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很容易聚集，因为它们形成疏水区域。许多应激条件，如热休克或缺氧，会减慢其折叠过程并导致未折叠/错误折叠的蛋白质在细胞中积累。分子伴侣，包括热休克蛋白（IISP），在这些条件下被诱导，与未折叠/错误折叠的蛋白质结合，并帮助它们被折叠。 正确折叠或重新折叠。分子伴侣对处于应激状态下的细胞的保护作用已有报道。血管生成因子如血管内皮生长因子 (VEGF) 在细胞应激条件下的表达涉及转录和翻译事件，以及诱导型内质网 (ER) 伴侣的重要作用。 VEGF 和 150-kDa 氧调节蛋白的共表达 (ORP) 是一种新型 ER 伴侣，在人类胶质母细胞瘤中表明血管生成与 ORP150 之间存在联系。用 ORP150 反义病毒稳定转染的 C6 神经胶质瘤细胞显示出选择性降低的 ORP150 表达。用 ORP150 反义 C6 神经胶质瘤转染子培养免疫受损小鼠后产生的肿瘤显示出与 野生型 C6 神经胶质瘤细胞随后在 8 天内显着消退。与源自过表达 ORP150 有义或载体对照的 C6 细胞的肿瘤相比，肿瘤床内血小板/内皮细胞粘附分子 1 阳性结构密度的降低与表达 ORP150 反义的 C6 神经胶质瘤中血管生成的减少一致。体外，抑制 ORP150 表达减少了 VEGF 向培养上清液中的释放；在 ORP150 反义转染子中，VEGF 在 ER 细胞内积累。这些发现证明了诱导型 ER 伴侣 ORP150 在肿瘤介导的血管生成（通过 VEGF 的处理）中发挥着关键作用，因此突出了适合治疗的血管生成机制的一个新方面。 热休克蛋白（HSP）/应激蛋白是由各种环境和生理刺激诱导的分子伴侣。 HSP 表达与细胞生长或转化调节之间关系的证据已经积累。 150 kDa 氧调节蛋白 (ORP150) 是 lisp 家族的新成员，在 内质网。我们检查了转导的反义 ORP150 cDNA 是否会降低致瘤性和血管生成性。描述了这些应激蛋白与癌症之间的关系以及抗癌基因治疗的可能性。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。