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1.
Resistance to chemotherapy in cancer is common. As gene expression profiling has been shown to anticipate chemotherapeutic resistance, we sought to identify cellular pathways associated with resistance to facilitate effective combination therapy. Gene set enrichment analysis was used to associate pathways with resistance in two data sets: the NCI-60 cancer cell lines deemed sensitive and resistant to specific chemotherapeutic agents (Adriamycin, cyclophosphamide, docetaxel, etoposide, 5-fluorouracil, paclitaxel, and topotecan) and a series of 40 lung cancer cell lines for which sensitivity to cisplatin and docetaxel was determined. Candidate pathways were further screened in silico using the Connectivity Map. The lead candidate pathway was functionally validated in vitro. Gene set enrichment analysis associated the matrix metalloproteinase, p53, methionine metabolism, and free pathways with cytotoxic resistance in the NCI-60 cell lines across multiple agents, but no gene set was common to all drugs. Analysis of the lung cancer cell lines identified the bcl-2 pathway to be associated with cisplatin resistance and the AKT pathway enriched in cisplatin- and docetaxel-resistant cell lines. Results from Connectivity Map supported an association between phosphatidylinositol 3-kinase/AKT and docetaxel resistance but did not support the association with cisplatin. Targeted inhibition of the phosphatidylinositol 3-kinase/AKT pathway with LY294002, in combination with docetaxel, resulted in a synergistic effect in previously docetaxel-resistant cell lines but not with cisplatin. These results support the use of a genomic approach to identify drug-specific targets associated with the development of chemotherapy resistance and underscore the importance of disease context in identifying these pathways.  相似文献   

2.
Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI) have poor efficacy in head and neck squamous carcinoma cells (HNSCC). Because the IGF-1 receptor (IGF1R) generates potent prosurvival signals and has been implicated in therapeutic resistance, its ability to induce resistance to EGFR-TKIs was studied in vitro. Five HNSCC cell lines showed reduced sensitivity to the EGFR-TKI gefitinib when the IGF1R was activated. In SCC-25 and Cal27 cells, gefitinib inhibited basal and EGF-stimulated EGFR, extracellular signal-regulated kinase (Erk), and Akt phosphorylation and reduced cell number. This correlated with initiation of apoptosis based on a 4-fold increase in PARP cleavage and a 2.5-fold increase in Annexin V positivity. The apoptotic response and reduction in cell number were blocked by IGF1R activation, which resulted in phosphorylation of both Erk and Akt. In both the cell lines, IGF1R-induced Erk, but not Akt, activation was eliminated by gefitinib. IGF1R-induced gefitinib resistance was unaffected by MAP/Erk kinase inhibition with U0126 but was partially impaired by inhibition of phosphoinositide-3-kinase with LY294002. The IGF1R-TKI PQ401 inhibited growth of SCC-25 and Cal27 cells alone and also acted synergistically with gefitinib. Thus, the IGF1R can make HNSCC cells resistant to EGFR-TKI treatment via a prosurvival mechanism. Of the 8 HNSCC tumor samples studied, all samples expressed the IGF1R and 5 showed detectable IGF1R phosphorylation, suggesting that this receptor may be relevant in vivo, and thus, combined EGFR/IGF1R inhibition may be necessary in some patients for effective targeted molecular therapy.  相似文献   

3.
Cisplatin resistance in human cancers   总被引:2,自引:0,他引:2  
Cancer chemotherapeutic agents primarily act by damaging cellular DNA directly or indirectly. Tumor cells, in contrast to normal cells, respond to cisplatin with transient gene expression to protect and/or repair their chromosomes. Repeated cisplatin treatments results in a stable resistant cell line with enhanced gene expression but lacking gene amplification for the proteins that will limit cisplatin cytotoxicity. Recently, several new human cell lines have been characterized for cisplatin resistance. These cell lines have led to a better understanding of the molecular and biochemical basis of cisplatin resistance. The c-fos proto-oncogene, a master switch for turning on other genes in response to a wide range of stimuli, has been shown to play an important role in cisplatin resistance both in vitro and in patients. Based on these studies, new strategies have been developed to circumvent and/or exploit clinical cisplatin resistance.  相似文献   

