溶瘤病毒抗神经胶质瘤治疗研究中的问题与对策

神经胶质瘤是人脑中最常见的原发性肿瘤,占中枢神经系统恶性肿瘤的81%,当前标准疗法仍是手术切除及术后放化疗。因神经胶质瘤具有高侵袭性、分子异质性、治疗后耐药肿瘤干细胞可再生,以及化疗药物难以通过血脑屏障（BBB）达到足够高的治疗浓度等特点,导致其预后非常差,患者中位存活期仅为15个月。近年来,新兴的溶瘤病毒免疫疗法治疗神经胶质瘤的研究备受关注并取得一定进展,但依然面临诸如BBB、免疫“冷”微环境、宿主抗病毒反应和肿瘤高度异质性等挑战。这些问题限制了溶瘤病毒疗法的深入发展及进一步应用,但也给基础与临床研究者带来新的研究机遇。因此,本文从穿越BBB、改善肿瘤微环境（TME）、调控溶瘤病毒介导的宿主免疫反应和适应肿瘤异质性等四个方面,阐述溶瘤病毒在抗神经胶质瘤治疗研究中的存在问题及对策。     

生物3D打印在肿瘤研究及组织工程中的应用

近年来生物3D打印技术快速发展,已成为肿瘤研究与组织工程领域用于组织构建、机制研究、药物评价及药物递送等研究的重要工具。本综述总结生物3D打印的基本原理及其在肿瘤研究和组织工程中的应用进展。生物3D打印是一种增材制造技术,通过数字控制逐层堆叠生物材料和活细胞以构建复杂的三维组织结构,其核心步骤是设计3D模型、选择合适的生物打印技术和材料、逐层打印、后期培养和功能化处理。在肿瘤研究中,生物3D打印可用于构建模拟肿瘤微环境的复杂模型,揭示肿瘤的发生、发展新机制。传统体外模型如二维细胞培养或动物模型难以准确模拟人类肿瘤的复杂性,而通过生物3D打印技术构建更仿生的3D肿瘤模型,模拟肿瘤细胞与免疫细胞、基质、血管等环境的动态相互作用,能够提供更接近真实肿瘤生长、侵袭及转移的研究平台。此外,生物3D打印为抗癌药物的开发、创新治疗策略的确立和个性化治疗方案的制订提供了创新平台,3D打印肿瘤模型能够提供更贴近临床的实验结果且具备高通量药物筛选的能力,可广泛应用于细胞毒类药物、靶向治疗药物和免疫治疗药物等多种类型的药物评价中;除药物开发外,生物3D打印还为肿瘤辅助治疗提供了新的解决思路。生物3D打印模型和支架,可用于个性化精准治疗,通过高效构建患者细胞构成的个性化3D模型预测患者对药物及放疗的敏感性,可建立局部支架,根据患者具体需求确定合适的药物剂型、剂量等。另外,3D打印支架可用于辅助药物递送,利用3D支架靶向递送药物或减弱药物引起的不良反应,还可辅助局部免疫检查点抑制剂疗法、局部细胞因子疗法、局部癌症疫苗疗法及局部嵌合抗原受体修饰的细胞疗法。在组织工程中,传统的组织修复方法通常难以应对复杂组织的构建需求,而生物3D打印为构建复杂组织结构和实现组织再生提供了全新的思路,骨与软骨、皮肤等结构较为基础且具备较高再生能力的组织和器官已逐渐进入临床实践,肝脏、心脏等复杂器官的修复和重建也已取得一定进展,但尚未实现临床转化。最后,本综述探讨了生物3D打印在上述领域面临的挑战及未来发展方向,以期为相关领域的研究人员提供有价值的参考。     

