Sphingosine-1-Phosphate Pathway Based Therapy for Neuroblastoma

1-磷酸鞘氨醇途径治疗神经母细胞瘤

• 批准号: 8345152
• 负责人: FERNANDO A FERRER
• 金额: $ 35.89万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-09 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8345152
• 关键词: 14-3-3 Proteins; Accounting; Angiogenic Factor; Apoptosis; Apoptotic; Award; Binding; Biological Availability; Biological Models; CCL2 gene; Caspase; Cell Death; Cell Line; Cell Survival; Cell physiology; Cells; Ceramides; Cessation of life; Chemotactic Factors; Child; Childhood Solid Neoplasm; Clinic; Collaborations; Combined Modality Therapy; Cytotoxic agent; Development; Disease; Drug usage; Equilibrium; Evaluation; Future; Generations; Genetic; Genetic Models; Goals; Grant; H218 Protein; Half-Life; Immunosuppressive Agents; Individual; Laboratories; Lipids; MYCN gene; Malignant Childhood Neoplasm; Malignant Neoplasms; Modeling; Mus; Neuroblastoma; Pathway interactions; Process; Property; Receptor Inhibition; Regulation; Reliance; Signal Transduction; Solid Neoplasm; Specificity; Sphingosine; Sphingosine-1-Phosphate Receptor; Stromal Neoplasm; Testing; Therapeutic Effect; Topoisomerase Inhibitors; Transgenic Model; Transgenic Organisms; Translating; Treatment Efficacy; Vascular Endothelial Growth Factors; Weight; Work; Xenograft Model; Xenograft procedure; analog; angiogenesis; base; cancer therapy; cell growth; chemotherapy; computerized data processing; cytotoxicity; design; drug development; efficacy evaluation; extracellular; high risk; improved; in vivo; information gathering; inhibitor/antagonist; knock-down; macrophage; neoplastic cell; neuroblastoma cell; novel; novel strategies; prevent; research study; small hairpin RNA; sphingosine 1-phosphate; sphingosine kinase; therapeutic target; tumor; tumor progression; tumorigenic

DESCRIPTION (provided by applicant): Neuroblastoma (NB) accounts for 15% of all childhood cancer deaths. Survival for children with high-risk disease remains below 40% and survival curves have plateaued despite treatment intensification strategies. As progress has stagnated, novel treatment approaches are clearly needed. Our laboratory has recently discovered that in NB two processes central to tumor progression are regulated by S1P signaling and can be targeted for therapeutic benefit. The first process concerns cell fate. Inside cells, the delicate balance between pro-survival (S1P) and apoptosis (sphingosine, ceramide) signaling is dictated by sphingosine kinases (SphK1 and SphK2). Neuroblastoma is unique among solid tumors we have evaluated in its disproportionate reliance on SphK2, which we strategically exploit, weighting the pendulum towards cell death. We have accomplished this by utilizing the sphingosine analogue FTY720, which is avidly and specifically phosphorylated by SphK2, thereby acting as a competitive inhibitor increasing pro-apoptotic sphingosine levels, while diminishing S1P levels. Secondly, we have found that extracellular S1P modulates tumor-stromal interactions. The S1P receptor, S1P2, contributes to the regulation of the tumor microenvironment in NB through expression of the potent macrophage chemoattractant MCP-1/CCL2, as well as, angiogenic factor VEGF. As we show, selective interference with this process disrupts establishment of a milieu favorable for tumor progression. Using this novel 2-pronged approach we demonstrate efficacy and synergy with current chemotherapy. Thus, our overarching hypothesis is that rational exploitation/inhibition of S1P signaling mechanisms will synergize with cytotoxic agents and result in enhanced treatment efficacy. This hypothesis is explored in 3 aims in this proposal. Aim 1: Evaluates the efficacy of FTY720 based NB tumor inhibition- In this aim we explore the hypothesis that FTY720 based interference with over-expressed SphK2 in NB cells results in enhanced apoptotic signaling through caspase independent pathways, favoring tumor cell death. These studies will evaluate FTY720 activity, alone and in combination with topoisomerse inhibitors across selected NB cell lines, xenografts and in the MYCN transgenic model. The mechanisms involved in FTY720 related cell death will also be defined as this information may aid in future drug development in aim 3. Aim 2: Utility of S1P2 interference/inhibition strategies for treating neuroblastoma-Through induction of MCP-1/CLL2 and VEGF, S1P2 regulates macrophage invasion and angiogenesis, respectively. We will test the therapeutic effect of S1P2 receptor inhibition alone (JTE-013) and in combination therapy against NB cell lines, xenografts and the MYCN transgenic model. To determine if S1P2 is critical to tumor development and progression we will use shRNA to knock down S1P2 in NB cell lines and xenograft models. Further, we will cross transgenic MYCN mice with S1p2-/- mice available in our lab, assessing the significance of S1P2 using a genetic model. Aim 3: Development and testing of FTY720 and JTE013 analogs with improved anti-cancer properties- The first generation drugs used in aims 1 and 2 have limitations and can be associated with off-target effects. In this final aim we will test analogs of FTY720, which are designed not to bind to the S1P receptors, to reduce immunosuppressive properties and other off target effects, yet are able to target intracellular SphK2 inhibition and thereby achieve tumor cytotoxicity. In the case of JTE-013, longer in-vivo half-life and enhanced inhibitory activity are focuses for 2nd generation compounds, several compounds designed with these endpoints are evaluated using our established models. The final aim will be performed in collaboration with Dr. Rolf Swenson of Arroyo Biosciences. PUBLIC HEALTH RELEVANCE: Present proposal explores a new approach to Neuroblastoma (NB) treatment, the disruption of essential S1P signaling processes. Sphingosine 1-phosphate (S1P) is a bioactive lipid that regulates many aspects of tumor cell growth and function. Findings from this proposal could rapidly translate into new compounds and subsequently into the clinic, which is our ultimate goal.

