Modrak David E, Cardillo Thomas M, Newsome Guy A, Goldenberg David M, Gold David V
Garden State Cancer Center, Center for Molecular Medicine and Immunology, Belleville, New Jersey 07109, USA.
Cancer Res. 2004 Nov 15;64(22):8405-10. doi: 10.1158/0008-5472.CAN-04-2988.
We have examined the mechanism by which sphingomyelin (SM) enhances chemotherapy in human pancreatic cancer cells, focusing on the correlation between ceramide metabolism and apoptosis. Dose response curves for gemcitabine in the absence or presence of 0.2 mg/mL SM provided IC(50) values of 78.3 +/- 13.7 and 13.0 +/- 3.0 nmol/L, respectively. The cytotoxic effect of the combined treatment was synergistic (combination index = 0.36). Using annexin-V staining, the percentage of apoptotic cells was 3.6 +/- 2.6% for the untreated cells, 6.5 +/- 3.8% for the 0.2 mg/mL SM-treated cells, and 19.9 +/- 12.9% for the 100 nmol/L gemcitabine-treated cells, but increased significantly to 42.1 +/- 12.7% with the combined treatment (P < 0.001, compared with gemcitabine-treated group). The percentage of cells losing mitochondrial membrane potential followed a similar trend. The ceramide content of untreated and gemcitabine-treated cells was not significantly different (0.46 +/- 0.29 and 0.59 +/- 0.34 pmol ceramide/nmole PO(4)). However, when 0.2 mg/mL SM was added, ceramide levels were 1.09 +/- 0.42 and 1.58 +/- 0.55 pmol ceramide/nmol PO(4), for the SM alone and SM with gemcitabine-treated cells, respectively (P = 0.038). Acidic SMase was activated by exposure to gemcitabine but not SM, whereas the activities of neutral SMase and glycosylceramide synthase did not change with either gemcitabine or SM. The data are consistent with gemcitabine-induced activation of acidic SMase and indicate that the addition of SM can yield increased production of ceramide, mitochondrial depolarization, apoptosis, and cell death. Because SM by itself is relatively nontoxic, addition of this lipid to agents that induce apoptosis may prove useful to enhance apoptosis and increase cytotoxicity in cancer cells.
我们研究了鞘磷脂（SM）增强人胰腺癌细胞化疗效果的机制，重点关注神经酰胺代谢与细胞凋亡之间的相关性。在不存在或存在0.2mg/mL SM的情况下，吉西他滨的剂量反应曲线分别给出IC(50)值为78.3±13.7和13.0±3.0nmol/L。联合治疗的细胞毒性作用具有协同性（联合指数=0.36）。使用膜联蛋白-V染色，未处理细胞的凋亡细胞百分比为3.6±2.6%，0.2mg/mL SM处理的细胞为6.5±3.8%，100nmol/L吉西他滨处理的细胞为19.9±12.9%，但联合治疗时显著增加至42.1±12.7%（与吉西他滨处理组相比，P<0.001）。丧失线粒体膜电位的细胞百分比呈现类似趋势。未处理和吉西他滨处理细胞的神经酰胺含量无显著差异（分别为0.46±0.29和0.59±0.34pmol神经酰胺/nmole PO(₄)）。然而，当加入0.2mg/mL SM时，单独SM处理的细胞和SM与吉西他滨联合处理的细胞的神经酰胺水平分别为1.09±0.42和1.58±0.55pmol神经酰胺/nmol PO(₄)（P=0.038）。酸性鞘磷脂酶通过暴露于吉西他滨而被激活，但不被SM激活，而中性鞘磷脂酶和糖基神经酰胺合酶的活性在吉西他滨或SM处理时均未改变。这些数据与吉西他滨诱导酸性鞘磷脂酶激活一致，并表明添加SM可导致神经酰胺生成增加、线粒体去极化、细胞凋亡和细胞死亡。由于SM本身相对无毒，将这种脂质添加到诱导细胞凋亡的药物中可能有助于增强癌细胞中的细胞凋亡并增加细胞毒性。
