5-Azacytidine and MS-275 in Myeloid Malignancies

5-氮杂胞苷和 MS-275 在骨髓恶性肿瘤中的应用

• 批准号: 6836977
• 负责人: STEVEN D GORE
• 金额: $ 28.46万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-15 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6836977
• 关键词: DNA methylation; acetylation; acute myelogenous leukemia; amidohydrolases; antineoplastics; azacitidine; chromatin immunoprecipitation; clinical research; clinical trial phase I; clinical trial phase II; combination chemotherapy; dosage; enzyme inhibitors; gene induction /repression; human subject; methyltransferase; microarray technology; myelogenous leukemia; neoplasm /cancer chemotherapy; neoplasm /cancer pharmacology; nucleic acid sequence; oral administration; patient oriented research; subcutaneous drug administration; western blottings

DESCRIPTION (provided by applicant): 5-azacytidine (5AC) is a highly active agent for the treatment of myelodysplastic syndromes (MDS, response rate 50 - 60%). 5AC retards the progression of MDS to acute myeloid leukemia (AML); however, the median duration of response to 5AC is less than 18 months. The mechanism of clinical activity of this agent may derive from its potent inhibition of DNA methyltransferase (DNMT). Exposure of cells to DNMT inhibitors (DNMTi) leads to reversal of methylation of CpG islands in the promoter regions of transcriptionally silenced genes and can lead to re-expression of such genes. More robust re-expression of methylated genes may be accomplished by the combination of DNMTi with histone deacetylase inhibitors (HDACi). MS-275, an orally bioavailable HDACi, induces histone hyperacetylation sustained up to three weeks following drug administration. This new reagent provides an outstanding opportunity to test the concept of combined DNMTi and HDACi in myeloid malignancies. The clinical trial forming the basis of this Quick Trials application will identify an optimal doses and schedule of subcutaneous 5AC and oral MS-275. The application requests funds to perform laboratory studies aimed at testing the hypothesis that combinations of 5AC with MS-275 lead to re-expression of silenced genes, and that such re-expression is associated with clinical response. Patients will be treated with a variety of doses and schedules of 5AC and MS-275 (administered on days 3 and 10 of each treatment cycle). The data will be evaluated to identify a "clinically optimal dose" which maximizes histone hyperacetylation and gene re-expression with minimal toxicity. Quantitative rt-PCR will be used to monitor the expression of two genes which are most commonly silenced through methylation in AML and MDS: p15INK4B and Ecadherin (E-CAD). Expression microarrays will be performed on isolated CD34+ cells to identify genes and molecular pathways whose expression is up- or down-regulated in response to this combination of drugs which may be related to response (whether methylated or not). Patient samples will be studied for changes in promoter methylation using realtime PCR modifications of methylation-specific PCR, and bisulfite sequencing of PCR products. Chromatin immunoprecipitation assays will identify changes in acetylation of histones associated with promoter sequences in p15 and p21WAF1/CIP1. Finally, Western analysis will be used to seek changes in phosphorylated H2AX, a marker of DNA damage, to evaluate the extent to which these drugs induce DNA strand breaks. Promising preliminary data from this trial will be followed directly by a randomized Phase II trial of 5AC plus MS-275 versus the same schedule of 5AC alone to investigate the clinical importance of the addition of the HDAC inhibitor.

描述（由申请人提供）：5-氮杂丁胺（5AC）是治疗骨髓增生综合征的高度活跃药物（MDS，响应率50-60％）。 5AC降低了MDS向急性髓样白血病（AML）的进展；但是，对5AC的响应持续时间少于18个月。该药物的临床活性机制可能来自其对DNA甲基转移酶（DNMT）的有效抑制。细胞暴露于DNMT抑制剂（DNMTI）会导致转录沉默基因启动子区域中CpG岛的甲基化逆转，并可能导致此类基因的重新表达。通过DNMTI与组蛋白脱乙酰基酶抑制剂（HDACI）的组合，可以实现甲基化基因的更强大的重新表达。 MS-275是一种口服可生物可用的HDACI，在药物给药后长达三周诱导组蛋白高乙酰化。这项新试剂为测试髓样恶性肿瘤中DNMTI和HDACI组合的概念提供了绝佳的机会。构成此快速试验申请的基础的临床试验将确定皮下5AC和口服MS-275的最佳剂量和时间表。该申请要求资金进行实验室研究，旨在测试5AC与MS-275的组合导致沉默基因的表达，并且这种重新表达与临床反应有关。患者将用5AC和MS-275的多种剂量和时间表进行治疗（在每个治疗周期的第3和10天进行）。将评估数据以确定“临床最佳剂量”，该剂量最大化组蛋白高乙酰化和基因以最小的毒性重新表达基因。定量RT-PCR将用于监测两个基因的表达，这些基因通常通过AML和MDS中的甲基化最常见：P15INK4B和Ecadherin（E-CAD）。表达微阵列将在分离的CD34+细胞上进行，以鉴定基因和分子途径，其表达响应于这种可能与反应有关的药物的组合响应（无论是否甲基化）。将研究使用甲基化特异性PCR的实时PCR修饰和PCR产物的亚硫酸盐测序的实时PCR修饰启动子甲基化的变化。染色质免疫沉淀分析将确定与p15和p21WAF1/CIP1中启动子序列相关的组蛋白乙酰化的变化。最后，西方分析将用于寻求DNA损伤标记的磷酸化H2AX的变化，以评估这些药物诱导DNA链断裂的程度。该试验中有希望的初步数据将直接进行5AC加上MS-275的随机II期试验，而仅相同的5AC时间表来研究添加HDAC抑制剂的临床重要性。