Development Of NSCLC Research Articles

Association of EGFR and KRAS mutations with expression of p-AKT, DR5 and DcR1 in non-small cell lung cancer.

The activation of AKT is one of the causes of resistance to epidermal growth factor receptor (EGFR)- tyrosine kinase inhibitors (TKIs). Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) combines with related receptors to trigger apoptosis or protect the cells against TRAIL apoptosis. This research focused on the association of EGFR and KRAS mutations with expression of AKT, p-AKT, DR5 and DcR1 in non-small cell lung cancer. 82 NSCLC patients were included in the study. xTAG liquichip techonolgy (xTAG-LCT) was applied to investigate the genetic mutation of EGFR and KRAS, Quantitative Real-time PCR was used to test the mRNA expression of AKT, DR5 and DcR1 and Western Blot was applied to test the protein expression of AKT, p-AKT, DR5, and DcR1. We found that of 82 patients, 31 cases had EGFR-activating mutations, more common in female, adenocarcinoma, and non-smoker patients; 9 cases had KRAS mutations, frequently found in patients with smoking history. The expression of AKT and p-AKT correlated with staging, tumor differentiation, and lymph node metastasis. The expression of DR5 in phase III and low differentiation tumor was significantly higher than that in phase I+II and high and median differentiation tumor; the expression of DcR1 in phase III and low differentiation tumor was significantly lower than that in phase I+II and high and median differentiation tumor. Compared with EGFR and KRAS wild type, in NSCLC tissue with EGFR and KRAS mutations, the expression of AKT and p-AKT was significantly higher. These results suggest that EGFR and KRAS mutation status was associated with the expression of AKT and p-AKT. AKT, p-AKT, DR5, and DcR1 all took part in the occurrence and development of NSCLC, and may become a reference index to evaluate the prognosis of NSCLC.

Association between the TP53 and CYP2E1*5B gene polymorphisms and non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is the most common form of lung cancer. Genetic polymorphisms in tumour suppressor genes and genes encoding xenobiotic metabolising enzymes alter the activity of their corresponding enzymes and are important individual susceptibility factors for NSCLC. Because of the lack of information in literature, the aim of our study was to investigate the role of the tumour suppressor gene TP53 (Arg72Pro) and the xenobiotic metabolising CYP2E1*5B gene polymorphisms on the risk of NSCLC development. The study population consisted of 172 patients and 172 controls (156 men and 16 women in each group). Genetic polymorphisms were determined with real-time polymerase chain reaction (PCR) and PCR restriction fragment length polymorphism (PCR-RFLP). Multivariate analysis showed a significant association with NSCLC for the combination between the TP53 codon72 Arg/Pro and the Pro/Pro genotypes (OR 2.21, 95 % CI 1.390-3.51; p=0.001). We also analysed whether combinations of these gene variants with GSTM1, GSTT1, GSTP1 exon 5 (Ile105Val), and GSTP1 exon 6 (Ala114Val) gene polymorphisms were associated with the NSCLC risk. A significant increase in the risk was observed for the following combinations: TP53 codon72 variant with GSTM1 null (OR 2.22, 95 % CI 1.23-4.04; p=0.009), GSTT1 null (OR 2.98, 95 % CI 1.49-5.94; p=0.002), and GSTP1 (Ala114Val) variant genotypes (OR 3.38, 95 % CI 1.54-7.41; p=0.002). Further studies with larger samples are needed to verify these findings.

Clinicopathological significance of WIF1 hypermethylation in NSCLC, a meta-analysis and literature review.

Methylation of the WIF-1 gene can lead to the loss of WIF-1 expression which has been observed in numerous types of cancer including NSCLC. However, the association and clinicopathological significance between WIF-1 promoter hypermethylation and NSCLC remains unclear. In the present study, we performed a meta-analysis to evaluate the clinicopathological significance of WIF-1 hypermethylation in NSCLC. A systematic literature search was carried out using Pubmed, EMBASE, Web of Science and CNKI. The Cochrane software Review manager 5.2 was used. The frequency of WIF-1 hypermethylation was significantly increased in NSCLC compared with normal lung tissue; the pooled OR was 8.67 with 95% CI 1.64-45.88, p = 0.01. The rate of WIF-1 hypermethylation was higher in SCC than in AC, OR was 1.74 with 95% CI 0.97-3.11, p = 0.06. In addition, WIF-1 loss was correlated with low 5-year survival rate. In summary, WIF-1 hypermethylation is a potential biomarker for diagnosis of NSCLC. WIF-1 hypermethylation is predominant in squamous cell carcinoma (SCC), suggesting that WIF-1 methylation contributes to the development of NSCLC, especially SCC.

