Cabot M C, Giuliano A E, Han T Y, Liu Y Y
John Wayne Cancer Institute, Saint John's Health Center, Santa Monica, California 90404, USA.
Cancer Res. 1999 Feb 15;59(4):880-5.
Resistance to chemotherapy is the major cause of cancer treatment failure. Insight into the mechanism of action of agents that modulate multidrug resistance (MDR) is instrumental for the design of more effective treatment modalities. Here we show, using KB-V-1 MDR human epidermoid carcinoma cells and [3H]palmitic acid as metabolic tracer, that the MDR modulator SDZ PSC 833 (PSC 833) activates ceramide synthesis. In a short time course experiment, ceramide was generated as early as 15 min (40% increase) after the addition of PSC 833 (5.0 microM), and by 3 h, [3H]ceramide was >3-fold that of control cells. A 24-h dose-response experiment showed that at 1.0 and 10 microM PSC 833, ceramide levels were 2.5- and 13.6-fold higher, respectively, than in untreated cells. Concomitant with the increase in cellular ceramide was a progressive decrease in cell survival, suggesting that ceramide elicited a cytotoxic response. Analysis of DNA in cells treated with PSC 833 showed oligonucleosomal DNA fragmentation, characteristic of apoptosis. The inclusion of fumonisin B1, a ceramide synthase inhibitor, blocked PSC 833-induced ceramide generation. Assessment of ceramide mass by TLC lipid charring confirmed that PSC 833 markedly enhanced ceramide synthesis, not only in KB-V-1 cells but also in wild-type KB-3-1 cells. The capacity of PSC 833 to reverse drug resistance was demonstrated with vinblastine. Whereas each agent at a concentration of 1.0 microM reduced cell survival by approximately 20%, when PSC 833 and vinblastine were coadministered, cell viability fell to zero. In parallel experiments measuring ceramide metabolism, it was shown that the PSC 833/vinblastine combination synergistically increased cellular ceramide levels. Vinblastine toxicity, also intensified by PSC 833 in wild-type KB-3-1 cells, was as well accompanied by enhanced ceramide formation. These data demonstrate that PSC 833 has mechanisms of action in addition to P-glycoprotein chemotherapy efflux pumping.
对化疗的耐药性是癌症治疗失败的主要原因。深入了解调节多药耐药性（MDR）的药物作用机制有助于设计更有效的治疗方案。在此，我们使用KB-V-1多药耐药人表皮样癌细胞和[3H]棕榈酸作为代谢示踪剂，证明MDR调节剂SDZ PSC 833（PSC 833）可激活神经酰胺合成。在短期实验中，加入PSC 833（5.0 microM）后，最早在15分钟时就产生了神经酰胺（增加40%），到3小时时，[3H]神经酰胺是对照细胞的3倍以上。一项24小时剂量反应实验表明，在1.0 microM和10 microM的PSC 833作用下，神经酰胺水平分别比未处理细胞高2.5倍和13.6倍。随着细胞内神经酰胺的增加，细胞存活率逐渐下降，这表明神经酰胺引发了细胞毒性反应。对用PSC 833处理的细胞中的DNA分析显示出寡核小体DNA片段化，这是细胞凋亡的特征。加入神经酰胺合酶抑制剂伏马菌素B1可阻断PSC 833诱导的神经酰胺生成。通过TLC脂质炭化评估神经酰胺含量证实，PSC 833不仅在KB-V-1细胞中，而且在野生型KB-3-1细胞中均显著增强了神经酰胺合成。用长春碱证明了PSC 833逆转耐药性 的能力。虽然每种药物在浓度为1.0 microM时可使细胞存活率降低约20%，但当同时给予PSC 833和长春碱时，细胞活力降至零。在测量神经酰胺代谢的平行实验中，结果表明PSC 833/长春碱组合可协同增加细胞内神经酰胺水平。在野生型KB-3-1细胞中，长春碱的毒性也因PSC 833而增强，同时神经酰胺形成也增加。这些数据表明，PSC
833除了具有P-糖蛋白化疗外排泵作用机制外，还有其他作用机制。