神经肿瘤干细胞在恶性胶质瘤放射治疗中的研究进展

脑胶质瘤是最常见的神经系统原发肿瘤。放射治疗是目前脑胶质瘤综合治疗最重要的治疗手段之一。然而经过放射治疗以后,部分肿瘤又会复发。随着肿瘤干细胞学说的确立,神经肿瘤干细胞的成功筛选和分离,研究者开始对其生物学特性进行了一系列基础研究,从肿瘤干细胞角度解释其放射生物学行为。放射肿瘤学研究者探求从治疗上特异于杀伤神经肿瘤干细胞,靶向作用于其相关信号转导通路,以期能为临床诊疗提供新的思路,最终使恶性胶质瘤临床放射治疗受益。     

MGMT基因甲基化检测在神经胶质瘤治疗中的应用

摘 要：神经胶质瘤是一种常见的、复杂的恶性肿瘤,现有的治疗方法仍不能明显提高患者的生存期。近年来,越来越多的研究表明MGMT启动子甲基化状态与神经胶质瘤诊断、治疗和预后密切相关。编码修复O6-甲基鸟嘌呤的MGMT可以帮助肿瘤细胞逃避烷化剂的细胞毒作用,但MGMT启动子常会因为发生甲基化降低基因的表达,最终阻碍DNA的修复。研究已证实MGMT基因甲基化是一种可靠的胶质瘤预测标志物,且MGMT基因甲基化的胶质瘤患者对化、放疗更敏感,生存期更长。根据患者的MGMT 基因甲基化状态,制定及实施不同的治疗方案,实现个体化治疗或许可以提高化、放疗疗效和改善预后。因此,本文对MGMT基因甲基化检测在神经胶质瘤治疗中的作用以及目前常用的MGMT基因启动子甲基化检测方法的研究进展进行了综述。     

胶质瘤干细胞与神经干细胞研究进展

多年来，对胶质瘤分子病因研究集中在基因突变体，虽已取得了长足的进步，但已知的突变体多达数十个，究竟是何种突变体，作用于何种细胞，产生何种类型的肿瘤，尚未有足够的实验数据佐证。Holland等研究表明，把某些突变体导人分化成熟或未分化的胶质祖细胞，虽然均可产生肿瘤，但组织类型不尽相同，而且未能得到“癌前期”变和WHO分类中的Ⅰ级胶质瘤，也就是说启动性突变体尚未确定。     

3D培养体系富集外周血中的循环肿瘤干细胞的研究

摘 要：［目的］ 探究建立3D培养体系富集外周血中的循环肿瘤干细胞。［方法］ 应用阴性筛选的方法富集外周血中的循环肿瘤细胞(circulating tumor cells,CTCs),随后利用3D培养体系将富集得到的细胞进行培养。采用免疫荧光、定量RT-PCR对3D培养的前列腺癌LNCaP细胞系、肺癌H2347细胞系的干性表型特征进行分析。使用肺癌患者的血液样本分离CTCs进行3D培养验证该方法的可行性。 ［结果］ 在光镜下可观察到,在3D培养体系中从外周血分离的前列腺癌LNCaP细胞增殖形成肿瘤细胞微球。免疫荧光结果显示,在3D培养条件下成团生长的前列腺癌LNCaP细胞表达EpCAM和CD133。在3D无血清培养条件下,LNCaP细胞系成球率为18.67%±3.06%。前列腺癌LNCaP细胞在3D培养体系中与普通2D培养相比,干性相关基因CD133、CD44和ABCG2表达水平明显增高,3D培养的肺癌H2347细胞与普通2D培养相比,干性相关基因CD133表达水平明显升高。肺腺癌患者肺静脉血分离CTCs 3D培养,可形成多个微型肿瘤细胞微球（称为肺癌类器官）。在光镜下观察,同一个患者培养获得的肺癌类器官呈现不同的形态结构特征。［结论］ 利用3D培养体系能够培养外周血中的CTCs,并富集得到CD133+的肿瘤干细胞。肺癌患者的CTCs 3D培养能够形成肺癌类器官,并能够展现CTCs的异质性。     

