Common hTERT Promoter Mutations Represent a Novel Therapeutic Target

 描述（由申请人提供）：端粒酶逆转录酶（hTERT）是端粒酶的催化亚单位，其经常在人类肿瘤中过表达。最近已经表明，hTERT启动子通常在恶性黑素瘤和胶质母细胞瘤（>75%的肿瘤）中突变。这些突变发生在富含G的区域中的四个特定位点，该区域已被证明形成四链体DNA并下调hTERT表达。我们已经表明，这些突变不稳定的四链体形成导致增加的hTERT表达和细胞增殖。用TMPYP4稳定四链体结构逆转了这些变化。用“链侵入”寡核苷酸处理其中hTERT启动子突变的细胞下调hTERT表达并显著抑制转化细胞的生长。本申请将检验这样的假设，即通过序列特异性寡核苷酸稳定突变的hTERT启动子四链体结构将导致hTERT的下调和具有突变序列的细胞的生长抑制。本申请的具体目的是：1.表征靶向突变或野生型hTERT启动子序列的寡核苷酸的基因和序列特异性。2.目的：探讨hTERT启动子靶向寡核苷酸与靶向药物和标准化疗药物的协同作用。3.表征hTERT链侵入寡核苷酸在动物模型中单独使用和与靶向治疗联合使用的治疗潜力。由于hTERT基因表达的抑制与多种肿瘤类型相关，因此拟议研究的结果将具有重要的临床意义。此外，拟议的实验将提供重要的新信息的作用，四链体形成的hTERT基因的表达调控。

 DESCRIPTION (provided by applicant): Telomerase reverse transcriptase (hTERT) is a catalytic subunit of the enzyme telomerase which is frequently overexpressed in human tumors. It has recently been shown that the hTERT promoter is commonly mutated in a malignant melanoma and glioblastoma (>75% of tumors). These mutations occur at four specific sites in a G-rich region which has been shown to form quadruplex DNA and to downregulate hTERT expression. We have shown that these mutations destabilize quadruplex-formation resulting in increased hTERT expression and cellular proliferation. Stabilization of the quadruplex structure with TMPYP4 reverses these changes. Treatment of cells in which the hTERT promoter is mutated with "strand-invading" oligonucleotides downregulates hTERT expression and dramatically inhibits the growth of transformed cells. This application will test the hypothesis tht stabilization of the mutated hTERT promoter quadruplex structure by sequence specific oligonucleotides will result in downregulation of hTERT and inhibition of growth by cells with the mutated sequence(s). The Specific Aims of this application are: 1. To characterize the gene and sequence specificity of oligonucleotides targeted to the mutated or wild type hTERT promoter sequences. 2. To characterize the potential synergy of strand invading hTERT promoter targeted oligonucleotides with targeted agents and standard chemotherapy drugs. 3. To characterize the therapeutic potential of hTERT strand invading oligonucleotides in animal models, alone and in conjunction with targeted therapies. The results of the proposed studies will have important clinical implications as inhibition of hTERT gene expression is relevant to a wide variety of tumor types. In addition, the proposed experiments will provide important new information about the role of quadruplex-formation in the regulation of hTERT gene expression.