Nakanishi T, Tamai I, Sai Y, Sasaki T, Tsuji A
Department of Pharmacobio-dynamics, Faculty of Pharmaceutical Sciences, Kanazawa University, Japan.
Cancer Res. 1997 Sep 15;57(18):4118-22.
To explore the feasibility of targeting human tumor cells via their transport systems, dipeptide uptake was studied in the human fibrosarcoma cell line HT1080 and the human fibroblast cell line IMR-90 by the use of hydrolysis-resistant glycylsarcosine (Gly-Sar). The uptake of [14C]Gly-Sar into HT1080 was time dependent. Kinetic analysis of the concentration dependence of the initial rate of [14C]Gly-Sar uptake showed that a carrier-mediated transport system with a K(m) of 11.4 +/- 3.3 mM and V(max) of 26.8 +/- 4.0 (nmol/15 min/mg protein) and a nonsaturable component (k(d) of 0.80 microl/15 min/mg protein) were responsible for the dipeptide uptake by HT1080 cells. The optimal pH for the maximal uptake was around 6.0. [14C]Gly-Sar uptake was inhibited by various di- and tripeptides and peptide-mimetic drugs, such as bestatin and cefadroxil. [14C]Gly-Sar uptake was not affected by the presence of amino acids or tetra- or pentapeptides. The uptake of cefadroxil was reduced significantly by unlabeled Gly-Sar. Moreover, Gly-Gly and Gly-Leu produced an increase in the apparent K(m) of the uptake of Gly-Sar without altering V(max). On the other hand, dipeptide uptake by IMR-90, which is a normal diploid cell line (not malignant), showed no saturable transport. These results suggest that HT1080 cells take up dipeptides via a pH-dependent transporter. This is the first report showing that a dipeptide transport system, which is similar but not identical to the well-characterized oligopeptide transporters PepT1 and PepT2, exists in fibroblast-derived tumor cells but not in normal fibroblasts. The present finding could be the basis of a novel strategy for the specific delivery of oligopeptide-mimetic anticancer drugs into tumor cells.
为了探究通过人类肿瘤细胞的转运系统靶向作用于肿瘤细胞的可行性，利用抗水解的甘氨酰肌氨酸（Gly-Sar），在人纤维肉瘤细胞系HT1080和人成纤维细胞系IMR-90中研究了二肽摄取。[14C]Gly-Sar进入HT1080细胞的摄取过程具有时间依赖性。对[14C]Gly-Sar摄取初始速率的浓度依赖性进行动力学分析表明，HT1080细胞摄取二肽是由一个K(m)为11.4±3.3 mM、V(max)为26.8±4.0（nmol/15分钟/毫克蛋白）的载体介导转运系统以及一个非饱和成分（k(d)为0.80微升/15分钟/毫克蛋白）负责的。最大摄取的最佳pH约为6.0。[14C]Gly-Sar摄取受到各种二肽和三肽以及肽模拟药物（如抑氨肽酶B和头孢羟氨苄）的抑制。[14C]Gly-Sar摄取不受氨基酸或四肽或五肽存在的影响。未标记的Gly-Sar可显著降低头孢羟氨苄的摄取。此外，甘氨酰甘氨酸和甘氨酰亮氨酸使Gly-Sar摄取的表观K(m)增加，而不改变V(max)。另一方面，正常二倍体细胞系（非恶性）IMR-90的二肽摄取未显示出饱和转运。这些结果表明，HT1080细胞通过一种pH依赖性转运体摄取二肽。这是首次报道在成纤维细胞来源的肿瘤细胞中存在一种与已充分表征的寡肽转运体PepT1和PepT2相似但不完全相同的二肽转运系统，而在正常成纤维细胞中不存在。目前的发现可能是将寡肽模拟抗癌药物特异性递送至肿瘤细胞的新策略的基础。
