Radioimmune & Inorganic Chemistry Section, Radiation Oncology Branch, National Cancer Institute, 10 Center Drive, Bethesda, Maryland 20892, United States.
Mol Pharm. 2012 Mar 5;9(3):374-81. doi: 10.1021/mp2003219. Epub 2011 Sep 21.
Herein we report the preparation along with the in vivo and in vitro MRI characterization of two generation four and five cystamine core dendrimers loaded with thirty and fifty-eight derivatized Gd-DOTA (G4SS30, G5SS58) respectively. Likewise the development and characterization of two half-dendrimers conjugated to the F(ab')(2) fragment of the monoclonal antibody (mAb) panitumumab functionalized with a maleimide conjugation functional group site (Ab-(G4S15)(4), Ab-(G5S29)(4)) are also described. The in vitro molar relaxivity of the Ab-(G4S15)(4) conjugate, measured at pH 7.4, 22 °C, and 3T showed a moderate increase in relaxivity as compared to Magnevist (6.7 vs 4.0 mM(-1) s(-1)) while the Ab-(G5S29)(4) conjugate was 2-fold higher (9.1 vs 4.0 mM(-1) s(-1)). The data showed that only a high injection dose (0.050 mmol Gd(3+)/kg) produced a detectable contrast enhanced contrast for the Ab-(G4S15)(4) conjugate while a lower dose (0.035 mmol Gd(3+)/kg) was sufficient for the Ab-(G5S29)(4) conjugate. The antibody-SMCC conjugate was purified by a Sephadex G-100 column, and the antibody-dendrimer-based agents were purified by spin filtration using a Centricon filter (50,000 MCO). The protein assay coupled with cysteine and Ellman's assay indicated an antibody to dendrimer ratio of 1:4. The in vivo blood clearance half-lives of the four agents measured at the jugular vein were ~12-22 min.
在此，我们报告了两种第四代和第五代半胱胺核树枝状大分子的制备及其体内外 MRI 特征，分别负载了三十和五十八个衍生的 Gd-DOTA（G4SS30、G5SS58）。同样，我们还描述了两种半树枝状大分子的开发和特征，这些半树枝状大分子与单克隆抗体（mAb）panitumumab 的 F(ab')(2)片段缀合，该片段用马来酰亚胺连接官能团位点（Ab-(G4S15)(4)、Ab-(G5S29)(4)）进行了功能化。在 pH 7.4、22°C 和 3T 下测量的 Ab-(G4S15)(4) 缀合物的体外摩尔弛豫率与 Magnevist 相比表现出适度增加（6.7 与 4.0 mM(-1) s(-1)），而 Ab-(G5S29)(4) 缀合物则增加了两倍（9.1 与 4.0 mM(-1) s(-1)）。数据表明，只有高注射剂量（0.050 mmol Gd(3+)/kg）才能产生可检测的 Ab-(G4S15)(4) 缀合物的对比增强对比，而低剂量（0.035 mmol Gd(3+)/kg）则足以满足 Ab-(G5S29)(4) 缀合物的需求。抗体-SMCC 缀合物通过 Sephadex G-100 柱进行纯化，而抗体-树枝状大分子基制剂通过离心超滤使用 Centricon 过滤器（50,000 MCO）进行纯化。蛋白质测定与半胱氨酸和 Ellman 的测定相结合表明抗体与树枝状大分子的比例为 1:4。在颈静脉测量的四种制剂的体内血液清除半衰期约为 12-22 分钟。
