Press O W, Shan D, Howell-Clark J, Eary J, Appelbaum F R, Matthews D, King D J, Haines A M, Hamann P, Hinman L, Shochat D, Bernstein I D
Department of Medicine (Division of Oncology), University of Washington, Seattle 98195-6043, USA.
Cancer Res. 1996 May 1;56(9):2123-9.
Radiolabeled antibodies have produced encouraging remissions in patients with chemotherapy-resistant hematological malignancies; however, the selection of therapeutic radionuclides for clinical trials remains controversial. In this study, we compared the internalization, lysosomal targeting, metabolism, and cellular retention of radiolabeled murine and humanized monoclonal antibodies targeting the CD33 antigen (monoclonal antibodies mP67 and hP67, respectively) on myeloid leukemia cell lines (HEL and HL-60) and of anti-carcinoma antibodies (monoclonal antibodies hCTM01 and hA33) targeting breast cancer and colorectal carcinoma cell lines (MCF7 and Colo 205, respectively). Each antibody was labeled with 125I (by the IodoGen method) and with 111In and 90Y using macrocyclic chelation technology. Targeted tumor cells were analyzed for retention and metabolism of radioimmunoconjugates using cellular-radioimmunoassays, Percoll gradient fractionation of cell organelles, SDS-PAGE, and TLC of cell lysates and culture supernatants. Our results suggest that antibodies are routed to lysosomes after endocytosis, where they are proteolytically degraded. [125I]monoiodotyrosine is rapidly excreted from cells after lysosomal catabolism of antibodies radioiodinated by conventional methods, whereas small molecular weight 111In and 90Y catabolites remain trapped in lysosomes. As a consequence of the differential disposition of small molecular weight catabolites, 111In and 90Y conjugates displayed superior retention of radioactivity compared with 125I conjugates when tumor cells were targeted using rapidly internalizing antibody-antigen systems (e.g., hP67 with HEL cells and hCTM01 with MCF7 cells). When tumor cells were targeted using antibody-antigen systems exhibiting slow rates of endocytosis (e.g., hP67 on HL-60 cells and hA33 on Colo 205 cells), little differences in cellular retention of radioactivity was observed, regardless of whether 125I, 111In, or 90Y was used.
放射性标记抗体已在化疗耐药的血液系统恶性肿瘤患者中产生了令人鼓舞的缓解效果；然而，用于临床试验的治疗性放射性核素的选择仍存在争议。在本研究中，我们比较了靶向CD33抗原的放射性标记鼠源和人源化单克隆抗体（分别为单克隆抗体mP67和hP67）在髓系白血病细胞系（HEL和HL - 60）上以及靶向乳腺癌和结肠癌细胞系（分别为MCF7和Colo 205）的抗癌抗体（单克隆抗体hCTM01和hA33）的内化、溶酶体靶向、代谢及细胞内滞留情况。每种抗体均用125I（通过IodoGen法）以及使用大环螯合技术用111In和90Y进行标记。使用细胞放射免疫分析、细胞器的Percoll梯度分级分离、SDS - PAGE以及细胞裂解物和培养上清液的TLC分析靶向肿瘤细胞中放射免疫缀合物的滞留和代谢情况。我们的结果表明，抗体在胞吞作用后被转运至溶酶体，在那里它们被蛋白水解降解。通过传统方法放射性碘化的抗体在溶酶体分解代谢后，[125I]单碘酪氨酸迅速从细胞中排出，而小分子量的111In和90Y分解代谢产物则被困在溶酶体中。由于小分子量分解代谢产物的处置差异，当使用快速内化抗体 - 抗原系统（例如hP67与HEL细胞以及hCTM01与MCF7细胞）靶向肿瘤细胞时，111In和90Y缀合物相比125I缀合物显示出更高的放射性滞留。当使用内吞速率较慢的抗体 - 抗原系统（例如HL - 60细胞上的hP67以及Colo 205细胞上的hA33）靶向肿瘤细胞时，无论使用125I、111In还是90Y，在细胞内放射性滞留方面观察到的差异都很小。