4.
Studies were undertaken to investigate acquired resistance to cisplatin in human ovarian cancer cells. The cell lines A2780 and A2780/CP70 were studied to assess their respective characteristics of drug accumulation and efflux, cytosolic inactivation of drug, and DNA repair. All experiments were performed using 1-h drug exposures. The A2780/CP70 cell line was 13-fold more resistant to cisplatin than A2780 cells. When studied at their respective IC50 doses, drug accumulation rates were similar for the two cell lines. However, the resistant cell line was twofold more efficient at effluxing drug, which was associated with reduced total drug accumulation for equivalent micromolar drug exposures. At equivalent levels of total cellular drug accumulation, the two cell lines formed the same levels of cisplatin-DNA damage, suggesting that cytosolic inactivation of drug does not contribute to the differential in resistance between these cell lines. Resistant cells were also twofold more efficient at repairing cisplatin-DNA lesions in cellular DNA and in transfected plasmid DNA. We conclude that in these paired cell lines, alterations in drug uptake/efflux and in DNA repair are the major contributing factors to acquired resistance to cisplatin.  相似文献   

5.
The broad nature of insulin resistant glucose metabolism in skeletal muscle of patients with type 2 diabetes suggests a defect in the proximal part of the insulin signaling network. We sought to identify the pathways compromised in insulin resistance and to test the effect of moderate exercise on whole-body and cellular insulin action. We conducted euglycemic clamps and muscle biopsies on type 2 diabetic patients, obese nondiabetics and lean controls, with and without a single bout of exercise. Insulin stimulation of the phosphatidylinositol 3-kinase (PI 3-kinase) pathway, as measured by phosphorylation of the insulin receptor and IRS-1 and by IRS protein association with p85 and with PI 3-kinase, was dramatically reduced in obese nondiabetics and virtually absent in type 2 diabetic patients. Insulin stimulation of the MAP kinase pathway was normal in obese and diabetic subjects. Insulin stimulation of glucose-disposal correlated with association of p85 with IRS-1. Exercise 24 hours before the euglycemic clamp increased phosphorylation of insulin receptor and IRS-1 in obese and diabetic subjects but did not increase glucose uptake or PI 3-kinase association with IRS-1 upon insulin stimulation. Thus, insulin resistance differentially affects the PI 3-kinase and MAP kinase signaling pathways, and insulin-stimulated IRS-1-association with PI 3-kinase defines a key step in insulin resistance.  相似文献   

6.
Because cytotoxic stress elicits various signaling pathways that may be implicated in cell survival or cell death, their alterations may have relevance in the development of platinum-resistant phenotype. Thus, in the present study, we investigated cell response to the epidermal growth factor receptor (EGFR) inhibitor gefitinib of ovarian carcinoma cell lines, including cells selected for resistance to cisplatin (IGROV-1/Pt1) and oxaliplatin (IGROV-1/OHP). Resistant sublines exhibited a marked decrease in sensitivity to gefitinib and resistance to apoptosis. Gefitinib was capable of inhibiting the phosphorylation of EGFR in all the studied cell lines. The Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) kinases, which act downstream of EGFR, were constitutively active in the three cell lines, but phospho-ERK1/2 levels were increased in the two resistant sublines. This feature was associated with reduced sensitivity to the MEK1/2 inhibitor U0126. Pretreatment of resistant cells with U0126 resulted in restoration of sensitivity to gefitinib. Gefitinib was more effective in inhibiting ERK1/2 and Akt phosphorylation in IGROV-1 cells than in IGROV-1/OHP and IGROV-1/Pt1 cells. Phospho-p38 was up-regulated in the resistant sublines, indicating the concomitant activation of distinct mitogen-activated protein kinases. The up-regulation of phospho-p38 was associated with a peculiar localization of EGFR, which, in resistant sublines, was mainly internalized. In conclusion, our results indicate that the development of resistance to platinum drugs is associated with multiple alterations including deregulation of survival pathways activated by EGFR resulting in a reduced cellular response to gefitinib.  相似文献   