Case-Control Study: Smoking History Affects the Production of Tumor Antigen–Specific Antibodies NY-ESO-1 in Patients with Lung Cancer in Comparison with Cancer Disease–Free Group

IntroductionLung cancer is the leading cause of cancer mortality worldwide; therefore, understanding the biological or clinical role of tumor-associated antigens and autoantibodies is of eminent interest for designing antitumor immunotherapeutic strategies. MethodsHere we prospectively analyzed the serum frequencies of New York esophageal squamous cell carcinoma 1 (NY-ESO-1), human epidermal growth factor 2/neu, and melanoma-associated antigen A4 (MAGE-A4) antibodies and expression of the corresponding antigens in tumors of 121 patients with NSCLC undergoing an operation without prior neoadjuvant chemotherapy and compared them with those in 57 control age-matched patients with no history of a malignant disease. ResultsWe found that only antibodies specific for NY-ESO-1 (19.8% [n = 24 of 121]) were significantly increased in the group of patients with NSCLC compared with in the controls. NY-ESO-1 seropositivity was significantly positively associated with an active smoking history in patients with NSCLC but not in smokers from the control group. In tumors, the frequency of NY-ESO-1 mRNA expression was 6.3% (in four of 64 patients), the frequency of human epidermal growth factor 2/neu (HER 2/neu) expression was 11.9% (five of 42), and the frequency of MAGE-A4 expression was 35.1% (20 of 57). MAGE-A4 expression in tumors correlated with smoking status and male sex in patients with NSCLC. Patients with squamous cell carcinoma displayed higher expression of NY-ESO-1 and MAGE-A4 in tumors than did patients with adenocarcinoma. On the other hand, 94.7% of nonsmoking patients in our study had adenocarcinoma (of whom 73.7% were women). ConclusionThese results confirm the reported high immunogenicity of NY-ESO-1 and suggest that a smoking-induced chronic inflammatory state might potentiate the development of NY-ESO-1–specific immune responses. Moreover, smoking might contribute to the expression of other cancer/testis antigens such as MAGE-A4 at early stages of NSCLC development.

MicroRNA-410 acts as oncogene in NSCLC through downregulating SLC34A2 via activating Wnt/β-catenin pathway.

SLC34A2 had been reported to be down-regulated in human NSCLC cells and patient tissues, and played a significant role in lung cancer. However, the mechanism of its unusual expressionin NSCLC has not been fully elucidated. In present study, we identified SLC34A2 was a direct target of miR-410 and could be inhibited by miR-410 transcriptionally and post-transcriptionally. MiR-410 promoted the growth, invasion and migration of NSCLC cells in vitro. An orthotopic xenograft nude mouse model further affirmed that miR-410 promoted NSCLC cell growth and metastasis in vivo. Moreover, restoring SLC34A2 expression effectively reversed the miR-410-mediated promotion of cell growth, invasion and migration in NSCLC cells. In addition, miR-410high /SLC34A2low expression signature frequently existed in NSCLC cells and tumor tissues. MiR-410 significantly increased the expression of DVL2 and β-catenin protein while decreased that of Gsk3β protein of Wnt/β-catenin signaling pathway, while SLC34A2 partly blocked the effects of miR-410 on those protein expressions. Hence, our data for the first time delineated that unusual expression of SLC34A2 was modulated by miR-410, and miR-410 might positivelycontribute to the tumorigenesis and development of NSCLC by down-regulating SLC34A2 and activating Wnt/β-catenin signaling pathway. MiR-410 might be a new potential therapeutic target for NSCLC.

