the close proximity of IFN genes to CDKN2A. The co-deletion of CDKN2A and the IFNAR1 gene in both canine and human gliomas strengthens the argument that IFNAR1 is a potentially significant gene in the biology of GBMs.An additional chromosomal amplification associated with the more malignant xenograft gliomas is seen in CFA 6, which shares evolutionarily conserved synteny with HSA 7. This is of particular interest since trisomy HSA 7 is a common finding in GBMs. This region contains the gene, Glioblastoma Amplified Sequence (GBAS), which has been reported to be amplified in up to 40% of human GBMs. Other commonly altered foci in human GBMs show striking conservation in our canine GSCs including those surrounding amplification of MDM4, MDM2, and in the genomic regions commonly deleted in HSA 6 and 13. A number of these CNAs that are conserved across both species contain well-known potential tumor suppressor or oncogenes. One such example is Gli1, a transcription factor involved in the transduction of sonic hedgehog signaling and that may have a role in the promotion of tumor cell invasion. The Gli1 gene (HSA 12q13.2-13.3) is amplified both in a subset of human GBMs as well as in our canine glioma (CFA 10). By contrast, it is of interest that amplification of the epidermal growth factor receptor (EGFR) seen in primary human GBMs was not seen in our canine tertiary tumors, consistent with its lack of amplification in secondary human GBMs. Thus, these foci of shared genomic alterations allow us to identify a series of genomic events responsible for driving both human and canine gliomagenesis with more certainty than would be possible through the genomic study of gliomas from only one species.

家犬是唯一一种可以在同一物种中研究所有这些方面的模型，就神经胶质瘤tic和成人NSCs而言，它们是来自同一个体的同基因样本。随着研究犬基因组能力的提高，犬胶质瘤形成中明确定义的品种易感性可能会阐明在人类胶质瘤中发现的生物学相关基因组改变，增强我们通过分子生物学对这种异质性肿瘤进行分类的能力，这可能会大大提高我们对传统病理分级模式的理解。为了更完整地定义在人类胶质母细胞瘤中经常发现的大染色体扩增或缺失中的最小改变区域，我们确定了共享CNAs区域（在人类GBMs和犬GSCs之间），并将这些区域映射到相应的犬染色体上。PTEN（位于hsa10q .23.3上）在人胶质母细胞中通常与hsa10q的大片段一起被删除。胶质瘤遗传学中一个长期存在的问题是，HSA 10q缺失的目标是单独的PTEN，还是与PTEN共同缺失的其他靶基因有助于致瘤表型。