描述（由申请人提供）：神经母细胞瘤 (NB) 占所有儿童癌症死亡人数的 15%。尽管采取了强化治疗策略，患有高危疾病的儿童的生存率仍低于 40%，并且生存曲线已趋于稳定。由于进展停滞，显然需要新的治疗方法。我们的实验室最近发现，在 NB 中，肿瘤进展的两个核心过程受到 S1P 信号传导的调节，并且可以作为治疗益处的目标。第一个过程涉及细胞命运。里面 在细胞中，促生存 (S1P) 和凋亡（鞘氨醇、神经酰胺）信号传导之间的微妙平衡由鞘氨醇激酶（SphK1 和 SphK2）决定。神经母细胞瘤在我们评估的实体瘤中是独一无二的，因为它对 SphK2 的过度依赖，我们战略性地利用它，使钟摆向细胞死亡倾斜。我们通过利用鞘氨醇类似物 FTY720 实现了这一目标，FTY720 被 SphK2 强烈且特异性地磷酸化，从而充当竞争性抑制剂，增加促凋亡鞘氨醇水平，同时降低 S1P 水平。其次，我们发现细胞外 S1P 调节肿瘤-基质相互作用。 S1P 受体 S1P2 通过表达强效巨噬细胞趋化剂 MCP-1/CCL2 以及血管生成因子 VEGF，有助于调节 NB 中的肿瘤微环境。正如我们所表明的，选择性干扰这一过程会破坏有利于肿瘤进展的环境的建立。使用这种新颖的双管齐下的方法，我们证明了与当前化疗的功效和协同作用。因此，我们的总体假设是，合理利用/抑制 S1P 信号传导机制将与细胞毒性药物产生协同作用，从而提高治疗效果。本提案通过 3 个目标探讨了这一假设。目标 1：评估基于 FTY720 的 NB 肿瘤抑制功效 - 在此目标中，我们探讨了这样的假设：基于 FTY720 的 NB 细胞中过表达 SphK2 的干扰导致通过 caspase 独立途径增强细胞凋亡信号传导，有利于肿瘤细胞死亡。这些研究将在选定的 NB 细胞系、异种移植物和 MYCN 转基因模型中单独评估 FTY720 活性，以及​​与拓扑异构体抑制剂联合使用的 FTY720 活性。 FTY720 相关细胞死亡涉及的机制也将被定义，因为该信息可能有助于未来目标 3 的药物开发。目标 2：利用 S1P2 干扰/抑制策略治疗神经母细胞瘤 - 通过诱导 MCP-1/CLL2 和 VEGF， S1P2 分别调节巨噬细胞侵袭和血管生成。我们将测试单独的 S1P2 受体抑制 (JTE-013) 和联合治疗对 NB 细胞系、异种移植物和 MYCN 转基因模型的治疗效果。为了确定 S1P2 是否对肿瘤的发生和进展至关重要，我们将使用 shRNA 敲低 NB 细胞系和异种移植模型中的 S1P2。此外，我们将转基因 MYCN 小鼠与我们实验室现有的 S1p2-/- 小鼠杂交，使用遗传模型评估 S1P2 的重要性。目标 3：开发和测试具有改善抗癌特性的 FTY720 和 JTE013 类似物 - 目标 1 和 2 中使用的第一代药物具有局限性，并且可能与脱靶效应相关。在这个最终目标中，我们将测试 FTY720 的类似物，其设计不结合 S1P 受体，以减少免疫抑制特性和其他脱靶效应，但能够靶向细胞内 SphK2 抑制，从而实现肿瘤细胞毒性。就 JTE-013 而言，第二代化合物的重点是更长的体内半衰期和增强的抑制活性，使用我们建立的模型评估了根据这些终点设计的几种化合物。最终目标将与 Arroyo Biosciences 的 Rolf Swenson 博士合作完成。 公共健康相关性：本提案探索了神经母细胞瘤 (NB) 治疗的新方法，即破坏重要的 S1P 信号传导过程。 1-磷酸鞘氨醇 (S1P) 是一种生物活性脂质，可调节肿瘤细胞生长和功能的许多方面。该提案的研究结果可以迅速转化为新的化合物，并随后进入临床，这是我们的最终目标。