Long noncoding RNAs: new insights into non-small cell lung cancer biology, diagnosis and therapy.

Recent advances in tiling array and high throughput analyses revealed that at least 87.3% of the human genome is actively transcribed, though <3% of the human genome encodes proteins. This unexpected truth suggests that most of the transcriptome is constituted by noncoding RNA. Among them, high-resolution microarray and massively parallel sequencing analyses identified long noncoding RNAs (lncRNAs) as nonprotein-coding transcripts. lncRNAs are largely polyadenylated and >200 nucleotides in length transcripts, involved in gene expression through epigenetic and transcriptional regulation, splicing, imprinting and subcellular transport. Although lncRNAs functions are largely uncharacterized, accumulating data indicate that they are involved in fundamental biological functions. Conversely, their dysregulation has increasingly been recognized to contribute to the development and progression of several human malignancies, especially lung cancer, which represents the leading cause of cancer-related deaths worldwide. We conducted a comprehensive review of the published literature focusing on lncRNAs function and disruption in nonsmall cell lung cancer biology, also highlighting their value as biomarkers and potential therapeutic targets. lncRNAs are involved in NSCLC pathogenesis, modulating fundamental cellular processes such as proliferation, cell growth, apoptosis, migration, stem cell maintenance and epithelial to mesenchymal transition, also serving as signaling transducers, molecular decoys and scaffolds. Also, lncRNAs represent very promising biomarkers in early-stage NSCLC patients and may become particularly useful in noninvasive screening protocols. lncRNAs may be used as predictive biomarkers for chemotherapy and targeted therapies sensitivity. Furthermore, selectively targeting oncogenic lncRNAs could provide a new therapeutic tool in treating NSCLC patients. lncRNAs disruption plays a pivotal role in NSCLC development and progression. These molecules also serve as diagnostic, prognostic and predictive biomarkers. Characterization of lncRNA genes and their mechanisms of action will enable us to develop a more comprehensive clinical approach, with the final goal to benefit our patients.

A Phase I Study of IGN523, a Novel Anti-CD98 Monoclonal Antibody in Patients with Relapsed or Refractory Acute Myeloid Leukemia (AML)