环状RNA在神经胶质瘤中的研究进展

环状RNA（circular RNA，circRNA）是1种特殊的内源性非编码RNA，由前体RNA的3′末端与5′末端首尾相连形成闭合环状结构。主要通过可变剪接、转录调控、编码蛋白、抗病毒免疫反应、微小RNA（microRNA，miRNA）海绵作用等发挥作用。circRNA在神经胶质瘤细胞增殖、凋亡、侵袭等方面发挥关键作用。随着基因芯片的问世，circRNA有望成为1种诊断神经胶质瘤的新型生物学标志物，也可为预防、靶向治疗神经胶质瘤提供新的理论依据。     

3D打印技术在肿瘤整形外科中的应用

3D打印技术作为一种新兴临床辅助技术手段,已广泛应用于肿瘤整形外科领域。本文总结了3D 打印技术在肿瘤整形外科中的应用流程 技术要求、应用范围 并结合团队白身临床应用基础 对国内外研室进展进行绕述,同时提出生物 3D打印技术在肿瘤整形外科中的研究思路和应用前景。     

分子影像学在神经胶质瘤的诊断与治疗中的应用

分子影像学是医学影像技术和分子生物学相结合的学科，利用现有的一些医学影像技术，如核医学（PET）、核磁共振（MRI）和光学成像方法（OCT）．对人体内部生理或病理过程在分子水平上进行无损伤的、实时的成像，对神经胶质瘤的诊断和治疗是分子影像学研究的一大方面，全文就分子影像学的原理及技术、分子影像学在胶质瘤诊疗中的应用加以综述。     

神经导航联合荧光素钠在低级别胶质瘤手术中的应用

目的:探讨神经导航联合荧光素钠术中显影辅助在低级别胶质瘤手术中的安全性和有效性。方法:将显微镜下常规手术的21例低级别胶质瘤(low-grade glioma, LGG)患者临床资料归为常规手术组,分别使用5 mg/kg(常规剂量组21例)和1 mg/kg(小剂量组25例)的荧光素钠(fluorescein sodium, FLS)联合神经导航手术的46例LGG患者纳入研究,比较3组手术效果及术后6个月KPS评分。结果:常规剂量组、小剂量组、常规手术组的肿瘤全切除率分别为81.0%(17/21)、76.0%(19/25)、42.9%(9/21),对3组肿瘤切除程度、手术时间、出血量、术后6个月KPS评分进行两两对比,常规剂量组、小剂量组分别与常规手术组比较差异均有显著性(P<0.05),常规剂量组与小剂量组比较差异均无显著性(P>0.05)。3组并发症比较差异均无显著性(P>0.05)。常规剂量组、小剂量组显影率分别为76.2%(16/21)、68.0%(17/25),2组比较差异均无显著性[(χ²=1.665,P=0.645)、(χ^2...     

Extracellular matrix in glioblastoma: opportunities for emerging therapeutic approaches

Extracellular matrix is a complex network of macromolecules that constitute a microenvironment of normal tissues and malignancies such as the primary brain tumor glioblastoma (GBM). The unique composition of the GBM ECM, compared with the brain, contributes to angiogenesis, invasion, and therapeutic resistance of GBM. On the other hand, components of tumor ECM and related aberrant signaling pathways offer opportunities for various therapeutic strategies that are under active investigations. Here we provide a comprehensive overview of emerging therapeutic approaches for GBM that target or utilize its unique ECM via antibodies or ligands, RNA interference, pharmacological agents and modification of ECM molecules. Furthermore, drug-loaded nanoparticles displaying ECM-directed antibodies or peptides enable tumor selective delivery of the payload. As an in vitro research platform, 3D tumor cell culture incorporating ECM can advance our understanding of tumor-ECM interactions.     

