RETINOID THERAPY OF NEUROBLASTOMA

神经母细胞瘤的视黄醇治疗

• 批准号: 6038551
• 负责人: CHARLES Patrick REYNOLDS
• 金额: $ 3.51万
• 依托单位: CHILDREN'S HOSPITAL OF LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-01-19 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6038551
• 关键词: 13 cis retinoate; SCID mouse; alkylating agents; antineoplastics; athymic mouse; cell line; cellular oncology; ceramides; cytotoxicity; drug resistance; drug screening /evaluation; fenretinide; fibroblasts; free radical oxygen; human tissue; myoblasts; neuroblastoma; transfection

Our pre-clinical studies showed that 13-cis-retinoic acid (13-cis-RA) can inhibit cell growth and decrease expression of the MYCN oncogene in neuroblastoma (NBL) cells in vitro. These data led us to conduct a phase I and then a randomized phase III trial which showed that high-dose, pulse 13-cis-RA significantly improved survival for high-risk NBL when given after intensive chemotherapy. Not all patients benefited from 13-cis-RA, and we found that some NBL cell lines derived from patients who were clinically resistant to 13-cis-RA are resistant to 13-cis-RA and trans-RA in vitro. We discovered that the RA derivative N-(4-hydroxyphenyl) retinamide (fenretinide; 4-HPR) is highly cytotoxic for NBL cell lines that are resistant to 13-cis-RA. The hypothesis of this proposal is that 4-HPR will further improve the survival of high-risk NBL patients by eradicating tumor cells, including those cells that are resistant to 13-cis-RA. This project will explore the molecular mechanisms by which 4-HPR selectively induces tumor cell death. We showed that 4-HPR caused large increases in the levels of ceramide (a lipid involved in apoptotic signaling) in NBL cell lines and that (like exogenous ceramide) 4-HPR cytotoxicity involved both apoptosis and necrosis. We will continue this work by determining if ceramide synthetic enzymes are induced by 4-HPR or if ceramide metabolic enzymes are down-regulated. To determine if cytotoxicity mediated by high-dose 4-HPR is due to increased ceramide we will transfect a NBL cell line with an inducible vector containing glucosylceramide synthase, which will shunt ceramide in 4-HPR treated cells to a non-toxic form. Cell lines will be identified and developed that are resistant to 4-HPR to determine if 4-HPR-resistance is mediated by a failure to increase ceramide in response to 4-HPR or by "shunting" the ceramide to non-toxic forms. Agents that inhibit ceramide shunting will be evaluated for their ability to enhance the cytotoxicity of 4-HPR for NBL cell lines. Combinations of agents with 4-HPR that have enhanced cytotoxicity against NBL cell lines in vitro will be tested for general toxicity against fibroblasts and myeloid progenitors. and for toxicity and efficacy against NBL xenografts. 4-HPR and enhancing agents will be evaluated for their efficacy against NBL cell lines that are resistant to 13-cis-RA and chemotherapeutic drugs. This project, in combination with our ongoing phase I trial of 4-HPR, will provide data to optimize clinical trial design employing 4-HPR against NBL. We anticipate that data generated will support developing a future clinical trial testing the ability of 4-HPR to improve outcome for high risk NBL when given after myeloablative therapy and 13-cis-RA. Elucidating mechanisms of action for 4-HPR should allow us to identify clinically useful agents that will enhance the anti-cancer activity 4-HPR for testing in future clinical trials.

我们的临床前研究表明，13-顺式维甲酸（13-顺式ra）可以抑制体外神经母细胞瘤（NBL）细胞的生长并降低MYCN癌基因的表达。这些数据使我们进行了一期和随后的随机三期试验，该试验表明，在强化化疗后给予高剂量，脉冲13-顺式ra可显著提高高风险NBL患者的生存率。并非所有患者都受益于13-顺式ra，我们发现一些来自临床对13-顺式ra耐药的患者的NBL细胞系在体外对13-顺式ra和反式ra耐药。我们发现RA衍生物N-（4-羟基苯基）维甲酸（fenretinide; 4-HPR）对耐13-顺式RA的NBL细胞系具有高度的细胞毒性。本提案的假设是，4-HPR将通过根除肿瘤细胞，包括对13-顺- ra耐药的细胞，进一步提高高危NBL患者的生存率。本项目将探索4-HPR选择性诱导肿瘤细胞死亡的分子机制。我们发现，4-HPR引起NBL细胞系中神经酰胺（一种参与凋亡信号传导的脂质）水平的大量增加，并且（像外源性神经酰胺一样）4-HPR细胞毒性涉及细胞凋亡和坏死。我们将继续这项工作，确定神经酰胺合成酶是否被4-HPR诱导，或者神经酰胺代谢酶是否被下调。为了确定高剂量4-HPR介导的细胞毒性是否是由于神经酰胺增加，我们将用含有葡萄糖神经酰胺合成酶的诱导载体转染NBL细胞系，该载体将4-HPR处理细胞中的神经酰胺转移到无毒形式。将鉴定和开发抗4-HPR的细胞系，以确定4-HPR抗性是由神经酰胺在4-HPR反应中未能增加还是通过将神经酰胺“分流”到无毒形式介导的。抑制神经酰胺分流的药物将被评估其增强4-HPR对NBL细胞系的细胞毒性的能力。对NBL细胞系体外增强细胞毒性的4-HPR药物组合将测试对成纤维细胞和髓系祖细胞的一般毒性。以及对NBL异种移植物的毒性和有效性。将评估4-HPR和增强剂对耐13-顺- ra和化疗药物的NBL细胞系的疗效。该项目与我们正在进行的4-HPR I期试验相结合，将为优化4-HPR治疗NBL的临床试验设计提供数据。我们预计，所产生的数据将支持开发未来的临床试验，测试4-HPR在清髓治疗和13-cis-RA后改善高风险NBL预后的能力。阐明4-HPR的作用机制将使我们能够确定临床有用的药物，以增强4-HPR的抗癌活性，以便在未来的临床试验中进行测试。