7.
In acute myeloid leukemia (AML), receptor tyrosine kinase ligands promote growth and survival and contribute to AML-associated marrow neoangiogenesis. We have tested simultaneous inhibition of vascular endothelial growth factor, fibroblast growth factor, and platelet-derived growth factor receptor signaling by novel indolinone derivatives using 14 myeloid, including 11 human leukemic, cell lines. Compounds inhibited colony formation of all cell lines in a dose-dependent fashion. Inhibitory concentrations for 50% of the colony formation/survival (IC50) for BIBF1000 were <100 nmol/L for 3 of 11, 相似文献   

8.
目的:探讨CML耐药细胞株K562/G耐药相关的FOXO3/Bcl-6信号通路及相关microRNA(miRNA)机制。方法:采用MTT法检测伊马替尼对K562/G的耐药性;采用Western blot法检测耐药株和敏感株细胞中FOXO3和Bcl-6蛋白的表达情况,实时荧光定量PCR检测FOXO3及Bcl-6 mRNA的表达。运用miRNA芯片技术筛查K562与K562/G细胞之间差异表达的miRNA,进一步筛选FOXO3/Bcl-6信号通路的靶向miRNA。结果:K562/G较K562细胞株的FOXO3及Bcl-6蛋白表达均明显升高(P<0.01);Bcl-6 mRNA的表达水平无明显差异,而K562/G细胞中FOXO3 mRNA表达明显升高(P<0.05)。miRNA芯片结果显示,K562/G与K562细胞之间有109条miRNA存在显著的差异,其中上调miRNA为81个,下调miRNA有28个。经生物信息学反向预测,其中miR-6718-5p、miR-5195-5p、miR-4711-3p、miR-4763-5p、miR-4664-5p和miR-3176与该信号通路相关。结论:伊马替尼耐药与FOXO3/Bcl-6信号通路相关,并且K562/G与K562细胞存在miRNA表达显著差异。  相似文献   

9.
Although >66% of melanomas harbor activating mutations in BRAF and exhibit constitutive activity in the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK)/extracellular signal-regulated kinase signaling pathway, it is unclear how effective MEK inhibition will be as a sole therapeutic strategy for melanoma. We investigated the anticancer activity of MEK inhibition in a panel of cell lines derived from radial growth phase (WM35) and vertical growth phase (WM793) of primary melanomas and metastatic melanomas (1205Lu, 451Lu, WM164, and C8161) in a three-dimensional spheroid model and found that the metastatic lines were completely resistant to MEK inhibition (U0126 and PD98059) but the earlier stage cell lines were not. Similarly, these same metastatic melanoma lines were also resistant to inhibitors of the phosphatidylinositol 3-kinase/Akt pathway (LY294002 and wortmannin). Under adherent culture conditions, the MEK inhibitors blocked growth through the induction of cell cycle arrest and up-regulation of p27, but this was readily reversible following inhibitor washout. However, when the phosphatidylinositol 3-kinase and MEK inhibitors were combined, the growth and invasion of the metastatic melanoma three-dimensional spheroids were blocked. Taken together, these results suggest that the most aggressive melanomas are resistant to strategies targeting one signaling pathway and that multiple signaling pathways may need to be targeted for maximal therapeutic efficacy. It is further suggested that BRAF mutational status is not predictive of response to MEK inhibition under three-dimensional culture conditions.  相似文献   