CD98 is a heterodimeric cell surface protein that is implicated in the pathogenesis of many hematological and solid malignancies. Elevated tumor CD98 expression has been correlated with poor prognosis. Previous studies demonstrated that CD98 functions in integrin signaling and amino acid transport processes that support tumor cell proliferation, anchorage independence, invasion, and metastasis. IGN523 is a humanized monoclonal antibody targeting CD98 with multiple mechanisms of action including antibody-dependent cellular cytotoxicity (ADCC), direct cell death induction, and interference with the normal amino acid transport function of CD98. IGN523 elicits potent anti-tumor activity against a variety of leukemic xenograft models (Hayes et al., Int. J. Cancer 2015). Here we present the results of a Phase I dose escalation study of IGN523 in adult patients with relapsed or refractory AML.IGN523 was administered intravenously (IV) as a single agent to 19 patients at weekly doses ranging from 0.1 mg/kg to 20 mg/kg, including 3 single-patient cohorts starting at 0.1, 0.3, and 1 mg/kg, 6 patients treated at 10 mg/kg, 3 at 20 mg/kg, and 7 at 15 mg/kg. Median age was 71 years (range 23-78). Patients received a median of one 4-week cycle of treatment (range <1-10 cycles).The most common treatment-related adverse events observed were part of infusion-related reactions (IRRs) occurring primarily with the first infusion, and were generally Grade 1 or 2 (CTCAE V4.03), including chills (52.6%), fever (pyrexia or febrile neutropenia, 36.9%), nausea (26.3%), and vomiting (26.3%). Premedication with acetaminophen, diphenhydramine, and hydrocortisone was instituted after a dose-limiting toxicity (DLT) of significant IRR with the first dose was observed in 1 of 3 patients treated at 10 mg/kg, and the cohort was expanded to 6 patients. Treatment-related Grade 3 and 4 AEs occurring in >1 patient included febrile neutropenia (26.1%), pyrexia (10.5%), and altered mental status (confusional state or mental status changes in 15.8% and 10.5% respectively). DLTs all occurred with the first dose of IGN523 and were observed in 1 of 6 patients at 10 mg/kg, 2 of 3 at 20 mg/kg, and 2 of 7 at 15 mg/kg. The maximum tolerated dose (MTD) of IGN523 administered weekly was 10 mg/kg.PK studies of IGN523 in all patients demonstrated evidence of target-mediated drug disposition at doses below 10 mg/kg/week, as pre-dose (trough) CD98 receptor occupancy (%RO) levels tended to be < 90%. By the 10 mg/kg dose cohort, pre-dose %RO levels were 50 to > 90% during the first few weeks of dosing, and consistently > 90% after 4-8 weeks of dosing. At 15 and 20 mg/kg/week doses, %RO was consistently > 90% from the first dose. Therefore, dose levels of 10 mg/kg/week and above should be suitable for most patients to maintain >90% receptor occupancy over time. Anti-drug antibodies (ADA) were not detected in any patient.Evidence of transient anti-leukemic activity was observed in 3 patients, including improvement in platelet count and reduction in marrow blast percentage, early reduction and control of blasts, and an episode of tumor lysis and peripheral blast count reduction. No complete or partial responses were observed.In conclusion, IGN523 was safe when administered at 10 mg/kg IV weekly with DLTs of significant first-dose infusional toxicity that may be manageable with prophylactic steroids or a modified dosing regimen. IGN523 demonstrated modest evidence of anti-leukemic activity as a single agent. Preclinical data demonstrating greater than additive activity of IGN523 when combined with cytarabine and significant monotherapy and combination activity with standard chemotherapy in patient-derived xenograft models of NSCLC and colorectal cancer support further development of IGN523 in combination with standard therapies in AML and solid tumors. DisclosuresBixby:Seattle Genetics, Inc.: Research Funding. Akard:Novartis: Research Funding, Speakers Bureau; BMS: Research Funding, Speakers Bureau; Celgene: Speakers Bureau; Millenium: Speakers Bureau; Ariad: Research Funding, Speakers Bureau; Gilead: Speakers Bureau; Pfizer: Research Funding; Incyte: Research Funding. Khoury:Pfizer: Honoraria; Ariad: Honoraria; Bristol-Meyer-Squibb: Honoraria. Becker:Igenica: Research Funding. Van Der Horst:Igenica: Employment. Ho:Igenica: Employment. Cortes:Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Research Funding; BerGenBio AS: Research Funding; BMS: Consultancy, Research Funding; Ariad: Consultancy, Research Funding; Astellas: Consultancy, Research Funding; Ambit: Consultancy, Research Funding; Arog: Research Funding; Celator: Research Funding; Jenssen: Consultancy.

Abstract C93: RYBP predicts survival of patients with non-small cell lung cancer and sensitizes chemotherapy

