The role of copper transporter ATP7A in platinum-resistance of esophageal squamous cell cancer (ESCC).
ABSTRACT: Purpose: Platinum derivatives, such as cisplatin (DDP), carboplatin and oxaliplatin, are widely used components of modern cancer chemotherapy including esophageal squamous cell cancer (ESCC). However, their roles are limited by the impact of intrinsic/acquired resistance mechanisms on tumor responses. Recent studies have shown that the mammalian copper transporters CTR1, ATP7A and ATP7B are involved in cisplatin-resistance to some cancers. Methods: The cytotoxicities of DDP in different cell lines were determined using the MTT assay. To determine whether knockdown the expression of ATP7A could reverse the platinum-resistance of EC109/DDP cells or not, we used RNA interference system to explore the role of ATP7A in platinum resistance. Results: We found that DDP-resistant cell sublines EC109/DDP (8.490 folds) showed cross-resistance to carboplatin (5.27 folds) and oxaliplatin (4.12 folds). ATP7A expressions in DDP-resistant cell sublines (EC109/DDP) were much higher than DDP-sensitive cell lines (EC109) at both mRNA and protein levels. ATP7A targeted small interfering RNA duplex at 100nM final concentration added into DDP-resistant cancer cells (EC109/DDP) markedly inhibited the expression of ATP7A as determined by Western blot (83.0%) and partially reversed DDP-resistance (37.09%), moreover, it also increased cell apoptosis at different DDP concentrations. Conclusions: These findings indicate that ATP7A high expression plays an important role in platinum-resistance of ESCC. This study sheds light on platinum resistance in ESCC patients and may have implications for therapeutic reversal of drug resistance.
Project description:The Sec61 protein translocon is a multimeric complex that transports proteins across lipid bilayers. We discovered that the Sec61? subunit modulates cellular sensitivity to chemotherapeutic agents, particularly the platinum drugs. To investigate the mechanism, expression of Sec61? was constitutively knocked down in 2008 ovarian cancer cells. Sec61? knockdown (KD) resulted in 8-, 16.8-, and 9-fold resistance to cisplatin (cDDP), carboplatin, and oxaliplatin, respectively. Sec61? KD reduced the cellular accumulation of cDDP to 67% of that in parental cells. Baseline copper levels, copper uptake, and copper cytotoxicity were also reduced. Because copper transporters and chaperones regulate platinum drug accumulation and efflux, their expression in 2008 Sec61?-KD cells was analyzed; ATP7A was found to be 2- to 3-fold overexpressed, whereas there was no change in ATP7B, ATOX1, CTR1, or CTR2 levels. Cells lacking ATP7A did not exhibit increased cDDP resistance upon knockdown of Sec61?. Sec61?-KD cells also exhibited altered ATP7A cellular distribution. We conclude that Sec61? modulates the cytotoxicity of many chemotherapeutic agents, with the largest effect being on the platinum drugs. This modulation occurs through effects of Sec61? on the expression and distribution of ATP7A, which was shown previously to control platinum drug sequestration and cytotoxicity.
Project description:Transporters are important mediators of specific cellular uptake and thus, not only for effects, but also for side effects, metabolism, and excretion of many drugs such as cisplatin. Cisplatin is a potent cytostatic drug, whose use is limited by its severe acute and chronic nephro-, oto-, and peripheral neurotoxicity. For this reason, other platinum derivatives, such as carboplatin and oxaliplatin, with less toxicity but still with antitumoral action have been developed. Several transporters, which are expressed on the cell membranes, have been associated with cisplatin transport across the plasma membrane and across the cell: the copper transporter 1 (Ctr1), the copper transporter 2 (Ctr2), the P-type copper-transporting ATPases ATP7A and ATP7B, the organic cation transporter 2 (OCT2), and the multidrug extrusion transporter 1 (MATE1). Some of these transporters are also able to accept other platinum derivatives as substrate. Since membrane transporters display a specific tissue distribution, they can be important molecules that mediate the entry of platinum derivatives in target and also nontarget cells possibly mediating specific effects and side effects of the chemotherapeutic drug. This paper summarizes the literature on toxicities of cisplatin compared to that of carboplatin and oxaliplatin and the interaction of these platinum derivatives with membrane transporters.
Project description:Platinum drugs are among the most effective anticancer agents, but their mode of action is still not fully understood. We therefore carried out a systematic investigation on the cellular activities of cisplatin, carboplatin and oxaliplatin in A498 kidney cancer cells. Cytotoxicity was higher for cisplatin and oxaliplatin compared to carboplatin, with induction of apoptosis as the preferred mode of cell death. Gene expression profiling displayed modulation of genes related to DNA damage response/repair, cell cycle regulation and apoptosis which was more pronounced upon oxaliplatin treatment. Furthermore, repression of specific DNA repair genes was restricted to oxaliplatin. Transcriptional level observations were further analyzed on the functional level. Uptake studies revealed low intracellular platinum accumulation and DNA platination upon carboplatin treatment. Removal of overall DNA platination was comparable for the three drugs. However, no processing of oxaliplatin-induced interstrand crosslinks was observed. Cisplatin and carboplatin influenced cell cycle distribution comparably, while oxaliplatin had no effect. Altogether, we found a similar mode of action for cisplatin and carboplatin, while the activity of oxaliplatin appeared to differ. This might be clinically relevant as due to the difference in mode of action oxaliplatin could be active in tumors which show resistance towards cisplatin and carboplatin.
