標的型磁振造影對比劑之合成及特性研究

Abstract

近年來，磁振造影(MRI)於人體之應用已發展至細胞及分子的層級。為了使MRI技術應用於細胞、分子與功能性影像，能夠標幟活體細胞的目標化、區域化及數量化之新一代MRI對比劑與技術是目前最迫切需要的。因此，目前兩種最主要的MRI對比劑的研究與發展方向：小型分子量之釓金屬錯合物及氧化鐵奈米粒子。高度表現於人類腫瘤上新穎的Legumain 蛋白酶(protease)與腫瘤侵犯及轉移有關， Legumain可當作標的至表現這些蛋白酶之腫瘤。本研究之目的為利用親脂性之釓金屬錯合物，並將此鍵結上目標化之Legumain，形成具目標化親脂性之釓金屬錯合物，可以非侵入的方式標幟與追蹤癌細胞。此外，許多研究顯示體內金屬離子的濃度不平衡會導致相當多疾病，例如阿茲海默症。因此為了早期發現，以降低這些疾病發病後對人體的傷害。本研究設計了檢測銅離子的磁振造影對比劑，藉由銅離子的配位而改變內層水分子數使弛緩率增加，以達到可偵測高濃度之銅離子之目的。釓金屬錯合物之物性、化性探討，包括熱力學穩定度、弛緩率(r1)、內層水分子數、水分子交換速率、轉動相關時間及與蛋白酶作用之探討是本研究的重點。最後，體外相關實驗與MR影像也將被研究探討。在氧化鐵奈米粒子方面，近年逐漸開發以紅外光的發光團在活體標靶上的研究，除了快速提供活體資訊外，也提高了影像的靈敏度與空間辨識度。因此，我們發展高弛緩率之奈米粒子，並在其奈米表面修飾上生物相容性聚合分子，用以鍵結近紅外光之發光團，且以此基礎應用在抗體(Anti-MUC4 antibody)標靶於細胞膜上有黏蛋白(MUC4)表現之腫瘤；此外另將β-galactose與螢光基團courmarin結合修飾於氧化鐵奈米粒子表面作為生物活化型對比劑，藉由酵素切割致使氧化鐵形成聚集而改變弛緩率。我們研究超順磁氧化鐵粒子之物、化性質，包括粒徑大小、穩定性、磁化率、XRD 晶型研究等。最後利用光學及MR 影像來證明已修飾超順磁氧化鐵奈米粒子之效能。

Recently, MRI has been developed to examine living organisms down to the cellular and molecule level.To exploit the advancement of MRI technique for cellular, molecule and functional imaging, there are increasing needs for developing new MRI contrast agents and techniques for cell and molecule labeling to report the localization, movement, mass, and functions of cells in vivo. Therefore, two major classes of contrast agents are available for MRI: small molecular weight Gd3+ chelates and iron oxide nanoparticles. The purpose of research is tuning the lipophilicity of Gd3+ complexes and conjugating with peptide substrates of Legumain protease. Legumain protease is greatly related to tumor invasion and metastasis, and is also highly expressed in majority of human tumors, which make them as the very representative cancer proteases. To find the optimum lipophilicity of Gd3+ complexes that can be stably incorporated into cell membranes may serve as a useful tool for tumor cell labeling and tracking. Ideally, these Gd3+ chelates should label intact cell membranes noninvasively at low concentrations and with fast kinetics, and should remain on labeled cells over a period of time to allow repetitive imaging. In addition, many evidence indicated that the metal ion imbalance in the living orgamisms will give rise to many diseases such as Alzheimer’s disease. For early detection to reduce the damage after the onset of these diseases, we design a contrast agent for the detection of copper ions. T1 relaxivity of contrast agent will increase by change the number of inner sphere water when Cu2+ presence. The chemical and physical properties of these Gd3+ complexes will be characterized, including thermodynamic stability constant, relaxivity (r1), the number of inner-sphere water and kinetic parameter. Finally, the MR imaging will be conducted as well. On the other hand, a NIR fluorochrome and a biological moiety that possess lock-and-key interactions can provide consistent in vivo imaging information with high sensitivity and spatial resolution. We have synthesized iron oxide nanoparticle with high relaxivity. Post-synthesis surface modification with PEG expose the amine group which have been used for bioconjugation of anti-MUC4 antibody bearing near-infrared (NIR) fluorophore which can specifically target MUC4 mucin expression tumor cell; The β-galactose and courmarin will conjugated on iron oxide nanoparticles as a biological activated contrast agent, after cutting by enzyme the iron oxide will aggregate and the relaxation rate will be changed .The geometry, structural features, and physical properties of magnetite nanoparticles will be characterized. To prove effectiveness of this MR probe, the optical and MR image will be used.