Genome Analysis through Reverse Translation of Human Bone Metastasis-Specific Protein

通过人骨转移特异性蛋白的反向翻译进行基因组分析

基本信息

批准号：
19591741
负责人：
TSURU Michiyo
金额：
$ 2.75万
依托单位：
Kurume University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2007
资助国家：
日本
起止时间：
2007 至 2010
项目状态：
已结题

项目摘要

Primary cancer control has become almost feasible, and controlling the spread of metastases to remote organs is the main focus of future cancer treatment. Even if a patient receives the best primary cancer treatment, metastasis of cancer occurs mostly five or ten years after the treatment when the cancer is considered to be completely cured. For the clinical test, a time-series follow-up study was conducted on blood samples obtained from each patient after the primary cancer was completely cured. Clinically, the target protein is present in the blood before the spread of definitive bone metastases (screening for bone metastases can be achieved by bone scintigraphy) and shows a decreasing trend after the initiation of chemotherapy and radiotherapy. A time-series follow-up was conducted on an individual basis. Regarding the purification method, the target peak was detected for the unattached fraction and the elution fraction (pH 7.0) was obtained after fractionating 200 μL of the blood s … More erum in an anion exchange column. Using this fraction as a partial purification fraction, the unattached and elution fractions (pH 7.0) were added to the cation exchange column and eluted by increasing the level of NaCl concentration. Consequently, the target peak was confirmed for 0.2 M and 0.3 M NaCl fractions.These fractions were then applied onto reverse-phase HPLC (2 mm) and eluted by the concentration gradient of acetonitrile. However, because the peak immediately adjacent to the target peak could not be separated and there was a possibility that high molecular protein, which could not be confirmed by SELDI may have existed, the fraction was applied onto the normal-phase HPLC system, where the target peak was successfully obtained. A tertiary structure analysis was performed on the amino acid sequence of the target protein using a super computer. First, an identified amino acid was synthesised. An antibody was generated by immunizing a rabbit with this amino acid, and the antibody was then purified. Furthermore, the result of tertiary structural analysis of this target protein has suggested that its synthesis for antibody pharmaceuticals can be expected. The target antigenic peptide and cancer cells were transplanted into the artery of a nude mouse, and systemic metastasis was confirmed by CT scan ten days after transplantation. Less

原发性癌症控制已经变得几乎是可行的，控制转移到偏远器官的传播是未来癌症治疗的主要重点。即使患者接受了最佳的原发性癌症治疗，癌症的转移大部分发生在治疗后五到十年后，当癌症被认为是完全治愈的。对于临床测试，对原发性癌症完全治愈后，对从每位患者获得的血液样本进行了时间序列随访研究。在临床上，靶蛋白在血液中呈现在确定性骨转移酶（可以通过骨闪烁图筛查可以实现骨转移）之前，并在化学疗法和放疗后显示出趋势下降。单独进行时间序列随访。关于纯化方法，检测到无附加部分的靶峰，并在分馏200μl血液S…中获得了模子分数（pH 7.0）。将此部分用作部分纯化部分，将未连接的和洗脱部分（pH 7.0）添加到阳离子交换柱中，并通过增加NaCl浓度水平来洗脱。因此，确认目标峰为0.2 m和0.3 m NaCl级分。然后将这些分数应用于反相HPLC（2 mm），并通过乙腈的浓度梯度洗脱。但是，由于无法分离与目标峰附近的峰，并且可能存在高分子蛋白（无法通过塞尔德斯证实的高分子蛋白，因此将馏分应用于正常相HPLC系统，在该系统中成功获得了目标峰。使用超级计算机对目标蛋白的氨基酸序列进行了三级结构分析。首先，合成了确定的氨基酸。通过用这种氨基酸免疫兔来产生抗体，然后纯化抗体。此外，该靶蛋白的三级结构分析的结果表明，可以预期其与抗体药物的合成。将靶抗原性胡椒和癌细胞移植到裸小鼠的动脉中，并在移植后十天通过CT扫描确认全身转移。较少的