Background:Radio- and chemotherapy (RT/CT) resistance hampers success in combating small and non-small cell lung cancers (SCLC/NSCLC). The underlying molecular mechanisms of RT/CT resistance of LCs are multifactorial and have been understood in part hitherto. miRNAs, key regulators of mRNAs, are well-recognised oncomirs; however, their role in regulating RT response remains poorly understood.Methods:Six human NSCLC and five SCLC cell lines with different SF2 values were investigated. Using microarray we examined whether expression of miRNAs is linked to the RT resistance of NSCLCs or SCLCs. Obtained data were validated by quantitative real-time PCR. Apoptosis and senescence were analysed using siRNA transfection, western blot and flow cytometry.Results:miRNA-21, miRNA-1827, miRNA-214, miRNA-339-5p, miRNA-625, miRNA-768-3p, miRNA-523-3p, miRNA-1227, miRNA-324-5p, miRNA-423-3p, miRNA-1301 and miRNA-1249 are differentially expressed in LC cells. miRNA-214 is upregulated in RT-resistant NSCLC cells relative to radiosensitive counterparts. Considering miRNA-214 as a putative regulator of RT resistance, we demonstrate that knockdown of miRNA-214 in radioresistant NSCLCs sensitised them to RT by stimulation of senescence. Consistently, overexpression of miRNA-214 in radiosensitive NSCLCs protected against RT-induced apoptosis. Protection was mediated by p38MAPK, as downregulation of this kinase could reverse the miRNA-214 overexpression-induced resistance of NSCLC cells.Conclusion:miRNA profiling of LC revealed putative RT resistance signalling circuits, which might help in sensitisation of LC to RT.

The prognosis of non-small cell lung cancer (NSCLC) is poor, since it has often metastasized to distant organs by the time of diagnosis. Therefore, biomarkers predicting metastasis are crucial. miRNAs play important roles in the regulation of different tumor cell processes, including metastasis. We recently showed that miRNA-214 is linked to a radioresistant phenotype of NSCLC. miRNA-214 has been linked to metastasis in other tumor types. Therefore, we examined the role of miRNA-214 in the metastatic potential of NSCLC. We showed that downregulation of miRNA-214 increased invasive potential, and conversely, overexpression of miRNA-214 decreased invasiveness of NSCLC cells in vitro. Gene expression and bioinformatic analyses of NSCLC cells with ablated miRNA-214, identified a number of metastasis-related target genes, including pregnancy-associated plasma protein A (PAPP-A), alpha protein kinase 2 (ALPK2), cyclin-dependent kinase 6 (CDK6) and tumor necrosis-factor alpha-induced protein 3 (TNFAIP3). These were validated on mRNA and protein level to be regulated by miRNA-214. Through immunoprecipitation we showed that only ALPK2 is directly regulated by miRNA-214. We also examined the protein expression of these four genes in NSCLC tumors with respect to metastatic potential. These results showed that NSCLC tumors express these proteins at moderate-high levels in the nucleus, cytoplasm and/or plasma membrane although with no significant correlation to the overall survival or the metastatic potential of the patients. However, we also showed that the membrane-localized PAPP-A had a higher expression level compared to the cytoplasm-localized. In conclusion, we show that low miRNA-214 expression is linked to a higher invasive potential of NSCLC cells.

BackgroundPlatinum compounds are the mainstay of chemotherapy for lung cancer. Unfortunately treatment failure remains a critical issue since about 60 % of all non-small cell lung cancer (NSCLC) patients display intrinsic platinum resistance.MethodsWe analyzed global gene expression profiles of NSCLC clones surviving a pulse treatment with cisplatin and mapped deregulated signaling networks in silico by Ingenuity Pathway Analysis (IPA). Further validation was done using siRNA.ResultsThe pooled cisplatin-surviving NSCLC clones from each of the biological replicates demonstrated heterogeneous gene expression patterns both in terms of the number and the identity of the altered genes. Genes involved in Wnt signaling pathway (Dickkopf-1, DKK1), DNA repair machinery (XRCC2) and cell-cell/cell-matrix interaction (FMN1, LGALS9) were among the top deregulated genes by microarray in these replicates and were validated by q-RT-PCR. We focused on DKK1 which previously was reported to be overexpressed in NSCLC patients. IPA network analysis revealed coordinate up-regulation of several DKK1 transcriptional regulators (TCF4, EZH2, DNAJB6 and HDAC2) in cisplatin-surviving clones from that biological replicate. Knockdown of DKK1 by siRNA sensitized for cisplatin in two different NSCLC cell lines and in ovarian A2780 cells, but not in the A2780 cis subline made resistant to cisplatin by chronic exposure, suggesting a role of DKK1 in intrinsic but not acquired platinum refractoriness.ConclusionsWe identified DKK1 as a possible marker of a cisplatin-refractory phenotype and as a potential novel therapeutic target to improve platinum response of NSCLC cells.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1635-9) contains supplementary material, which is available to authorized users.

Cetuximab combined with irinotecan when administered biweekly is safe and effective for treatment of pretreated elderly patients with mCRC.

Non small cell lung cancer (NSCLC) constitutes the majority of lung cancer diagnoses and platinum-based chemotherapy regimen is the standard treatment. Unfortunately, a large proportion do not respond or relapse on such treatment regimens for yet only partly revealed mechanisms. We profiled residual clones of NSCLC cells after treatment with cisplatin. The mRNA and miRNA pattern was compared between untreated and cisplatin-resistant NSCLC clones using Affymetrix gene and miRNA expression arrays, respectively. We found that around 1100 genes were up regulated (at least two fold) and about 1200 genes were down regulated (at least two fold) in the cisplatin-resistant clones compared with untreated NSCLCs. We then used the Gene Set Enrichment Analysis (GSEA) to find the over- and underrepresented gene sets in large datasets like, Gene Trail and Gene Ontology. We found that cisplatin-refractory NSCLCs showed increased expression of genes involved in different cellular mechanisms including DNA metabolic processes, DNA replication and DNA-dependent DNA replication initiation for example, XRCC2, FANCI, FANCD2, RRM1, MRE11A, BLM, RRM2 and TOP2A which were validated on RNA, protein and functional level for their role in cisplatin sensitivity of NSCLC. We also found a few miRNAs that were up regulated in cisplatin refractory NSCLC by a least 1.3-fold e.g. miRNA-25 star and miRNA-1290. Interestingly, only 14 miRNAs were down regulated (by at least 1.3 fold) in the surviving cisplatin refractory NSCLC cells. Among these miRNAs were (miR-29b, miR-210, miR-486-3p, miR-324-5p, miR-572, miR-30a, e, miR-424, miR-449a, b, miR-301a, miR-153, miR-150 star and miR-34c-5p). We then looked for the predicted targets of these miRNAs using Target scan and microcosm and we found a large list of genes to be potential targets of the miRNAs among them are genes involved in apoptosis, cell cycle regulation, Wnt signaling pathway and other cellular processes (ex. VDAC, SMAD2, RAN, etc.). Correlation analysis between miRNA gene targets and gene expression data is ongoing with the aim to identify and validate putative targets on RNA, protein and functional level. In conclusion, our data show that cisplatin refractory NSCLC tumors have dysregulated mRNAs and miRNAs. Among the dysregulated genes, are certain genes involved in DNA metabolism, DNA replication and DNA-dependent DNA replication initiation which may be novel targets for improved platinum-based therapy response of NSCLC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 163. doi:10.1158/1538-7445.AM2011-163