10.
Interleukin (IL)-7 is essential for normal T cell development. Previously, we have shown that IL-7 increases viability and proliferation of T cell acute lymphoblastic leukemia (T-ALL) cells by up-regulating Bcl-2 and down-regulating the cyclin-dependent kinase inhibitor p27kip1. Here, we examined the signaling pathways via which IL-7 mediates these effects. We investigated mitogen-activated protein kinase (MEK)-extracellular signal-regulated kinase (Erk) and phosphatidylinositol-3-kinase (PI3K)-Akt (protein kinase B) pathways, which have active roles in T cell expansion and have been implicated in tumorigenesis. IL-7 induced activation of the MEK-Erk pathway in T-ALL cells; however, inhibition of the MEK-Erk pathway by the use of the cell-permeable inhibitor PD98059, did not affect IL-7-mediated viability or cell cycle progression of leukemic cells. IL-7 induced PI3K-dependent phosphorylation of Akt and its downstream targets GSK-3, FOXO1, and FOXO3a. PI3K activation was mandatory for IL-7-mediated Bcl-2 up-regulation, p27kip1 down-regulation, Rb hyperphosphorylation, and consequent viability and cell cycle progression of T-ALL cells. PI3K signaling was also required for cell size increase, up-regulation of CD71, expression of the glucose transporter Glut1, uptake of glucose, and maintenance of mitochondrial integrity. Our results implicate PI3K as a major effector of IL-7-induced viability, metabolic activation, growth and proliferation of T-ALL cells, and suggest that PI3K and its downstream effectors may represent molecular targets for therapeutic intervention in T-ALL.  相似文献   

11.
Cyclin D1 is essential for Neu-induced cell growth and is induced by growth factors through Ras-dependent and Ras-independent signaling pathways (1). Because flavopiridol, a novel flavanoid cyclin-cyclin-dependent kinase inhibitor, may function through Ras-dependent and/or -independent pathways, we hypothesized that treatment of breast cancer cells with inhibitors of Neu signaling and flavopiridol might synergize to inhibit proliferation. Human breast cancer cell lines, which express high levels of endogenous Neu receptor, were treated with the anti-Neu antibody, trastuzumab, together with flavopiridol and subject to MTT assay. Cell lines were assayed for alterations in cell cycle by fluorescence-activated cell sorter and signaling proteins by Western blot. Flavopiridol and trastuzumab synergistically inhibited DNA synthesis, cellular proliferation, and contact-dependent growth. Cytotoxic synergy was observed independent of the sequence of addition of the two drugs to cultured cells. In SKBR3 cells, a combination of trastuzumab and flavopiridol inhibited the Ras-MAPK-Akt pathway, decreased cyclin D1 abundance, and kinase activity to a greater extent than either drug alone. Compared with single-agent treatment, combination treatment selectively inhibited Akt and pRB phosphorylation. Cytotoxic synergy was observed with flavopiridol plus LY294002 (selective phosphatidylinositol 3-kinase inhibitor) but not with PD98059 (selective mitogen-activated protein kinase kinase 1 inhibitor) suggesting that Akt inhibition may be important in synergy. Zinc-induced overexpression of cyclin D1 in T-47D deltaMTcycD1 cells were more resistant to drug-induced cell death when compared with vector-transfected T-47D deltaMT cells. Cyclin D1 overexpression reverses drug treatment induced cell cycle arrest in SKBR3 cells. Flavopiridol and trastuzumab yield cytotoxic synergy in human breast cancer cells overexpressing Neu. Cyclin D1 overexpression results in combination drug resistance. In addition, inhibition of Akt may prove to be a useful therapeutic strategy in combination with flavopiridol.  相似文献   

12.
Organ preservation protocols in head and neck squamous cell carcinoma (HNSCC) are limited by tumors that fail to respond. We observed that larynx preservation and response to chemotherapy is significantly associated with p53 overexpression, and that most HNSCC cell lines with mutant p53 are more sensitive to cisplatin than those with wild-type p53. To investigate cisplatin resistance, we studied two HNSCC cell lines, UM-SCC-5 and UM-SCC-10B, and two resistant sublines developed by cultivation in gradually increasing concentrations of cisplatin. The cisplatin-selected cell lines, UM-SCC-5PT and UM-SCC-10BPT, are 8 and 1.5 times more resistant to cisplatin than the respective parental cell lines, respectively. The parental lines overexpress p53 and contain p53 mutations but the cisplatin-resistant cell lines do not, indicating that cells containing mutant p53 were eliminated during selection. Bcl-x(L) expression increased in the cisplatin-resistant lines relative to the parental lines, whereas Bcl-2 expression was high in the parental lines and decreased in the cisplatin-resistant lines. Thus, cisplatin selected for wild-type p53 and high Bcl-x(L) expression in these cells. We tested a small-molecule BH3 mimetic, (-)-gossypol, which binds to the BH3 domain of Bcl-2 and Bcl-x(L), for activity against the parental and cisplatin-resistant cell lines. At physiologically attainable levels, (-)-gossypol induces apoptosis in 70% to 80% of the cisplatin-resistant cells but only in 25% to 40% of the parental cells. Thus, cisplatin-resistant cells seem to depend on wild-type p53 and Bcl-x(L) for survival and BH3 mimetic agents, such as (-)-gossypol, may be useful adjuncts to overcome cisplatin resistance in HNSCC.  相似文献   