卵巢癌肿瘤微环境的三维细胞培养模型研究进展

卵巢癌(Ovarian cancer,OC)是一种具有侵袭性和致命性的癌症。越来越多的研究表明,肿瘤微环境(Tumor microenvironment,TME)通过各种机制参与促进卵巢癌发生、发展、免疫抑制和炎性反应。TME包括肿瘤血管和淋巴管,以及肿瘤细胞、间质细胞、免疫细胞和细胞外基质(ECM)等。本文结合近年来的文献报道,综述了目前常用的卵巢癌TME三维细胞培养模型,以及总结了许多不包含原始基质细胞的三维模型,旨在总结和寻找卵巢癌治疗的新途径和新方法。     

新型肝癌组织的体外三维培养模型

目的:利用肝癌组织进行体外的三维（three-dimension,3D）组织块培养,通过添加PD98059和霍乱毒素（cholera toxin,CT）优化培养条件,建立一种高活性的肝癌组织体外培养模型。方法:收集中山大学肿瘤防治中心肝胆科的肝癌组织标本,经清洗、冻存、复苏等处理后分为对照组、PD组、CT组、PD+CT组,同时进行体外的二维（two-dimension,2D）和三维培养。各组组织采用DNA断裂的原位末端标记法（TUNEL）检测组织细胞凋亡情况,qRT-PCR检测BCL-2、BID、Caspase 3、Cyclin D1、CDK2、ELK-1、C-FOS、C-MYC、C-JUN等相关基因的mRNA表达水平,免疫荧光染色检测BCL-2和cleaved caspase-3蛋白表达情况,筛选并确定最优的体外培养方法。结果:3D培养组的组织细胞凋亡均少于2D培养组;PD98059通过上调BCL-2蛋白的表达,下调cleaved caspase-3蛋白的表达,激活MEK/ERK下游基因,从而减少体外培养的组织细胞凋亡,增强其抗凋亡能力;霍乱毒素通过上调Cyclin D1、CDK2的基因表达,促进其增殖。结论:我们开发了一种新型肝癌组织的体外三维培养模型,它可以作为进一步的药物试验和人源性异种移植(PDX)模型的工具。     

Cancer cell heterogeneity and plasticity: A paradigm shift in glioblastoma

Phenotypic plasticity has emerged as a major contributor to intra-tumoral heterogeneity and treatment resistance in cancer. Increasing evidence shows that glioblastoma (GBM) cells display prominent intrinsic plasticity and reversibly adapt to dynamic microenvironmental conditions. Limited genetic evolution at recurrence further suggests that resistance mechanisms also largely operate at the phenotypic level. Here we review recent literature underpinning the role of GBM plasticity in creating gradients of heterogeneous cells including those that carry cancer stem cell (CSC) properties. A historical perspective from the hierarchical to the nonhierarchical concept of CSCs towards the recent appreciation of GBM plasticity is provided. Cellular states interact dynamically with each other and with the surrounding brain to shape a flexible tumor ecosystem, which enables swift adaptation to external pressure including treatment. We present the key components regulating intra-tumoral phenotypic heterogeneity and the equilibrium of phenotypic states, including genetic, epigenetic, and microenvironmental factors. We further discuss plasticity in the context of intrinsic tumor resistance, where a variable balance between preexisting resistant cells and adaptive persisters leads to reversible adaptation upon treatment. Innovative efforts targeting regulators of plasticity and mechanisms of state transitions towards treatment-resistant states are needed to restrict the adaptive capacities of GBM.     

DNA methylation based glioblastoma subclassification is related to tumoral T-cell infiltration and patient survival

