Three-dimensional Model of Human Ewing Sarcoma

人类尤文肉瘤三维模型

• 批准号: 9130807
• 负责人: Fred Kurtis Kasper
• 金额: $ 32.27万
• 依托单位: RICE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-11 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9130807
• 关键词: Affect; Antineoplastic Agents; Architecture; Biomechanics; Biomimetics; Bioreactors; Blood Vessels; Bone neoplasms; Carcinoma; Cell Communication; Cell Differentiation process; Cell Survival; Cells; Characteristics; Chondrosarcoma; Clinical; Clinical Trials; Coculture Techniques; Colon Carcinoma; Complement; Complex; Cues; Deposition; Dimensions; Drug Targeting; Drug resistance; Endothelial Cells; Engineering; Ewings sarcoma; Exhibits; Extracellular Matrix; FRAP1 gene; Feedback; Goals; Health; Human; Hybrids; IGF1 gene; Institutional Review Boards; Insulin-Like Growth Factor I; Invaded; Laboratories; Left; Ligands; Malignant Neoplasms; Malignant neoplasm of lung; Mesenchymal Stem Cells; Microscopic; Modeling; Molecular Abnormality; Nature; Neoplasm Metastasis; Nutrient; Oxygen; Pathway interactions; Patients; Pattern; Perfusion; Polymers; Pre-Clinical Model; Proteomics; Publishing; Reporting; Research; Resistance; Role; SDZ RAD; Sampling; Signal Pathway; Signal Transduction; Specimen; Testing; Tissue Engineering; Tumor Cell Line; Tumor-Derived; Ursidae Family; Vascular Endothelial Growth Factors; bone; caprolactone; clinical investigation; clinically relevant; drug testing; in vivo; innovation; mTOR Inhibitor; malignant breast neoplasm; monolayer; neoplastic cell; novel; novel strategies; osteogenic; osteosarcoma; protein expression; resistance mechanism; scaffold; shear stress; stem; targeted treatment; therapeutic target; three dimensional cell culture; three-dimensional modeling; tumor; tumor growth; tumor microenvironment; tumorigenic; vasculogenesis

DESCRIPTION (provided by applicant): The overall goal of the proposed research is to apply static and flow perfusion bioreactor culture of bone sarcoma cells, grown upon a tissue-engineered polymer/extracellular matrix (ECM) hybrid model that reliably mimics key features of the bone tumor niche, to advance our understanding of the IGF-1R/mTOR cancer- related pathway and its clinically-relevant resistance mechanisms. Our laboratory has reported in vivo-like IGF- 1R/mTOR expression patterns, closely related to those observed in human Ewing sarcoma tumors (ES), when established ES cell lines are grown upon innovative biologically inert 3D electrospun poly(�-caprolactone) (PCL) microfiber scaffolds rather than upon traditional plastic monolayers. The present proposal seeks to elucidate the precise mechanisms by which cell-cell and cell-ECM interactions stimulate an activated IGF- 1R/mTOR signaling state within this engineered 3D bone tumor microenvironment, as those interactions are critically important in initiating and maintaining ES. In parallel with determining the influence of those parameters under static cell culture, 3D scaffolds and culture conditions will be adapted to better emulate the native bone microenvironment: (a) varied flow perfusion rates will be used to assess the effect of shear stress upon cell retention while facilitating uniform distribution of Ewing cells within the scaffold, (b) PCL scaffolds will incorporate IGF-1 to mimic the high concentration of IGF-1 naturally released as tumors invade surrounding bone, (c) the effect of an osteogenic ECM upon the IGF-1R/mTOR signaling cascade will be determined using decellularized scaffolds upon which heterotypic mesenchymal stem cells, differentiated toward an osteoblastic lineage, are first grown, and (d) ES cells will be co-cultured with endothelial cells (EC) to determine how heterotypic cells interact within a fabricated 3D bone tumor model to elicit viable ES tumors. Finally, to determine the mechanism(s) by which Ewing sarcoma evades sensitivity to combined mTOR/IGF-1R targeted therapy, freshly-derived tumor specimens (obtained from Ewing sarcoma patients treated with Medi- 573/everolimus in an IRB-approved clinical trial), will be grown in primary culture within 3D scaffolds and compared to 2D culture and patient-derived tumor explants (PDTX) by proteomic analysis of the IGF- 1R/mTOR pathway and putative resistance mechanisms. This novel approach of studying Ewing sarcoma within an ex vivo preclinical model of the bone microenvironment presents tremendous potential for understanding chemotherapeutic efficacy and for determining resistance mechanisms to biologically targeted therapies.

描述（由申请人提供）：拟议研究的总体目标是应用骨肉瘤细胞的静态和流动灌注生物反应器培养，在组织工程聚合物/细胞外基质（ECM）混合模型上生长，可靠地模拟骨肿瘤生态位的关键特征，以促进我们对IGF-1R/mTOR癌症相关途径及其临床相关耐药机制的理解。我们的实验室报告了体内样IGF- 1R/mTOR表达模式，与在人类尤文氏肉瘤肿瘤（ES）中观察到的表达模式密切相关，当建立的ES细胞系生长在创新的生物惰性3D电纺丝聚己内酯（PCL）微纤维支架上而不是传统的塑料单层上时。目前的建议旨在阐明细胞-细胞和细胞- ecm相互作用在这种工程化的3D骨肿瘤微环境中刺激激活的IGF- 1R/mTOR信号状态的确切机制，因为这些相互作用在启动和维持ES中至关重要。在确定这些参数在静态细胞培养下的影响的同时，将调整3D支架和培养条件，以更好地模拟天然骨微环境：(a)不同的血流灌注率将用于评估剪切应力对细胞保留的影响，同时促进支架内尤文氏细胞的均匀分布；(b) PCL支架将加入IGF-1来模拟肿瘤侵入周围骨骼时自然释放的高浓度IGF-1； (c)成骨性ECM对IGF-1R/mTOR信号级联的影响将使用去细胞化支架来确定，在去细胞化支架上异型间充质干细胞，(d)胚胎干细胞将与内皮细胞（EC）共培养，以确定异型细胞如何在制造的3D骨肿瘤模型中相互作用，从而引发可存活的胚胎干细胞肿瘤。最后，为了确定Ewing肉瘤逃避mTOR/IGF-1R联合靶向治疗敏感性的机制，将在3D支架内的原代培养中培养新鲜来源的肿瘤标本（来自在irb批准的临床试验中接受Medi- 573/依维莫司治疗的Ewing肉瘤患者），并通过对IGF-1R /mTOR途径的蛋白质组学分析和推测的耐药机制，将其与2D培养和患者来源的肿瘤外植体（PDTX）进行比较。这种在骨微环境的离体临床前模型中研究尤文氏肉瘤的新方法为了解化疗疗效和确定生物靶向治疗的耐药机制提供了巨大的潜力。