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Study of telomerase as a useful target for diagnosis and therapy of neuroblastoma with the aim of clinical application

研究端粒酶作为神经母细胞瘤诊断和治疗有用靶点的临床应用

基本信息

批准号：
11470368
负责人：
HIYAMA Eiso
金额：
$ 9.47万
依托单位：
HIROSHIMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11470368/
关键词：
neuroblastoma telomerase hTERT hTR prognosis ribozyme MYCN antisense RNA antisense RNA テメロメラーゼテロメア予後

项目摘要

We examined telomere length, telomerase activity, and the expression levels of several human telomerase components such as RNA component(hTR), human telomerase reverse transcriptase (hTERT), and human telomerase associated protein 1 (hTEP1) in 241 frozen neuroblastoma tissues by the TRAP assay and RT-PCR using fluorescent thermal cycler. Unfavorable neurobalstomas were distinguishable from favorable ones by the levels of telomerase activity and hTERT expression. Some tumors with low or undetectable telomerase activity showed alternatively spliced fragments of hTERT.While the amount of the full-length fragment was in relation to the telomerase activity in general, the sensitivity and the specificity of quantitative RT-PCR in consideration of alternative splicing were not satisfactory.Then we tried in situ hybridization of hTR and immunohistochemical detection of hTERT expression using anti-hTERT antibody. By these methods, in favorable tumors, these expression levels were low and some t … More umor cells lacked these signals. On the other hand, in the tumors with high telomerase activity, almost all tumors cells exhibits strong signals of hTR and hTERT.In the tumors with mnoderate levels of telomerase activity, small amounts of tumor cells showed high expressions of hTR and hTERT.Among 12 tumors with high telomerase and high hTERT expressions, shortened telomere was detected in 10, which were considered as adequate cases for antitelomerase therapy. Then we tried risozyme treatment for hTR, antisense hTERT RNA treatment, and anti hTERT antibody treatment in the cultured neuroblastoma cells with high telomerase activity. In these antitelomerase treatments brought the growth inhibition and the induction of apoptosis, suggesting the correlation between telomerase reduciton without telomerase activity and apoptosis.These results suggested that the quatitative and in situ analyses of human telomerase are useful markers for evaluation of malignant grades of neuroblastoma and for indication of antitelomerase therapies. Less

采用TRAP法和RT-PCR技术检测了241例神经母细胞瘤冰冻组织的端粒长度、端粒酶活性以及端粒酶RNA组分（hTR）、端粒酶逆转录酶（hTERT）和端粒酶相关蛋白1（hTEP 1）的表达水平。不良的神经母细胞瘤可以通过端粒酶活性和hTERT表达水平来区分。在端粒酶活性较低或检测不到的肿瘤中发现hTERT的可变剪接片段，而全长片段的多少一般与端粒酶活性有关，但考虑可变剪接的定量RT-PCR的敏感性和特异性均不理想，于是我们尝试了hTR的原位杂交和hTERT抗体的免疫组化检测。通过这些方法，在良好的肿瘤中，这些表达水平很低，一些肿瘤细胞的表达水平很低。 ...更多信息肿瘤细胞缺乏这些信号。在端粒酶活性高的肿瘤中，几乎所有的肿瘤细胞都有较强的hTR和hTERT信号，在端粒酶活性中等的肿瘤中，有少量的肿瘤细胞hTR和hTERT高表达，在12例端粒酶和hTERT高表达的肿瘤中，有10例检测到端粒缩短，这些肿瘤都可以进行抗端粒酶治疗。在培养的端粒酶活性高的神经母细胞瘤细胞中，我们尝试了risozyme处理hTR，反义hTERTRNA处理，以及抗hTERT抗体处理。这些抗端粒酶治疗均能抑制神经母细胞瘤的生长并诱导细胞凋亡，提示端粒酶活性降低与细胞凋亡之间存在相关性，提示端粒酶定量和原位分析可作为神经母细胞瘤恶性程度的评价指标和抗端粒酶治疗的参考指标。少