在狗中，PTEN位于CFA 26的端粒末端，而CFA 26在二级和三级异种移植的GSCs中被删除。在CFA 26中，我们的犬GSCs中与PTEN共同缺失的基因包括Dkk1，它的功能是抑制Wnt信号传导，并可能抑制分化潜能或克隆生长潜能。此外，环鸟苷单磷酸（cGMP）依赖性激酶（PRKG1）也在犬三期肿瘤中与PTEN共缺失，并且与PRKG2一起被认为是结肠癌和胶质瘤中潜在的肿瘤抑制基因。PRKG1还通过血管扩张剂刺激磷酸化蛋白（VASP）抑制血管生成。因此，犬和人GSCs中PRKG1的缺失可能在恶性胶质瘤的异常细胞周期控制和血管生成表型中起机制作用。除了含有PTEN的CFA 26缺失（HSA 10q 89.2-90.9Mb）外，犬第三代异种移植物肿瘤也表现出CFA 4的扩展缺失，该缺失与PTEN位点旁边的HSA 10区域一致，并且在人类GBMs中经常缺失（HSA 10q, 59.6-88.7和91-91.16Mb）。因此，虽然二级和三级犬GSC在CFA 26处表现出与PTEN相对应的缺失，但在我们的犬GSC群体中，异种移植恶性肿瘤的进展与CFA 4的额外、单独的染色体缺失有关，该缺失包含人类基因组上与PTEN紧邻的合成区域。我们检测了CFA 4中与hsa10相对应的共缺失片段中的基因，并在该区域确定了几个可能在胶质瘤形成中起作用的基因。ANXA7（位于HSA 10q21.1-q21.2），在人类GBMs中经常被删除，以前被认为是一种独立于其与PTEN的人类染色体邻近性的肿瘤抑制基因。据推测，ANXA7的缺失稳定了EGFR信号，从而增加了EGFR信号，并与患者生存负相关。在犬胶质瘤中，由cfa4缺失引起的ANXA7缺失，与PTEN基因（cfa26）分离，支持了这两个基因在GBMs中作为潜在肿瘤抑制因子的独立和重要作用，并证明了犬作为胶质瘤基因组预测模型的能力。CFA 4中与HSA 10q合成区相对应的其他共缺失基因涉及其他肿瘤抑制基因候选基因，包括BMPR1A和CCAR1。我们和其他人已经表明，BMP信号的中断可能会影响GSCs的致瘤潜力和分化能力，并且BMPR1A缺失已知会使个体易患结肠癌。CCAR1或细胞周期和凋亡调节蛋白1（也称为细胞周期和凋亡调节蛋白1或CARP-1）位于HSA 10q21-q22上，据报道可抑制人乳腺癌细胞的克隆生长、致瘤性和侵袭，并且在EGFR抑制后诱导细胞凋亡是不可或缺的。CCAR1的缺失可能在一定程度上解释了一些患者在治疗上对EGFR抑制反应失败或使肿瘤细胞在低生长因子浓度条件下存活。我们对犬GSCs和人类GBMs之间共享的其他常见改变的基因组病灶进行了类似的分析。含有CDK2NA的犬GSC基因组缺失与HSA 9p21-p22的一个非常小的区域进化相关，突出了CDKN2A在胶质瘤生物学中的重要性，并降低了在HSA 9中发现的大CNAs中共同缺失的大量其他基因（乘客基因）的潜在重要性。事实上，犬染色体上含有CDKN2A的小位点两侧的区域被扩增，这表明CDKN2A缺失在胶质瘤生物学中具有特定作用，同时可能排除HSA 9中一些相关的共缺失基因。在我们的犬GSCs中包含p16/Ink4a位点的最小缺失区域中，也缺失了几个干扰素基因（IFNB1, IFNA5， IFNA13和IFNA7）。这可能与Cerami等人最近通过自动网络分析（Human Interaction network）发现IFNAR1基因是PIK3R1模块中的连接基因有关。该模块连接了几个IFNA基因和IFNB1与IFNAR1，在TCGA数据库中发现在25%的GBMs中发生了改变，但正如作者所表达的那样，由于IFN基因与CDKN2A非常接近，因此其意义未知。犬和人胶质瘤中CDKN2A和IFNAR1基因的共缺失强化了IFNAR1在GBMs生物学中是一个潜在的重要基因的观点。在CFA 6中发现了与更恶性的异种移植物胶质瘤相关的额外染色体扩增，它与HSA 7具有进化保守的同源性。这是特别有趣的，因为hsa7三体在GBMs中是常见的发现。该区域包含胶质母细胞瘤扩增序列（GBAS）基因，据报道，该基因在高达40%的人类GBMs中被扩增。人类GBMs中其他常见改变的病灶在我们的犬GSCs中显示出惊人的保守性，包括MDM4， MDM2扩增周围的区域，以及HSA 6和13中常见缺失的基因组区域。在这两个物种中保守的许多CNAs含有众所周知的潜在肿瘤抑制基因或致癌基因。Gli1就是这样一个例子，它是一种参与超音hedgehog信号转导的转录因子，可能在促进肿瘤细胞侵袭中起作用。Gli1基因（HSA 12q13.2-13.3）在人类GBMs和犬胶质瘤亚群中都有扩增（CFA 10）。相比之下，令人感兴趣的是，在原发性人GBMs中发现的表皮生长因子受体（EGFR）扩增在犬三期肿瘤中未见，这与在继发性人GBMs中缺乏扩增一致。因此，这些共享基因组改变的焦点使我们能够确定一系列负责驱动人类和犬胶质瘤形成的基因组事件，比通过仅来自一个物种的胶质瘤的基因组研究更确定。