13.
Recombinant human erythropoietin is widely used to treat anemia associated with cancer and with the myelosuppressive effects of chemotherapy, particularly platinum-based regimens. Erythropoietin is the principal regulator of erythroid cell proliferation, differentiation, and apoptosis. Recently, the antiapoptotic and proliferative effects of erythropoietin on nonhematopoietic cells were also established. We now show the effect of erythropoietin treatment on the response of A2780 and SKOV3 ovarian carcinoma cell lines to photodynamic therapy (PDT) using hypericin. SKOV3 exhibited an increased resistance to hypericin when cells were treated with erythropoietin. This resistance was reversed by treatment of SKOV3 cells with the specific Janus kinase 2 kinase inhibitor AG490 or the tyrosine kinase inhibitor genistein. These results support a role for the specific erythropoietin-induced Janus kinase 2/STAT signal transduction pathway in PDT resistance. Evidence of erythropoietin signaling was obtained by the demonstration of Akt phosphorylation in both A2780 and SKOV3 cells. Erythropoietin-treated SKOV3 cells exhibited decreased apoptosis induced by hypericin, an effect that was blocked by the phosphoinositide 3-kinase/Akt inhibitor wortmannin. These results may have important implications for ovarian cancer patients undergoing PDT and receiving erythropoietin.  相似文献   

14.
Hepatocellular carcinoma (HCC) is one of the most common potentially lethal human malignancies worldwide. Sorafenib, a tyrosine kinase inhibitor, was recently approved by the United States Food and Drug Administration for HCC. In this study, we established two sorafenib-resistant HCC cell lines from Huh7, a human HCC cell line, by long-term exposure of cells to sorafenib. Sorafenib induced significant apoptosis in Huh7 cells; however, Huh7-R1 and Huh7-R2 showed significant resistance to sorafenib-induced apoptosis at the clinical relevant concentrations (up to 10 μM). Thorough comparisons of the molecular changes between Huh7 and resistant cells showed that the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway played a significant role in mediating acquired resistance to sorafenib in Huh7-R1 and Huh7-R2 cells. Phospho-Akt and p85 (a regulatory subunit of PI3K) were up-regulated, whereas tumor suppressor phosphatase and tensin homolog were down-regulated in these resistant cells. In addition, ectopic expression of constitutive Akt in Huh7 demonstrated similar resistance to sorafenib. The knockdown of Akt by RNA interference reversed resistance to sorafenib in Huh7-R1 cells, indicating the importance of Akt in drug sensitivity. Furthermore, the combination of 8-[4-(1-aminocyclobutyl)phenyl]-9-phenyl-1,2,4-triazolo[3,4-f][1,6]naphthyridin-3(2H)-one dihydrochloride (MK-2206), a novel allosteric Akt inhibitor, and sorafenib restored the sensitivity of resistant cells to sorafenib-induced apoptosis. In conclusion, activation of PI3K/Akt signaling pathway mediates acquired resistance to sorafenib in HCC, and the combination of sorafenib and MK-2206, an Akt inhibitor, overcomes the resistance at clinical achievable concentrations.  相似文献   

