Inhibition of potentially anti-apoptotic proteins by antisense protein kinase C-α (Isis 3521) and antisense bcl-2 (G3139) phosphorothioate oligodeoxynucleotides:: Relationship to the decreased viability of T24 bladder and PC3 prostate cancer cells
Inhibition of potentially anti-apoptotic proteins by antisense protein kinase C-α (Isis 3521) and antisense bcl-2 (G3139) phosphorothioate oligodeoxynucleotides:: Relationship to the decreased viability of T24 bladder and PC3 prostate cancer cells
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DOI：
10.1124/mol.60.6.1296
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发表时间：
2001-12-01
影响因子：
3.6
通讯作者：
Stein, CA
中科院分区：
文献类型：
--
作者：
Benimetskaya, L;Miller, P;Stein, CA
ISIS 3521和G3139分别是针对蛋白激酶C(PKC)-α和bcl2 mRNAs的20和18聚硫代寡核苷酸。用Isis 3521的全长和3‘端截断突变体处理T24膀胱和PC3前列腺癌细胞，可下调PKC-a蛋白和mRNA的表达。然而，在15-聚体和更短的水平上，不再观察到mRNA表达的下调。此外，通过3-(4，5-二甲基噻唑-2-基)2，5-二苯基四氮唑溴化法测定，这些较短的低聚物的细胞存活率不会随着紫杉醇浓度的增加而降低。这些观察结果不仅表明PKC-α蛋白的表达可以被RNaseH依赖和非依赖的机制下调，而且PKC-α的下调本身不足以使细胞对化疗敏感。G3139下调BCL-2蛋白和mRNA的表达，也下调PKC-α蛋白和mRNA的表达，但不下调PKC-β1、-epsilon或-Zeta的表达。然而，PKC-α和bcl-2的下调并不相关。当使用载体Eufetin 5时，在50 nM的寡核苷酸浓度下，只有bcl2在T24和PC3细胞中下调。100 nm时，bcl2和PKC-α的表达均下调，只有在该浓度下才能观察到对紫杉醇和卡铂的“化疗增敏”。相反，bcl2的下调似乎与relA的下调有关(P65)。然而，这对于化学增敏也是不够的，即使它会导致核因子-kappaB控制下的基因表达的丧失，并导致细胞从塑料表面分离。这些结果强调了细胞内对抗肿瘤药物启动化疗增敏要求的复杂性。
Isis 3521 and G3139 are 20- and 18-mer phosphorothioate oligonucleotides, respectively, targeted to the protein kinase C (PKC)-alpha and bcl-2 mRNAs. Treatment of T24 bladder and PC3 prostate carcinoma cells with full-length and 3'-truncation mutants of Isis 3521 causes down-regulation of PKC-a protein and mRNA. However, at the level of a 15-mer and shorter, downregulation of mRNA expression is no longer observed. Further, no diminution in cellular viability, as measured by 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide assay, in response to increasing concentrations of paclitaxel, can be observed for these shorter oligomers. These observations not only indicate that PKC-alpha protein expression can be down-regulated by both RNase H-dependent and -independent mechanisms but also that down-regulation of PKC-alpha is insufficient by itself to "chemosensitize" cells. G3139, which down-regulates bcl-2 protein and mRNA expression, also down-regulates PKC-alpha protein and mRNA expression but not that of PKC-beta1, -epsilon, or -zeta. However, the down-regulation of PKC-alpha and bcl-2 are not linked. When the carrier Eufectin 5 is employed, only bcl-2 is down-regulated in both T24 and PC3 cells at 50 nM oligonucleotide concentration. At 100 nM, both bcl-2 and PKC-alpha expression are down-regulated, and only at this concentration can "chemosensitization" to paclitaxel and carboplatin be observed. In contrast, the down-regulation of bcl-2 seems to be linked with that of RelA (p65). However, this too is also not sufficient for chemosensitization, even though it leads to the loss of expression of genes under the putative control of nuclear factor-kappaB and to detachment of the cells from plastic surfaces. These results underscore the complexity of the intracellular requirements for the initiation of chemosensitization to anti-neoplastic agents.