Rothbarth Joost, Koevoets Cindy, Tollenaar Rob A E M, Tilby Mike J, van de Velde Cornelis J H, Mulder Gerard J, Kuppen Peter J K
Department of Surgery, Leiden University Medical Center, Albinusdreef 2, Leiden 2333, The Netherlands.
Biochem Pharmacol. 2004 May 1;67(9):1771-8. doi: 10.1016/j.bcp.2003.12.038.
Melphalan is a chemotherapeutic drug that exerts its cytotoxic effect mainly through the formation of DNA adducts. We report the specific immunohistochemical detection and visualisation of melphalan-DNA adducts using the monoclonal antibody MP5/73 in cultured tumour cells and solid tumour tissue from colorectal liver metastases from patients treated with melphalan. The human colon cancer cell lines HT29, SW480 and SW1116, and the rat colon cancer cell line CC531 were exposed to different concentrations of melphalan. In addition, tumour samples from 17 patients with colorectal liver metastases treated by isolated hepatic perfusion with high dose melphalan (200mg) were collected. Cell lines and tumour samples were stained with the MP5/73 antibody against melphalan-DNA adducts and cell viability was determined by an MTT assay. Melphalan-DNA adducts could be visualised by immunohistochemistry in both cultured cells and solid tumour tissue. A correlation between melphalan exposure concentration, the subsequent melphalan-DNA adduct staining intensity, and melphalan cytotoxicity existed for each individual cell line, but the level of both parameters independently differed between cell lines. Specific staining for melphalan-DNA adducts also was feasible in the human solid tumour tissue. There was considerable variation in melphalan-DNA adduct staining, staining intensity, and distribution in the tumour stroma and the tumour epithelium among the different patients. Melphalan-DNA adducts appeared to be more intense in tumour cells at the border of the tumour nodules than in tumour cells in the centre. Thus, visualisation of melphalan-DNA adducts by immunohistochemistry allows the study of distribution of melphalan-DNA adducts in solid tumours.
美法仑是一种化疗药物，其细胞毒性作用主要通过形成DNA加合物来发挥。我们报告了使用单克隆抗体MP5/73在接受美法仑治疗患者的培养肿瘤细胞和结直肠肝转移实体瘤组织中特异性免疫组织化学检测和可视化美法仑-DNA加合物的情况。将人结肠癌细胞系HT29、SW480和SW1116，以及大鼠结肠癌细胞系CC531暴露于不同浓度的美法仑。此外，收集了17例接受高剂量美法仑（200mg）经肝动脉隔离灌注治疗的结直肠肝转移患者的肿瘤样本。细胞系和肿瘤样本用抗美法仑-DNA加合物的MP5/73抗体染色，并通过MTT法测定细胞活力。美法仑-DNA加合物可通过免疫组织化学在培养细胞和实体瘤组织中可视化。每个细胞系中美法仑暴露浓度、随后的美法仑-DNA加合物染色强度和美法仑细胞毒性之间存在相关性，但这两个参数的水平在不同细胞系之间独立存在差异。美法仑-DNA加合物的特异性染色在人实体瘤组织中也是可行的。不同患者之间美法仑-DNA加合物染色、染色强度以及在肿瘤基质和肿瘤上皮中的分布存在相当大的差异。美法仑-DNA加合物在肿瘤结节边缘的肿瘤细胞中似乎比在中心的肿瘤细胞中更强烈。因此，通过免疫组织化学可视化美法仑-DNA加合物有助于研究美法仑-DNA加合物在实体瘤中的分布。
