Studies for Gene Therapy of Sphingolipidosis

鞘脂沉积症基因治疗研究

• 批准号: 10044321
• 负责人: ETO Yoshikatsu
• 金额: $ 3.39万
• 依托单位: The Jikei University School of Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10044321/
• 关键词: lipidosis; Gene analysis; Gene therapy; neuronal cells; 造血幹細胞; アデノウイルスベクター; 先天代謝異常症; 遺伝子治療; 中枢神経症状; 動物モデル

Gene analysis of Japanese lipidodes patients :We analyzed gene from various Japanese patients with lipidoses and found that the distribution of gene mutations was quite different from that from other ethnic groups.Gene therapy :The deficiency of human β-glucuronidase(HBG) results in MPS type VII(Sly syndrome). In this study, we tested the ability to target CD18/CD11b positive cells with gene therapy for murine MPS VII. We harvested bone marrow cells from syngeneic normal mice and cultivated in CSF-1 containing medium for 2 weeks. More than 95% of the cells were double positive for CD18/CD11b by flowcytometry. We gave 2×10ィイD16ィエD1 cells to the non-myeloablated Sly mouse. One week post-transplantation, donor cells populated liver and spleen. The HBG activity increased from 0.9±0.7 to 28.4±12.5u/mg and 0.7±0.4 to 29.7±23.1u/mg in liver and spleen respectively. However, the pathology was unchanged. The HBG activity in liver and spleen decreased b 5 weeks to 3.7±1.5 and 2.3±0.5 respectivel … More y, but the pathological improvements were significant and glycosaminoglycan storage was largely cleared. Next, the CD18/CD11b cells were collected from Sly mouse by the method above, transduced by HBG expressing retrovirus(MFG-HBG), and given to non-myeloablated Sly mouse. By 5 weeks post-transplantation, HBG activity was only marginally above the control level. However, many HBG positive cells were observed and pathological improvement was significant. These data suggest that CD18/CD11b cells transplantation is promising for treatment of murine MPS VII without myeloablation, and CD18/CD11b cells may be a good targets for gene therapy for Sly syndrome.Gene transfer to neuronal cells :Because significant prenatal pathology occurs in genetic diseases, postnatal therapy is often not sufficient to correct them, especially if damage has occurred in the central nervous system. In such diseases, prenatal treatment will be required. Adenovirus mediated gene transfer may be useful for this purpose because of its wide host range and efficient gene transfer to various organs. However, most of the gene transfer studies have been carried out at a relatively late stage of embryogenesis. In this study, a recombinant adenovirus expressing the E. coli lacZ gene (AxCALacZ) was administered into the rat embryos at the 8th to 12th day of gestation. The results of the study revealed that the lacZ gene was expressed in many different organs including liver, heart, skin and brain. These results suggest that the adenovirus vector may be useful for the prenatal gene therapy of many genetic diseases. Less

基因治疗：人类β-葡萄糖醛酸酶（HBG）缺乏导致MPS VII型（Sly综合征）。在这项研究中，我们测试了小鼠MPS VII基因治疗靶向CD 18/CD 11b阳性细胞的能力。我们从同系正常小鼠收获骨髓细胞，并在含有CSF-1的培养基中培养2周。流式细胞仪检测结果显示95%以上的细胞为CD 18/CD 11b双阳性。我们将2×10 ~（-16）μ g/ml的D16细胞给予非骨髓清除的Sly小鼠。移植后一周，供体细胞植入肝脏和脾脏。肝、脾中HBG活性分别从0.9±0.7上升到28.4±12.5u/mg和0.7±0.4上升到29.7±23.1u/mg。然而，病理学没有改变。肝、脾HBG活性B 5周后分别降至3.7±1.5和2.3±0.5。 ...更多信息 y，但病理学改善是显著的，糖胺聚糖储存在很大程度上被清除。接着，通过上述方法从Sly小鼠收集CD 18/CD 11b细胞，用表达HBG的逆转录病毒（MFG-HBG）转导，并给予非骨髓清除的Sly小鼠。移植后5周，HBG活性仅略高于对照水平。然而，观察到许多HBG阳性细胞，病理学改善显著。这些数据表明，CD 18/CD 11b细胞移植是有希望的治疗小鼠MPS VII没有骨髓消融，和CD 18/CD 11b细胞可能是一个很好的靶点的基因治疗Sly syndrome.Gene transfer to neuronal cells：因为显着的产前病理发生在遗传性疾病，出生后的治疗往往是不足以纠正他们，特别是如果损害已经发生在中枢神经系统。在这些疾病中，需要进行产前治疗。腺病毒介导的基因转移可用于此目的，因为其广泛的宿主范围和有效的基因转移到各种器官。然而，大多数基因转移研究都是在胚胎发生的相对晚期阶段进行的。本研究用重组腺病毒载体表达E.将大肠杆菌lacZ基因（AxCALacZ）注入孕8 ~ 12天的大鼠胚胎。研究结果表明，lacZ基因在许多不同的器官中表达，包括肝脏、心脏、皮肤和大脑。这些结果表明，腺病毒载体可用于许多遗传性疾病的产前基因治疗。少

项目成果

Ida H., Rennert OM., Eto Y., et al.: "Clinical and genetic studies of Japanese homozygotes for the・・・"Hum Genet. 105. 120-6 (1999)

Ida H.、Rennert OM.、Eto Y. 等人：“日本纯合子的临床和遗传学研究......”Hum Genet. 105. 120-6 (1999)

Oishi K., , Ida H., Eto Y., et al.: "Clinical and molecular of Japanese patients with neuronal・・・"Molecular Genetics and Metabolism. 66. 344-348 (1999)

Oishi K., , Ida H., Eto Y., et al.：“日本神经元患者的临床和分子......”分子遗传学和代谢 66. 344-348 (1999)

Ida H., Rennert OM., Eto Y., et al.: "Clinical and genetic studies of Japanese homozygotes for the・・・"Hum Genet. 105. 120-126 (1999)

Ida H.、Rennert OM.、Eto Y. 等人：“日本纯合子的临床和遗传学研究......”Hum Genet. 105. 120-126 (1999)

Ohashi,T., Lizuka,S., Sly,W.S., Machiki,Y., Eto,Y.: "Efficient and persistent expression of β-glucuronidase gene in CD34+cells from human umbiical cord blood by retroviral vector."Eur J Haematol. 61(4). 235-239 (1998)

Ohashi,T.、Lizuka,S.、Sly,W.S.、Machiki,Y.、Eto,Y.：“通过逆转录病毒载体在人脐带血 CD34+ 细胞中高效、持久地表达 β-葡萄糖醛酸酶基因。”Eur J苏木醇。61(4)。

Yokoo T., Ohashi T., Eto Y., et al.: "Prophyaxis of Antibody-Induced Actue Glomerulonephritis・・・"Hum Gene Ther. 10. 2673-8 (1999)

Yokoo T.、Ohashi T.、Eto Y. 等：“预防抗体诱发的急性肾小球肾炎……”Hum Gene Ther。