Vijaykumar V, Topp E M
Department of Pharmaceutical Chemistry, University of Kansas, Lawrence 66045-2504, USA.
Pharm Res. 1995 Dec;12(12):1907-16. doi: 10.1023/a:1016424321378.
The purpose of the study was to develop a cell culture system representing a layer of cells in a solid tumor, and to use the system to study factors affecting the diffusion of a binding monoclonal antibody.
A mouse melanoma cell line, B16F10, was grown on Matrigel coated Transwell inserts. The diffusion of a binding monoclonal antibody which recognizes the murine transferrin receptor (a-TfR) was investigated, and compared to that of an isotype-matched, nonbinding control.
At a cell density of 6.4 x 10(5) cells/cm2, 37 degrees C and a donor antibody concentration of 10 nM, the permeability of the a-TfR antibody was approximately half that of the control. Blocking experiments, in which the cell layer was pretreated with excess a-TfR, resulted in equal permeabilities for a-TfR and control antibodies, suggesting that the difference in permeabilities observed in the absence of blocking is due to a-TfR binding. Experiments at 4 degrees C and in the presence of metabolic inhibitors also equalized the permeation rates of the two antibodies, indicating that internalization of the antibody/antigen complex is important in retarding the permeation of the binding antibody.
In this system, the diffusion of the a-TfR antibody is retarded by binding and internalization processes. The results have implications for the use of immunoconjugates in cancer chemotherapy, and for the delivery of other binding macromolecules to tissue sites of action.
本研究的目的是开发一种能代表实体瘤中细胞层的细胞培养系统，并利用该系统研究影响结合型单克隆抗体扩散的因素。
将小鼠黑色素瘤细胞系B16F10接种在基质胶包被的Transwell小室上。研究了识别小鼠转铁蛋白受体的结合型单克隆抗体（α-TfR）的扩散情况，并与同型匹配的非结合对照抗体进行比较。
在细胞密度为6.4×10⁵个细胞/cm²、37℃且供体抗体浓度为10 nM时，α-TfR抗体的通透性约为对照抗体的一半。在阻断实验中，用过量的α-TfR预处理细胞层，结果α-TfR抗体和对照抗体的通透性相等，这表明在未进行阻断时观察到的通透性差异是由于α-TfR结合所致。在4℃以及存在代谢抑制剂的条件下进行的实验也使两种抗体的渗透速率相等，这表明抗体/抗原复合物的内化在阻碍结合型抗体的渗透中起重要作用。
在该系统中，α-TfR抗体的扩散受到结合和内化过程的阻碍。这些结果对免疫缀合物在癌症化疗中的应用以及其他结合型大分子向组织作用部位的递送具有启示意义。
