DEMELOPMENT OF RAKIOLABELED SMALLER ANTI-TENASCIN-CANTIBODY FOR CLINICAL USE

RAKIO标记的较小抗腱蛋白抗体的开发用于临床用途

• 批准号: 17390342
• 负责人: IRIE Toshiaki
• 金额: $ 9.67万
• 依托单位: National Institute of Radiological Sciences
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17390342/
• 关键词: DIAGNOSTIC IMAGING; REMODELING; EXTRACELLUAR MATRIX; GENETIC ENGINEERING; ANTIBODY

Hybridoma cells of mouse monoclonal anti-tenascin-C (TNC) antibody, Fab fragments of anti-TNC antibody, and refined TNC were prepared to make new radiolabeled smaller antibody against TNC. The DNA fragments encoding variable domains of heavy and light chains (V_H and V_L, respectively) in a mouse anti-TNC antibody (4F1OTT and 4C8MS) were cloned and combined to give the scFv gene fragment encoding the sequence of 5'-VH-linker-VL-FLAG-Cys-3'. The total RNA was extracted from hybridoma cells secreting mouse anti TNC antibody and reverse-transcribed to produce the first-strand cDNA encoding the V_H and V_L,. The V_H and VL DNA fragments were gel-purified and were spliced by overlap extension PCR. Peri plasmic expression of this gene in E. coli cells provided soluble scFv proteins that showed similar affinity to TNC to that of the Fab fragment of progenitor antibody.The bifunctional chelating agent (EMCS-Bz-EDTA) was synthesized and reacted with a cysteine residue, which was induced at C-te … More rminal of the scFv. 111In-anti-TNC-scFv was injected intravenously in rats after producing myocardial infarction (Ml). Biodistribution of each organ was measured. By autoradiography high radioactivity of ^<111>In-anti-TNC-scFv was observed in the granulation tissue around the necrotic area in the acute MI rat, which corresponds to the localization of TNC molecule detected by immunohistochemistry.To validate clinical usefulness, ^<111>In-anti-TNC-Fab was injected intravenously to MI rat. Dual-isotope single-photon emission computed tomography imaging (SPECT) was performed. Two kinds of mIn-anti-TNC-Fab accumulated higher than ^<111>In-non-specific-Fab antibody at necrotic area. A rat with larger accumulation of win-anti-INC-Fab at heart showed myocardial enlargement after MI by echocardiography. In addition, ^<111>In-anti-TNC-Fab was injected into experimental glioma mice and revealed the feasibility of in vivo imaging for tumor. This tracer was useful for evaluating various diseases. Long term safety after injection of ^<111>In-anti-TNC-Fab was also certified.This research is planning to apply a patent. Less

制备了小鼠抗腱生蛋白-C（TNC）单克隆抗体、抗TNC抗体Fab片段和精制TNC的杂交细胞，以制备新的放射性标记的更小的抗TNC抗体。将编码小鼠抗TNC抗体（4F 1 OTT和4C 8 MS）的重链和轻链可变区（分别为V_H和V_L）的DNA片段克隆并组合，得到编码5 '-VH-接头-VL-FLAG-Cys-3'序列的scFv基因片段。从分泌小鼠抗TNC抗体的杂交瘤细胞中提取总RNA，反转录得到编码V_H和V_L的第一链cDNA。将VH和VL DNA片段进行凝胶纯化，并通过重叠延伸PCR进行拼接。该基因在E.用自制的双功能螯合剂EMCS-Bz-EDTA与在C-te诱导的半胱氨酸残基反应，获得了与TNC具有相似亲和力的可溶性单链抗体，并将该双功能螯合剂与C-te诱导的半胱氨酸残基反应，获得了与TNC具有相似亲和力的单链抗体。 ...更多信息 scFv的重组。111 In-抗-TNC-scFv在大鼠心肌梗死（MI）后静脉注射。测量各器官的生物分布。放射自显影显示，<111>急性心肌梗死大鼠坏死区周围的肉芽组织中有高放射性的抗TNC单链抗体，与免疫组化检测到的TNC分子的定位一致<111>。进行双同位素单光子发射计算机断层扫描成像（SPECT）。两种mIn-抗TNC-Fab抗体在坏死区的蓄积量均高于mIn<111>-非特异性Fab抗体。在心脏中具有较大的win-抗-INC-Fab积累的大鼠通过超声心动图显示MI后的心肌增大。此外，将<111>抗TNC-Fab抗体注射到实验性胶质瘤小鼠中，并揭示了肿瘤体内成像的可行性。该示踪剂可用于评估各种疾病。注射后的长期安全性<111>也得到了证实。这项研究正在计划申请专利。少

项目成果

Eplerenone attenuates myocardial fibrosis in the angiotensin II-induced hypertensive mouse: Involvement of tenascin-C induced by aldosterone-mediated inflammation

• DOI: 10.1097/fjc.0b013e318033dfd4
• 发表时间: 2007-05-01
• 期刊: JOURNAL OF CARDIOVASCULAR PHARMACOLOGY
• 影响因子: 3
• 作者: Nishioka, Tomohiro;Suzuki, Maiko;Imanaka-Yoshida, Kyoko
• 通讯作者: Imanaka-Yoshida, Kyoko

Technetium-99m-labeled long chain fatty acid analogues metabolized by beta-oxidation in the heart.

• DOI: 10.1021/jm061017g
• 发表时间: 2007-01
• 期刊: Journal of medicinal chemistry
• 影响因子: 7.3
• 作者: T. Uehara;Tomoe Uemura;S. Hirabayashi;Sayaka Adachi;K. Odaka;H. Akizawa;Y. Magata;T. Irie;Y. Arano

Assessment of macrocyclic triamine ligands as synthons for organometallic (99m)Tc radiopharmaceuticals.

大环三胺配体作为有机金属 (99m)Tc 放射性药物合成子的评估。

• 发表时间: 2008
• 期刊: Inorg Chem. 47
• 作者: Suzuki K;Shimmura N;Thipyapong K;Uehara T;Akizawa H;Arano Y.
• 通讯作者: Arano Y.

Assessment of macrocyclic triamine ligands as synthons for organometalic(99m)Tc radiopharmaceuticals.

大环三胺配体作为有机金属(99m)Tc放射性药物合成子的评估。

• 发表时间: 2008
• 期刊: Inorg Chem. 47
• 作者: Suzuki K;Shimmura N;Thipyapong K;Uehara T;Aldzawa H;Arano Y.
• 通讯作者: Arano Y.

Development of radiolabeled smaller anti-tenascin-C antibody for clinical use : comparison between two antibodies by dual injection.

开发用于临床的放射性标记的较小抗生腱蛋白-C 抗体：通过双重注射比较两种抗体。

• 发表时间: 2007
• 期刊: Innervision. 22
• 作者: odaka K;Uehara T;Kobayashi N;Imanaka-Yoshida K;Arano Y;Tanada S;Irie T.
• 通讯作者: Irie T.