HLTF gene silencing: a novel determinant of sensitivity to autophagy inhibition

HLTF 基因沉默：自噬抑制敏感性的新决定因素

• 批准号: 8664818
• 负责人: RAVI K AMARAVADI
• 金额: $ 33.24万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8664818
• 关键词: Advanced Malignant Neoplasm; Antineoplastic Agents; Autophagocytosis; Biological Assay; Biological Markers; Blood; Cancer Patient; Cancer cell line; Cell Line; Cells; Cessation of life; Characteristics; Chloroquine; Clinical; Clinical Treatment; Clinical Trials; Colon Carcinoma; DNA Damage; DNA Methylation; DNA Repair; Data; Development; Disease; Enrollment; Frequencies; Future; Gene Silencing; Genes; Genome; Genomics; Goals; Human; Hydroxychloroquine; Institution; Knowledge; Light; Link; Lysosomes; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of pancreas; Measures; Mediating; Metabolic; Methylation; Microarray Analysis; Molecular; Outcome; Pathway interactions; Patients; Performance; Prevalence; Process; Prognostic Marker; Reactive Oxygen Species; Refractory; Regimen; Research Personnel; Resistance; SMARCA3 gene; Sampling; Serum; Specific qualifier value; Stable Disease; Stomach Carcinoma; Stress; Testing; Therapeutic; Tissue Microarray; Tumor Suppressor Genes; Tumor Tissue; Validation; Work; assay development; base; cancer cell; cancer therapy; cohort; colon cancer cell line; demethylation; helicase; improved; inhibition of autophagy; inhibitor/antagonist; lung Carcinoma; mRNA Expression; malignant breast neoplasm; malignant stomach neoplasm; melanoma; new therapeutic target; novel; overexpression; oxidative DNA damage; pre-clinical; promoter; public health relevance; repaired; response; transcription factor; tumor

DESCRIPTION (provided by applicant): Autophagy is a lysosome-dependent degradative process that protects cancer cells from metabolic and therapeutic stress. Autophagy is up regulated in most advanced cancers and has identified as a new target for cancer therapy. Autophagy inhibition with chloroquine (CQ) derivatives augments the efficacy of many anticancer therapies. Numerous clinical trials are testing the combination of variety of anticancer agents with hydroxychloroquine (HCQ), but there currently is no biomarker that can focus the development of HCQ combinations or regimens involving novel emerging autophagy inhibitors into patient subsets that will most likely benefit from this strategy. To identify a candidate biomarker of sensitivity to autophagy inhibition, a microarray analysis of differentially expressed genes in HCQ-sensitive and HCQ-resistant human cancer cell lines was conducted. The most down regulated gene in HCQ-sensitive cells was helicase- like transcription factor (HLTF), an understudied tumor suppressor gene involved in multiple aspects of maintaining genomic integrity during replication stress. HLTF expression is silenced by promoter methylation in 20-40% of lung, colon, and gastric carcinomas. In a large panel of cell lines HLTF gene silencing was found almost exclusively in cell lines that were sensitive to HCQ. Forced expression of HLTF in HLTF silenced cells conferred resistance to HCQ. The link between autophagy inhibition and HLTF may be through oxidative DNA damage that was observed soon after HCQ treatment. A methylation-specific PCR assay was able to detect HLTF methylation status of tumors in the serum samples of patients with melanoma and breast cancer indicating that HLTF gene silencing is common across multiple malignancies and a serum assay may be able to classify patients as HLTF gene silenced or expressed. This proposal will test the hypotheses that a) HLTF gene silencing confers sensitivity to autophagy inhibitors by allowing oxidative DNA damage to go unrepaired~ b) A clinical grade assay for HLTF promoter methylation in the serum or tumors of patients can be a sensitive and specific assay for HLTF gene silencing c) HLTF gene silencing is common in a number of malignancies and predicts of clinical response in patients treated with HCQ. Knowledge gained from completion of these specific aims will establish a new mechanistic framework that links autophagy inhibition to the DNA damage response. This work will shed light on the functions of commonly silenced tumor suppressor gene HLTF and will determine which malignancies should be a target for the development of autophagy inhibitors. Finally, as novel autophagy inhibitors are currently being developed for clinical trials in cancer patients, completion of these aims will provide the supporting data necessary for future development of a CLIA approved predictive assay that could be used to enroll the patients most likely to benefit from autophagy inhibitors.

描述（由申请人提供）：自噬是一种溶酶体依赖性的降解过程，可保护癌细胞免受代谢和治疗应激。自噬在大多数高级癌症中受到调节，并已确定为癌症治疗的新靶标。用氯喹（CQ）衍生物抑制自噬抑制增强了许多抗癌疗法的功效。许多临床试验正在测试各种抗癌的组合 羟基氯喹（HCQ）的试剂，但目前没有生物标志物可以集中精力的HCQ组合或方案，涉及新兴的自噬抑制剂到患者子集中，这很可能会从这种策略中受益。 为了确定对自噬抑制敏感性的候选生物标志物，对差异表达的微阵列分析 进行了抗HCQ敏感和耐HCQ的人类癌细胞系中的基因。 HCQ敏感细胞中最下调的基因是解旋酶类转录因子（HLTF），这是一种研究了在复制应激过程中维持基因组完整性的多个方面的研究研究的肿瘤抑制基因。 HLTF表达被20-40％的肺，结肠和胃癌中的启动子甲基化沉默。 在一系列细胞系中，HLTF基因沉默几乎完全在对HCQ敏感的细胞系中。 HLTF在HLTF中的强迫表达沉默的细胞赋予了对HCQ的抗性。自噬抑制与HLTF之间的联系可能是通过HCQ治疗后不久观察到的氧化DNA损伤。甲基化特异性PCR分析能够检测到黑色素瘤和乳腺癌患者血清样品中肿瘤的HLTF甲基化状态，表明HLTF基因沉默在多种恶性肿瘤中很常见，并且血清测定可能能够将患者分类为HLTF基因沉默或表达。 This proposal will test the hypotheses that a) HLTF gene silencing confers sensitivity to autophagy inhibitors by allowing oxidative DNA damage to go unrepaired~ b) A clinical grade assay for HLTF promoter methylation in the serum or tumors of patients can be a sensitive and specific assay for HLTF gene silencing c) HLTF gene silencing is common in a number of通过HCQ治疗的患者的恶性肿瘤和预测临床反应。 这些特定目标从完成后获得的知识将建立一个新的机械框架，该框架将自噬抑制与DNA损伤响应联系起来。这项工作将阐明通常沉默的肿瘤抑制基因HLTF的功能，并将确定哪些恶性肿瘤应成为自噬抑制剂发展的靶标。 最后，由于目前正在开发用于癌症患者的临床试验的新型自噬抑制剂，因此这些目标的完成将为将来开发CLIA认可的预测性测定所必需的支持数据，该数据可用于招募最有可能受益于自噬抑制剂的患者。