Determinants of differential doxorubicin sensitivity between SCLC and NSCLC

详细信息    查看全文

• 作者：Sharad S. Singhal ; Dilki Wickramarachchi ; Jyotsana Singhal ; Sushma Yadav ; Yogesh C. Awasthi and Sanjay Awasthi
• 关键词：Protein-kinase-C ; Lung cancer ; Doxorubicin ; Drug-resistance ; RLIP76 ; siRNA
• 刊名：FEBS Letters
• 出版年：2006
• 出版时间：17 April 2006
• 年：2006
• 卷：580
• 期：9
• 页码：2258-2264
• 全文大小：192 K

文摘

Doxorubicin (DOX) transport activity of Ral-interacting protein (RLIP76) in non-small cell lung cancer (NSCLC) is approximately twice that of in small cell lung cancer (SCLC). Since protein-kinase-C (PKC)α mediated phosphorylation of RLIP76 causes doubling of the specific activity of RLIP76, and NSCLC cells are known to have greater PKCα activity, we examined the contribution of PKC mediated phosphorylation of RLIP76 towards intrinsic DOX-resistance in human NSCLC. Expression of a deletion mutant RLIP76^{delPKCα-sites} followed by depletion of the wild-type RLIP76 using a siRNA targeted at one of the deleted regions resulted in generation of cells expressing only the mutant protein, which could not be phosphorylated by PKCα. DOX-transport activity of the mutant RLIP76 purified from NSCLC and SCLC was similar and comparable to that of RLIP76 purified from the wild-type SCLC. However, this activity was significantly lower than that of RLIP76 purified from the wild-type NSCLC. After siRNA mediated depletion of PKCα, DOX-transport activities of RLIP76 purified from SCLC and NSCLC were indistinguishable. Depletion of PKCα inhibited the growth of NSCLC more than SCLC cells (70 ± 3 % vs. 43 ± 5 % , respectively). PKCα-depletion lowered the IC₅₀ of NSCLC cell lines for DOX to the same level as that observed for SCLC. RLIP76^−/− mouse embryonic fibroblasts (MEFs) were significantly more sensitive to DOX as compared with RLIP76^+/+ MEFs (IC₅₀ 25 vs. 125 nM, respectively). However, PKCα-depletion did not affect DOX-cytotoxicity towards RLIP76^−/− MEFs, as opposed to RLIP76^+/+ MEFs which were sensitized by 2.2-fold. These results demonstrate that RLIP76 is a primary determinant of DOX-resistance, and that PKCα mediated accumulation defect and DOX-resistance in NSCLC is primarily due to differential phosphorylation of RLIP76 in SCLC and NSCLC.