- 學位論文
鎝99m(I)標誌Trastuzumab單株抗體作為乳癌細胞造影劑之研究
Study on 99mTc(I) Labeled Trastuzumab Monoclonal Antibody as a Specific Marker of HER2-overexpressed Cells for Breast Cancer Imaging
摘要
Herceptin是近年來常見的抗乳癌藥物，其主要成份為trastuzumab。Trastuzumab是由DNA重組製成的人類單株抗體(Humanized Monoclonal Antibody，mAb)，具高度專一性地親和作用於細胞膜上的人類上皮生長因子接受器第二蛋白(Human Epidermal Growth Factor Receptor 2 Protein，HER2)，因而阻礙生長因子與HER2受體結合使細胞增生速度下降，此外，抗體亦能誘導殺手細胞（Killer cell）和巨噬細胞（Macrophage）殺死過度HER2表現的乳癌細胞，達到治療之目的。 本研究發展出新的鎝-99m標誌方式，並利用trastuzumab對HER2之專一性，將放射性核種鎝-99m(I)標誌於trastuzumab單株抗體，作為HER2過量表現乳癌細胞之造影劑。 利用無機化學合成方法製備的三羰基鎝化合物（[99mTc(CO)3(OH2)3]＋）為前驅物並將其標誌於抗體，得到99mTc(I)-trastuzumab，再利用高效能液相層析、分子大小排阻層析法、免疫反應分析與動物實驗來測試其標誌效率、產率、穩定度、對乳癌之活性及專一性。經實驗證明，99mTc(I)-trastuzumab在血清中相當穩定並且仍保有trastuzumab本身之專一性與活性；在生物分佈實驗中，亦發現99mTc(I)-trastuzumab與HER2親和之高度專一性。
並列摘要
Herceptin, the common anti-breast cancer medicine prevailed in recent years, contains mainly trastuzumab. Trastuzumab, the DNA recombinated humanized monoclonal antibody (mAb), can recognize specifically Human Epidermal Growth Factor Receptor 2 protein (HER2) on cell membrane. HER2 is a 185 kDa transmembrane receptor tyrosine kinase which is overexpressed in 25-30% of breast cancers. As HER2 is blocked by trastuzumab mAb, the growth factor will greatly reduce its binding with the receptor leading to lower the cell proliferation. Moreover, trastuzumab mAb will induce natural killer cells and macrophage to kill the cancer cells to cure breast cancer. The study is to develop a novel 99m Tc labeled trastuzumab construct and via the high affinity of trastuzumab toward HER2 to adopt as an imaging agent for HER2 overexpressed breast cancer. For the radiolabeling, the tricarbonyl techenium compound, [99mTc(CO)3(OH2)3]+, was synthesized and utilized as the precursor to conjugate rapidly with the monoclonal antibody, trastuzumab, by coordinating with N, O and S donors. The labeled mAb, 99mTc(I)-trastuzumab, was characterized by high performance liquid chromatography (HPLC) and size-exclusion chromatography (SEC) for its labeling yield and radiochemical purity. In addition, immunoreactivity assay and animal model experiment were undertaken to confirm its in vitro and in vivo stability, immunoreactivity and specific affinity with HER2 overexpressed cells. In conclusion, 99mTc(I)-trastuzumab is stable in serum and retains the inherent property and immunoreactivity of trastuzumab. The result of biodistribution also reveals that 99mTc(I)-trastuzumab is of high affinity with HER2-positive BT-474 cells and related animal tumor, in constrast with much lower affinity with HER2-negative MCF-7 models.