Seidenfeld J, Sprague W S
Department of Pharmacology, Northwestern University Medical School, Chicago, Illinois 60611.
Cancer Res. 1990 Feb 1;50(3):521-6.
We have reported that 2-difluoromethylornithine (DFMO)-induced polyamine (PA) depletion sensitized five chloroethylnitrosourea (CENU)-resistant, O6-alkylguanine repair-proficient (Mer+) human tumor cell lines to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), but failed to alter BCNU efficacy in a single CENU-sensitive, repair-deficient (Mer-) line. Further, alkaline elution assays of DNA interstrand cross-links (ISC) found no BCNU-induced ISC in either PA-depleted or control Mer+ cells, suggesting that targets other than ISC may be involved in the DFMO/BCNU drug interaction. To verify that DFMO-induced enhancement of BCNU action segregates with Mer phenotype, we tested three additional Mer- lines for effects of DFMO pretreatment on BCNU efficacy. We found no potentiation of BCNU by PA depletion in any of our human Mer- lines. We also used streptozotocin (STZ) to deplete the repair capacity of Mer+ cell lines, thus converting their BCNU sensitivity to near that of Mer- cells. Combined pretreatment with DFMO then STZ did enhance BCNU cell kill relative to STZ pretreatment alone. Exogenous putrescine restored BCNU sensitivity of (DFMO plus STZ)-pretreated cells to that of cells pretreated with STZ alone. Measurements of O6-alkylguanine DNA alkyltransferase activity verified that in at least one of the Mer+ lines (HT-29), STZ did deplete repair capacity to below detectable limits. These results suggest that in HT-29 cells, STZ and DFMO probably act via differing mechanisms to potentiate BCNU. Our observations also imply that targets for CENUs may differ between Mer+ and Mer- cells, with importance of ISC possibly limited to Mer- cells. Our data further suggest that PA depletion may potentiate CENUs only at targets critical in Mer+ cells. We also noted that 48-h treatments with DFMO markedly reduced clonogenicity of Mer- cells. Exogenous putrescine restored Mer- cell survival after DFMO to near that of controls. In contrast, Mer+ cells showed little, if any, effect of DFMO treatment on plating efficiency. These results suggest that PA depletion may be cytocidal to some Mer- cells.
我们曾报道，二氟甲基鸟氨酸（DFMO）诱导的多胺（PA）耗竭使五种对氯乙基亚硝脲（CENU）耐药、O6 - 烷基鸟嘌呤修复 proficient（Mer +）的人肿瘤细胞系对1,3 - 双（2 - 氯乙基）-1 - 亚硝脲（BCNU）敏感，但未能改变BCNU对单一CENU敏感、修复缺陷（Mer -）细胞系的疗效。此外，DNA链间交联（ISC）的碱性洗脱分析发现，在PA耗竭的或对照的Mer +细胞中均未检测到BCNU诱导的ISC，这表明ISC以外的靶点可能参与了DFMO/BCNU药物相互作用。为了验证DFMO诱导的BCNU作用增强与Mer表型相关，我们测试了另外三种Mer -细胞系，观察DFMO预处理对BCNU疗效的影响。我们发现，在任何一种人Mer -细胞系中，PA耗竭均未增强BCNU的作用。我们还使用链脲佐菌素（STZ）来耗尽Mer +细胞系的修复能力，从而使其对BCNU的敏感性接近Mer -细胞。相对于单独的STZ预处理，联合使用DFMO然后STZ预处理确实增强了BCNU对细胞的杀伤作用。外源性腐胺使（DFMO加STZ）预处理的细胞对BCNU的敏感性恢复到单独用STZ预处理的细胞的水平。对O6 - 烷基鸟嘌呤DNA烷基转移酶活性的测量证实，在至少一种Mer +细胞系（HT - 29）中，STZ确实将修复能力耗尽至低于可检测水平。这些结果表明，在HT - 29细胞中，STZ和DFMO可能通过不同机制增强BCNU的作用。我们的观察结果还意味着，CENUs的靶点在Mer +和Mer -细胞之间可能不同，ISC的重要性可能仅限于Mer -细胞。我们的数据进一步表明，PA耗竭可能仅在Mer +细胞中的关键靶点上增强CENUs的作用。我们还注意到，用DFMO处理48小时显著降低了Mer -细胞的克隆形成能力。外源性腐胺使DFMO处理后的Mer -细胞存活率恢复到接近对照水平。相比之下，Mer +细胞对DFMO处理的平板效率几乎没有影响（如果有影响的话也很小）。这些结果表明，PA耗竭可能对一些Mer -细胞具有细胞毒性。