Project description:Cisplatin, carboplatin, and oxaliplatin are widely used anticancer drugs. Their efficacy is strongly reduced by development of cell resistance. Down-regulation of CTR1 and up-regulation of the Cu-ATPases, ATP7A and ATP7B, have been associated to augmented drug resistance. To gain information on translocation of Pt drugs by human Cu-ATPases, we performed electrical measurements on the COS-1 cell microsomal fraction, enriched with recombinant ATP7A, ATP7B, and selected mutants, and adsorbed on a solid supported membrane. The experimental results indicate that Pt drugs activate Cu-ATPases and undergo ATP-dependent translocation in a fashion similar to that of Cu. We then used NMR spectroscopy and ESI-MS to determine the binding mode of these drugs to the first N-terminal metal-binding domain of ATP7A (Mnk1).
Project description:The formation of acquired drug resistance is a major reason for the failure of anti-cancer therapies after initial response. Here, we introduce a novel model of acquired oxaliplatin resistance, a sub-line of the non-MYCN-amplified neuroblastoma cell line SK-N-AS that was adapted to growth in the presence of 4000 ng/mL oxaliplatin (SK-N-ASrOXALI4000). SK-N-ASrOXALI4000 cells displayed enhanced chromosomal aberrations compared to SK-N-AS, as indicated by 24-chromosome fluorescence in situ hybridisation. Moreover, SK-N-ASrOXALI4000 cells were resistant not only to oxaliplatin but also to the two other commonly used anti-cancer platinum agents cisplatin and carboplatin. SK-N-ASrOXALI4000 cells exhibited a stable resistance phenotype that was not affected by culturing the cells for 10 weeks in the absence of oxaliplatin. Interestingly, SK-N-ASrOXALI4000 cells showed no cross resistance to gemcitabine and increased sensitivity to doxorubicin and UVC radiation, alternative treatments that like platinum drugs target DNA integrity. Notably, UVC-induced DNA damage is thought to be predominantly repaired by nucleotide excision repair and nucleotide excision repair has been described as the main oxaliplatin-induced DNA damage repair system. SK-N-ASrOXALI4000 cells were also more sensitive to lysis by influenza A virus, a candidate for oncolytic therapy, than SK-N-AS cells. In conclusion, we introduce a novel oxaliplatin resistance model. The oxaliplatin resistance mechanisms in SK-N-ASrOXALI4000 cells appear to be complex and not to directly depend on enhanced DNA repair capacity. Models of oxaliplatin resistance are of particular relevance since research on platinum drugs has so far predominantly focused on cisplatin and carboplatin.
Project description:Oxaliplatin (OHP) treatment of colorectal cancer (CRC) frequently leads to resistance. OHP resistance was induced in CRC cell lines LoVo-92 and LoVo-Li and a platinum-sensitive ovarian cancer cell line, A2780, and related to cellular platinum accumulation, platinum-DNA adducts, transporter expression, DNA repair genes, gene expression arrays, and array-CGH profiling. Pulse (4 h, 4OHP) and continuous exposure (72 h, cOHP) resulted in 4.0 to 7.9-fold and 5.0 to 11.8-fold drug resistance, respectively. Cellular oxaliplatin accumulation and DNA-adduct formation were decreased and related to OCT1-3 and ATP7A expression. Gene expression profiling and pathway analysis showed significantly altered p53 signaling, xenobiotic metabolism, role of BRCA1 in DNA damage response, and aryl hydrocarbon receptor signaling pathways, were related to decreased ALDH1L2, Bax, and BBC3 (PUMA) and increased aldo-keto reductases C1 and C3. The array-CGH profiles showed focal aberrations. In conclusion, OHP resistance was correlated with total platinum accumulation and OCT1-3 expression, decreased proapoptotic, and increased anti-apoptosis and homologous repair genes.