BackgroundHistologically classified glioblastomas (GBM) can have different clinical behavior and response to therapy, for which molecular subclassifications have been proposed. We evaluated the relationship of epigenetic GBM subgroups with immune cell infiltrations, systemic immune changes during radiochemotherapy, and clinical outcome.Methods450K genome-wide DNA methylation was assessed on tumor tissue from 93 patients with newly diagnosed GBM, treated with standard radiochemotherapy and experimental immunotherapy. Tumor infiltration of T cells, myeloid cells, and Programmed cell death protein 1 (PD-1) expression were evaluated. Circulating immune cell populations and selected cytokines were assessed on blood samples taken before and after radiochemotherapy.ResultsForty-two tumors had a mesenchymal, 27 a receptor tyrosine kinase (RTK) II, 17 RTK I, and 7 an isocitrate dehydrogenase (IDH) DNA methylation pattern. Mesenchymal tumors had the highest amount of tumor-infiltrating CD3+ and CD8+ T cells and IDH tumors the lowest. There were no significant differences for CD68+ cells, FoxP3+ cells, and PD-1 expression between groups. Systemically, there was a relative increase of CD8+ T cells and CD8+ PD-1 expression and a relative decrease of CD4+ T cells after radiochemotherapy in all subgroups except IDH tumors. Overall survival was the longest in the IDH group (median 36 mo), intermediate in RTK II tumors (27 mo), and significantly lower in mesenchymal and RTK I groups (15.5 and 16 mo, respectively).ConclusionsMethylation based stratification of GBM is related to T-cell infiltration and survival, with IDH and mesenchymal tumors representing both ends of a spectrum. DNA methylation profiles could be useful in stratifying patients for immunotherapy trials.     

Isolinderalactone inhibits glioblastoma cell supernatant-induced angiogenesis

Glioblastoma multiforme (GBM) is the most frequently occurring malignant brain tumor in adults and is characterized by a high degree of vascularization. Glioblastoma cells communicate with their microenvironment and stimulate blood vessel formation to support tumor progression. It has previously been reported that isolinderalactone induces apoptosis in GBM cells and suppresses the growth of glioblastoma xenograft tumors in vivo. Furthermore, isolinderalactone has been shown to inhibit the hypoxia-driven upregulation of vascular endothelial growth factor (VEGF) in U-87 GBM cells and strongly reduce VEGF-triggered angiogenesis in vitro and in vivo. In the present study, the direct angiogenic effect of GBM and the effect of isolinderalactone on tumor angiogenesis were investigated. Culture supernatants were obtained from U-87 cells under normoxic or hypoxic conditions to provide normoxic conditioned medium (NCM) and hypoxic conditioned medium (HCM) respectively. The NCM and HCM were each used to treat to human brain microvascular endothelial cells (HBMECs), and their effects were observed using wounding migration and tube formation assays. HCM increased the migration and capillary-like tube formation of HBMECs when compared with NCM, and treatment with isolinderalactone suppressed the HCM-driven angiogenesis in vitro. Additionally, isolinderalactone decreased HCM-triggered angiogenic sprouting in HBMECs in a 3D microfluidic device after the application of an HCM-containing interstitial fluid flow. Furthermore, isolinderalactone strongly reduced HCM-triggered angiogenesis in an in vivo Matrigel plug assay in mice. These findings provide evidence of angiogenesis inhibition by isolinderalactone, and demonstrate the anti-angiogenic effect of isolinderalactone against the direct angiogenic effect of GBM tumor cells.     

基于Oncomine数据库及GEPIA数据库分析CHI3L2基因在胶质母细胞瘤中的表达及与预后的相关性

目的:基于Oncomine数据库及GEPIA数据库分析CHI3L2基因在胶质母细胞瘤中的表达及与预后的相关性。方法:检索Oncomine和GEPIA数据库中相关胶质母细胞瘤的数据集,分析胶质母细胞瘤组织与正常对照组织之间基因表达的差异,采用GEPIA数据库进行在线生存分析。结果:Oncomine数据库中共收集了449项关于CHI3L2基因在肿瘤与正常组织中表达比较的研究结果,表达差异有统计学意义的研究结果有23项,其中有9项研究结果呈高表达,14项研究结果呈低表达。与对照组相比,在胶质母细胞瘤组织中CHI3L2的表达显著高于正常组织（P＜0.05）。结论:CHI3L2在胶质母细胞瘤组织中呈高表达,且与胶质母细胞瘤患者预后相关,为临床胶质母细胞瘤的治疗及基因靶向药物的研制提供重要理论依据。     

Interactions between cancer cells and immune cells drive transitions to mesenchymal-like states in glioblastoma

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