Identification of organic anion transporter LST-2 which expressed in the gastrointestinal cancers: clinical implication for anticancer drug delivery.

背景与目的：发展肿瘤靶向化疗的一种方法是利用肿瘤细胞对药物的摄取增加。在这种情况下，癌细胞中较高浓度的这种药物是导致差异杀伤的原因，而正常人体细胞中较低浓度的药物则会减少副作用。本研究的目的是分离一种在正常细胞中弱表达但在癌细胞中大量表达的有机阴离子转运体，将抗癌药物输送到细胞内。方法：以LST-1基因为探针，筛选人肝脏cDNA文库，进行Northern印迹分析。制备LST-2特异性抗体，进行免疫组织化学包括免疫电子显微镜分析。采用非洲爪哇卵母细胞表达系统进行功能分析。我们还建立了一个稳定表达LST-2的永久细胞系，以检测MTX摄取与敏感性的关系。结果：分离到的LST-2与人LST-1的同源性为79.7%。正常情况下，LST-2仅在肝脏中表达，中央静脉周围肝细胞基底膜的免疫反应最强。虽然LST-2在肝脏中弱表达，但在胃、结肠癌和胰腺癌中大量表达，另一方面，SHE-LST-1只在肝细胞系中表达，LST-2以饱和和剂量依赖的方式转运甲氨蝶呤，此外，将LST-2基因导入哺乳动物细胞增强了对甲氨蝶呤的敏感性。结论：LST-2是决定甲氨蝶呤敏感性的首选候选分子之一，可能成为胃肠道肿瘤抗癌药物输送的良好靶点。

Background & Aims: One approach to the development of targeted cancer chemotherapy exploits increased uptake of the agent into neoplastic cells. In this scenario, higher concentrations of the agent in cancer cells are responsible for differential killing while the low concentration in normal human cells decreases side effects. The aim of this study is to isolate an organic anion transporter which in weakly in normal, but abundantly expressed in cancer cells, to deliver the anti-cancer drugs to the cells.Methods: A human liver cDNA library was screened with LST-1 cDNA as a probe, Northern blot analyses were performed using the isolated cDNA (termed LST-2). A LST-2 specific antibody was raised and immunohistochemical analyses including immuno-electron microscopy were performed. Xenopus oocyte expression system was used for functional analysis. We also established a permanent cell line which consistently expresses LST-2 to examine the relationship between MTX uptake and sensitivity.Results: The isolated cDNA, LST-2 has 79,7 % of overall homology wish human LST-1. LST-2 exclusively expressed in the liver under normal conditions and immunoreactivity was highest at the basolateral membrane of the hepatocytes around the central vein. Although its weak expression in the liver, LST-2 is abundantly expressed in the gastric, colon and pancreatic cancers, On she other hand, she LST-1 was only detected in a hepatic cell line, LST-2 transports methotrexate in a saturable and dose dependent manner Furthermore, introduction of the LST-2 gene into mammalian cells potentiates sensitivity to methotrexate.Conclusion: LST-2 is one of the prime candidate molecule for determining methotrexate sensitivity and may be a good target on deliver anti-cancer drugs to the gastrointestinal cancers.