Project description:Copper transporter 1 (CTR1), copper transporter 2 (CTR2), copper-transporting p-type adenosine triphosphatase 1 and 2 (ATP7A and ATP7B) are key mediators of cellular cisplatin, carboplatin and oxaliplatin accumulation. In this meta-analysis, we aimed to evaluate the relation of CTR1, CTR2, ATP7A and ATP7B to overall survival (OS), progression-free survival (PFS), disease-free survival (DFS) and treatment response (TR) of cancer patients who received chemotherapy based on published literatures, the Gene Expression Omnibus (GEO) and the Cancer Genome Atlas (TCGA) datasets. Hazard ratios (HRs) and odds ratios (ORs) were pooled using random-effect models. Subgroup analysis and sensitivity analysis were conducted; heterogeneity and publication bias were assessed. Twelve literatures and eight datasets with 2149 patients were included. Our results suggested that high CTR1 expression was associated with favorable OS, PFS, DFS and TR in cancer patients who underwent chemotherapy with acceptable heterogeneity. The relationship of CTR1 to cancer prognosis remained significant in the subgroup of patients who underwent platinum-based chemotherapy, the patients with ovarian cancer and those with lung cancer. The significance of these relationships was not influenced by geological region of publication, data origin or detection method. However, there was no evidence for relation of CTR2, ATP7A or ATP7B to OS, PFS, DFS or TR. Test of publication bias and sensitivity analysis suggested a robustness of all the summary effect estimates. In conclusion, high CTR1 level predicts prolonged survival and enhanced response to chemotherapy in cancer patients who underwent chemotherapy and CTR1 might be a potential target to circumvent chemotherapy resistance.
Project description:<h4>Background</h4>Increasing evidence has suggested the involvement of long non-coding RNA taurine upregulated gene 1 (TUG1) in chemoresistance of cancer treatment. However, its function and molecular mechanisms in esophageal squamous cell carcinoma (ESCC) chemoresistance are still not well elucidated. In the present study, we investigate the functional role of TUG1 in cisplatin (DDP) resistance of ESCC and discover the underlying molecular mechanism.<h4>Results</h4>Our study revealed that TUG1 was up-regulated in DDP-resistant ESCC tissues and cells. High TUG1 expression was correlated with poor prognosis of ESCC patients. TUG1 knockdown improved the sensitivity of ECA109/DDP and EC9706/DDP cells to DDP. Moreover, TUG1 could epigenetically suppress PDCD4 expression via recruiting enhancer of zeste homolog 2. PDCD4 overexpression could mimic the functional role of down-regulated TUG1 in DDP resistance. PDCD4 knockdown counteracted the inductive effect of TUG1 inhibition on DDP sensitivity of ECA109/DDP and EC9706/DDP cells. Furthermore, TUG1 knockdown facilitated DDP sensitivity of DDP-resistant ESCC cells in vivo.<h4>Conclusion</h4>TUG1 knockdown overcame DDP resistance of ESCC by epigenetically silencing PDCD4, providing a novel therapeutic target for ESCC.
Project description:Cisplatin and its platinum analogs, carboplatin and oxaliplatin, are some of the most widely used cancer chemotherapeutics. Although cisplatin and carboplatin are used primarily in germ cell, breast and lung malignancies, oxaliplatin is instead used almost exclusively to treat colorectal and other gastrointestinal cancers. Here we utilize a unique, multi-platform genetic approach to study the mechanism of action of these clinically established platinum anti-cancer agents, as well as more recently developed cisplatin analogs. We show that oxaliplatin, unlike cisplatin and carboplatin, does not kill cells through the DNA-damage response. Rather, oxaliplatin kills cells by inducing ribosome biogenesis stress. This difference in drug mechanism explains the distinct clinical implementation of oxaliplatin relative to cisplatin, and it might enable mechanistically informed selection of distinct platinum drugs for distinct malignancies. These data highlight the functional diversity of core components of front-line cancer therapy and the potential benefits of applying a mechanism-based rationale to the use of our current arsenal of anti-cancer drugs.
Project description:We have recently reported that vatalanib, an orally active small molecule multi-tyrosine kinase inhibitor (Hess-Stumpp et al., 2005 ), can sensitize multidrug resistant (MDR) colon cancer cells to chemotherapy under hypoxia by inhibiting two MDR transporters ABCB1 and ABCG2 (To et al., 2015 ). This data article describes the possible circumvention of resistance to specifically platinum (Pt)-based anticancer drugs by vatalanib via inhibition of two other efflux transporters ABCC2 and ATP7A. Data from the flow cytometric transporter efflux assay showed specific inhibition of ABCC2 activity by vatalanib in stable transfected cells and ABCC2-overexpressing oxaliplatin-resistant colon cancer cells HCT116/Oxa. We also performed the transporter ABCC2 ATPase assay and showed an increase in ATP hydrolysis by ABCC2 in the presence of vatalanib. ATP7A mRNA expression was also shown to be upregulated in HCT116/Oxa cells. Vatalanib was shown to suppress this upregulated ATP7A expression. Data from the cellular Pt accumulation assay showed a lower Pt accumulation in HCT116/Oxa cells than the parental sensitive HCT116 cells. Vatalanib was shown to increase cellular Pt accumulation in a concentration-dependent manner. Combination of oxaliplatin and vatalanib was shown to restore the suppressed apoptosis in HCT116/Oxa cells.