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17.
PRIMA-1 (p53 reactivation and induction of massive apoptosis) is a chemical compound that was originally identified as a selective mutant p53-dependent growth suppressor by screening a library of low-molecular-weight compounds. However, its mechanism of action is unknown. In this study, we examined toxicity of PRIMA-1 to three premalignant human colorectal adenoma cell lines (RG/C2, BR/C1, and AA/C1) and four colorectal carcinoma cell lines (DLD-1, SW480, LOVO, and HCT116) and its mechanism of action. It selectively induced apoptosis only in the mutant p53 premalignant and malignant colon cell lines, but was not toxic to the wild-type p53 premalignant and malignant colon cell lines. Using stable transfectants of temperature-sensitive p53 mutant Ala(143) in null p53 H1299 lung cancer cells, we found that PRIMA-1 induced significantly more apoptosis in cells with mutant p53 conformation (37 degrees C) than the wild-type p53 conformation (32.5 degrees C). Cell cycle analysis indicated that its inhibition of cell growth was correlated with induction of G(2) arrest. Western blot analysis showed PRIMA-1 increased p21 and GADD45 expression selectively in the mutant p53 cells. However, Fas, Bcl-2 family proteins, and caspases were not involved in PRIMA-1-induced cell death. The c-Jun-NH(2)-kinase (JNK) inhibitor SP 600125, but not p38 mitogen-activated protein kinase inhibitor SB 203580 or extracellular signal-regulated kinase inhibitor PD 98059, blocked PRIMA-1-induced apoptosis. Transfection with a dominant-negative phosphorylation mutant JNK, but not a dominant-negative p38 or wild-type JNK, inhibited PRIMA-1-induced cell death, suggesting that the JNK pathway plays an important role in PRIMA-1-induced apoptosis. PRIMA-1 is a highly selective small molecule toxic to p53 mutant cells and may serve as a prototype for the development of new p53-targeting agents for therapy of premalignant and malignant cells.  相似文献   

18.
Malignant mesothelioma is an aggressive cancer with no known cure, which has become a therapeutic challenge. Onconase is one of few chemotherapeutic agents that have been studied in patients with malignant mesothelioma that has the advantage of low toxicity and limited side effects. Here, we evaluate the effect of Onconase on killing of malignant mesothelioma cells and how the phosphatidylinositol 3-kinase/AKT (PI3-K/AKT) survival pathway influences this effect. Our results show that Onconase induces apoptosis in malignant mesothelioma cell lines and that this effect is tumor cell specific. Malignant mesothelioma cell lines with the highest AKT activation, which correlated with the presence of the SV40 large and small T antigen (SV40+), were the most resistant to the drug. Finally, a cooperative effect was observed between small molecule inhibitors of PI3-K and Onconase in the killing of malignant mesothelioma cells. Our results suggest that kinase screening of individual malignant mesotheliomas for endogenous levels of activated PI3-K/AKT may be predictive of the efficacy of Onconase and possibly other chemotherapeutic agents.  相似文献   

19.
Several studies in the last years evidenced that deregulation of proapoptotic and antiapoptotic pathways are key players in the onset and maintenance of chemoresistance in advanced ovarian cancers. To characterize the signaling events and molecules involved in the acquisition of cisplatin resistance, we used the human ovarian cancer cell line A2780 and its derivative cisplatin-resistant subline A2780 CIS. We found that the mitochondrial intrinsic apoptotic pathway, induced by cis-dichlorodiammineplatinum (CDDP) in A2780 wild-type cells, was compromised in the resistant subline CIS. The analysis of expression of proteins involved in mitochondria-dependent apoptosis revealed a role of Bax and p73 but not p53. Indeed, we found that CDDP treatment induced the up-regulation of p53 in both sensitive and resistant A2780 cell lines. By contrast, p73 and Bax expressions were compromised in resistant cells. Pretreatment of resistant A2780 CIS cells with the histone deacetylase inhibitor trichostatin A overcomes apoptosis resistance to CDDP by restoring both p73 and Bax but not p53 expression. Altogether, these data indicate that p73, but not p53, is involved in the regulation of apoptosis susceptibility to cisplatin in A2780 ovarian cancer cells and evidence a key contribution of histone deacetylase activation in the acquisition of chemotherapy resistance in human ovarian cancer cells.  相似文献   

20.
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