Experimental research for the availability of gene therapy using ORP150, a novel molecular chaperone

使用新型分子伴侣 ORP150 进行基因治疗可用性的实验研究

• 批准号: 12671522
• 负责人: OGAWA Satoshi
• 金额: $ 2.05万
• 依托单位: GRADUATE SCHOOL OF MEDICINE, KANAZAWA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671522/
• 关键词: Stress response; Unfolded protein response; angiogenesis; Molecular chaperone; Gene therapy; Adenovirus vecto; Tumor growth; 癌遺伝子治療; 分子シャペロン

Newly synthesized protein and immature proteins are easily aggregated because they ej(pose hydrophobic regions. Many stress conditions, such as heat shock or hypoxia, slow down their folding process and cause accumulation of unfolded/misfolded proteins in the cell. Molecular chaperones, including heat shock'proteins (IISPs), are induced in these conditions, bind to unfolded/misfolded proteins, and help them to be folded or reflolded properly. The protective role of molecular chaperones for the cells under stress has been reported.Expression of angiogenic factors such as vascular endothelial growth factor (VEGF) under conditions of cell stress involves both transcriptionaland translational events, as well as an important role for inducible endoplasmicreticulum (ER) chaperones. Coexpression of VEGF and 150-kDaoxygen-regulated protein (ORP), a novel ER chaperone, in human glioblastomasuggested a link between angiogenesis and ORP150. C6 gliomacells stably transfected with ORP150 antisense … More displayed selectively reduced ORP150 expression. Tumors raised after in culation of immunocompromisedmice with ORP150 antisense C6 glioma transfectants demonstratedan initial phase of growth comparable to wild-type C6 gliomacells which was followed by marked regression within 8 days. Decreaseddensity of platelet/endothelial cell adhesion molecule 1-positive structureswithin the tumor bed was consistent with reduced angiogenesis in C6 gliomas expressing ORP150 antisense, compared with tumors derived from C6 cells overexpressing ORP150 sense or vector controls. In vitro,inhibition of ORP150 expression decreased release of VEGF into culturesupernatants ; in ORP150 antisense transfectants, VEGF accumulatedintracellularly within the ER. These findings demonstrate a critical rolefor the inducible ER chaperone ORP150 in tumor-mediated angiogenesisvia processing of VEGF, and, thus, highlight a new facet of angiogenicmechanisms amenable to therapeutic manipulation in tumors.Heat shock proteins (HSPs)/stress proteins are molecular chaperflnes that are induced by various environmental and physiological stimuli. Evidence of the relations between the expression of HSPs and the regulation of cell growth or transformation has accumulated. The 150-kDa oxygen-regulated protein (ORP150), a new member of lisp family, functions as a molecular chaperone in the endoplasmic reticulum. We have examined whether transduced antisense ORP150 cDNA reduces tumorigenicity and angiogenicity. Relations between these stress proteins and cancer and possibilities for anticancer gene therapy are described. Less

新合成的蛋白质和未成熟的蛋白质容易聚集，因为它们形成疏水区域。许多应激条件，如热休克或缺氧，减缓其折叠过程，并导致未折叠/错误折叠蛋白在细胞中积累。分子伴侣，包括热休克蛋白（IISP），在这些条件下诱导，结合未折叠/错误折叠的蛋白质，并帮助它们正确折叠或重新折叠。分子伴侣对应激状态下细胞的保护作用已有报道，在细胞应激条件下，血管生成因子如血管内皮生长因子（VEGF）的表达涉及转录和翻译事件，诱导型内质网（ER）分子伴侣也发挥重要作用。VEGF和150-kDa氧调节蛋白（ORP），一种新的ER伴侣，在人胶质母细胞瘤中的共表达表明血管生成和ORP 150之间存在联系。稳定转染ORP 150反义核酸的C6胶质瘤细胞 ...更多信息 显示选择性降低的ORP 150表达。用ORP 150反义C6神经胶质瘤转染子培养免疫功能低下的小鼠后，肿瘤生长的初始阶段与野生型C6神经胶质瘤细胞相当，随后在8天内明显消退。与过表达ORP 150正义或载体对照的C6细胞相比，在表达ORP 150反义的C6胶质瘤中，肿瘤床内血小板/内皮细胞粘附分子1阳性结构的密度降低与血管生成减少一致。在体外，抑制ORP 150表达减少了VEGF释放到培养上清液中;在ORP 150反义转染子中，VEGF在ER内细胞内聚集。这些研究结果表明，诱导型ER伴侣蛋白ORP 150在肿瘤介导的血管生成中通过加工VEGF发挥关键作用，因此，突出了肿瘤血管生成机制的一个新方面，适合于治疗操作。热休克蛋白的表达与细胞生长或转化的调节之间的关系的证据已经积累。150-kDa氧调节蛋白（ORP 150）是lisp家族的新成员，在内质网中起分子伴侣的作用。我们已经检查了转导的反义ORP 150 cDNA是否降低致瘤性和血管生成性。这些应激蛋白和癌症之间的关系和抗癌基因治疗的可能性进行了描述。少

项目成果

Kitao Y., Ozawa K., Miyazaki M., Kobayashi T., Yanagi H., Okabe M., Ikawa M., Yamashima T., Tohyama M., Stern D., Hori O., and Ogawa S.: "Expression of 150 kDa Oxygen Regulated Protein (ORP150), a Molecular Chaperone in the Endoplasmic Reticulum, Rescues

Kitao Y.、小泽 K.、宫崎 M.、小林 T.、柳 H.、冈部 M.、井川 M.、山岛 T.、远山 M.、斯特恩 D.、堀 O. 和小川 S.：“

Tamatani 他: "ORP150 protects against hypoxia/ischemia-induced neuronal death"Nature Med.. 7. 317-323 (2001)

Tamatani 等人：“ORP150 可以防止缺氧/缺血引起的神经元死亡”Nature Med.. 7. 317-323 (2001)

Miyagi他: "Antitumor Effect of Reduction of 150-kDa Oxygen-Regulated Protein Expression in Human Prostate Cancer Cells"Mol. Urol.. 5. 79-80 (2001)

Miyagi 等人：“人前列腺癌细胞中 150-kDa 氧调节蛋白表达减少的抗肿瘤作用”Mol Urol.. 5. 79-80 (2001)

Ozawa K, Tsukamoto Y, Hori O, Kitao Y, Yanagi Stern D, and Ogawa S.: "Regulation of tumor angiogeensis by ORP150, an inducible endoplasmic reticulum chaperone."Can. Res.. 61. 4206-4213 (2001)

Ozawa K、Tsukamoto Y、Hori O、Kitao Y、Yanagi Stern D 和 Okawa S.：“ORP150（一种诱导型内质网伴侣）对肿瘤血管生成的调节”。

Tsukamoto 他: "Expression of a novel RNA splicing factor, RA301/Tra2beta, in vascular lesions and its role in smooth muscle cell proliferation"Am. J. Pathol. 158. 1685-1694 (2001)

Tsukamoto 等人：“新型 RNA 剪接因子 RA301/Tra2beta 在血管病变中的表达及其在平滑肌细胞增殖中的作用”Am. J. Pathol。