Multidrug-resistant human KB carcinoma cells are highly resistant to the protein phosphatase inhibitors okadaic acid and calyculin A. Analysis of potential mechanisms involved in toxin resistance.
Multidrug-resistant human KB carcinoma cells are highly resistant to the protein phosphatase inhibitors okadaic acid and calyculin A. Analysis of potential mechanisms involved in toxin resistance.
复制标题
多重耐药人 KB 癌细胞对蛋白磷酸酶抑制剂冈田酸和花萼蛋白 A 具有高度耐药性。分析毒素耐药性的潜在机制。
DOI：
10.1002/ijc.2910530225
复制
发表时间：
1993
影响因子：
6.4
通讯作者：
Kuo,JF
中科院分区：
文献类型：
--
作者：
Chambers,TC;Raynor,RL;Kuo,JF
在这项研究中，我们发现多药耐药（MDR）人KB-VI细胞对冈田酸和calyculin A的细胞毒性具有高度抗性，这两种毒素来自海绵，是I型和2A型蛋白磷酸酶（PP 1和PP 2A）的有效抑制剂。细胞毒性和集落形成试验表明，相对于亲本药物敏感性KB-3细胞，KB-VI细胞对冈田酸的耐药性高35倍，对calyculin A的耐药性高70倍。毒素的细胞毒性与以染色体分散和过度浓缩为特征的有丝分裂阻滞相关，KB-3细胞比KB-VI细胞更敏感，calyculin A比冈田酸更有效。KB-VI细胞对冈田酸和calyculin A的抗性被维拉帕米完全逆转，表明毒素可能通过P-糖蛋白（P-gp）转运。为了进一步评估与P-糖蛋白相互作用的可能性，这些毒素被用作[3 H]叠氮平光亲和标记P-糖蛋白的潜在调节剂。长春碱与[3 H]azidopine有效竞争P-gp光标记，calyculin A的效力比长春碱低100倍，冈田酸在浓度高达50 μM时不抑制光标记。为了确定耐药机制是否涉及毒素敏感性磷酸酶活性的差异，在两种细胞系的提取物中测定了活性，发现KB-VI细胞中的活性略高于KB-3细胞（1.6倍）。我们的研究结果表明，MDR KB‐VI细胞对这些磷酸酶抑制剂具有新的显著耐药性，并表明耐药性的主要机制可能涉及P-gp在明显不同于结合azidopine的位点的毒素转运。
In this study we show that multidrug‐resistant (MDR) human KB‐VI cells are highly resistant to the cytotoxicity of okadaic acid and calyculin A, 2 toxins from marine sponges that are potent inhibitors of type‐I and type‐2A protein phosphatases (PP1 and PP2A). Cytotoxicity and colony‐forming assays indicated that, relative to parental drug‐sensitive KB‐3 cells, KB‐VI cells are 35‐fold more resistant to okadaic acid and 70‐fold more resistant to calyculin A. Cytotoxicity of the toxins was associated with mitotic arrest characterized by chromosome scattering and over‐condensation, with KB‐3 cells being more sensitive than KB‐VI cells and calyculin A being more potent than okadaic acid. The resistance of KB‐VI cells to both okadaic acid and calyculin A was completely reversed by verapamil, suggesting that the toxins may be transported by P‐glycoprotein (P‐gp). To further assess the possibility of an interaction with P‐gp, the toxins were employed as potential modulators of the photoaffinity labeling of P‐gp by [3H]azidopine. Relative to vinblastine, which effectively competed with [3H]azidopine for P‐gp photolabeling, calyculin A was 100‐fold less potent and okadaic acid did not inhibit photolabeling at concentrations up to 50 μM. To determine whether the resistance mechanism involved differences in toxin‐sensitive phosphatase activity, the activity was assayed in extracts from both cell lines and found to be slightly higher (1.6‐fold) in KB‐VI than in KB‐3 cells. Our results demonstrate a novel, marked resistance of MDR KB‐VI cells to these phosphatase inhibitors and suggest that a major mechanism of resistance may involve toxin transport by P‐gp at sites apparently different from those which bind azidopine.