Abstract Non-small cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancers and associates with a poor prognosis. Despite several targeted therapies have been developed with promising results, the therapeutic options for NSCLC are limited and the 5-year survival rate remains poor (∼15%). Therefore, there is an urgent need for a better understanding of its pathogenesis and exploring novel therapeutic targets. Ring1 and YY1 binding protein (RYBP) is a member of polycomb group (PcG) proteins that typically act as transcriptional repressors via epigenetic modification of chromatin and participate in the establishment and maintenance of cell fates. Recent studies have shown that RYBP also possesses PcG-independent functions that promote apoptosis and cell cycle arrest in cancer cells. Our previous study has shown that RYBP stabilizes p53 through inhibiting MDM2 activity. Recent study indicates that RYBP is downregulated in human cancers, including NSCLC, but the underlying mechanisms are largely unknown. The present study was designed to demonstrate the molecular role of RYBP in NSCLC development and progression, chemotherapy, and patient survival. We systemically investigated the expression of RYBP in human NSCLC tissues and matched non-cancerous samples and evaluated the associations between the RYBP expression in NSCLCs and patient survial. We also generated a replication-deficient recombinant adenovirus driving the expression of RYBP (Adv-RYBP) and carried out in vitro and in vivo studies to explore the molecular basis for the tumor suppressing role of RYBP in NSCLC. We first demonstrated that the RYBP mRNA and protein expressions levels were significantly down-regulated in NSCLC tissues compared with matched noncancerous lung tissues. The low expression of RYBP was significantly associated with the poor prognosis in NSCLC patients. We then identified that enforced RYBP expression decreased cell viability, inhibited colony formation and induced apoptosis in NSCLC cells, while RYBP knockdown led to the opposite effects. RYBP affected the expression of several apoptosis-related proteins; and promoting apoptosis was one of the main mechanisms of RYBP-mediated cell growth inhibition in NSCLC. Moreover, AdRYBP led to a decreased NSCLC tumor xenograft growth. Additionally, clinically used chemotherapeutic agents induced the expression of RYBP, and RYBP sensitized NSCLC cells to chemotherapy in vitro and in vivo. In conclusion, our results reveal that RYBP is a potential biomarker for the diagnosis and prognosis of NSCLC and that reactivating RYBP in cancer cells may provide an effective and safe therapeutic approach to NSCLC therapy. (Supported by NIH/NCI grants R01 CA186662). Citation Format: wei wang, Sukesh Voruganti, Jiang-Jiang Qin, Biyun Qian, Ruiwen Zhang. RYBP predicts survival of patients with non-small cell lung cancer and sensitizes chemotherapy. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C93.

Over-expressed RPL34 promotes malignant proliferation of non-small cell lung cancer cells

Ribosomal protein L34 (RPL34) was reported to be involved in the regulation of cell proliferation of prokaryotes, plant and animal cells. In the present study, we analyze the expression and function of RPL34 in NSCLC. Immunohistochemical analysis, qPCR and Western blot were used to detect the expression of RPL34 in NSCLC tissues and cells lines. Flow cytometry was used to detect cell activity of NSCLC cell line H1299 under lentivirus-mediated RNAi on RPL34. Cell proliferation and colony formation assays were used to analyze the role of RPL34 in NSCLC cell proliferation. We found that expression of ribosomal protein RPL34 was significantly up-regulated in NSCLC tissues compared to adjacent normal tissues. Lentivirus-mediated shRNA knockdown of RPL34 in NSCLC cell line H1299 resulted in a strong decrease of proliferation, and a moderate but significant increase of apoptosis and S-phase arrest. These data indicate that over-expressed RPL34 may promote malignant proliferation of NSCLC cells, thus playing an important role in development and progress of NSCLC.

Expression profile analysis of microRNAs and downregulated miR-486-5p and miR-30a-5p in non-small cell lung cancer.

Lung cancer is the leading cause of cancer-related mortality worldwide and although there have been improvements in treatment there is a low survival rate. The aim of the present study was to investigate the effect of microRNA (miRNA) on cell pathways. A miRNA microarray was used to profile miRNAs of lung cancer tissues. It was identified that 33 miRNAs with >2.0-fold change and FDR <0.05 were differentially expressed between the adjacent non-cancerous lung tissues and non-small cell lung cancers NSCLCs (P<0.005). The data were optimized in combination with physical interaction analysis to obtain crucial miRNAs. The results showed that differentially expressed miRNAs were associated with biological processes such as cell migration, protein phosphorylation and neuron differentiation, and signaling pathways such as MAPK, TGF-β and PI3K/Akt signaling pathways. Validation of significant miRNAs in independent 40 paired NSCLC tissues demonstrated that the expression level of miR-486-5p and miR-30a-5p was significantly downregulated in another 40 paired lung cancer tissues. Taken together, the results provided strong evidence of the possible involvement of miRNAs in the development and progression of NSCLC. Thus, the results are of importance for clinical investigators and for those who design miRNA‑based novel cancer therapeutics.

The effects and mechanisms of SLC34A2 in tumorigenesis and progression of human non-small cell lung cancer.

BackgroundSLC34A2 with highest expressions in lung, small intestine and kidney encoded a type 2b sodium-dependent phosphate transporter (NaPi-IIb). In lung, SLC34A2 only expressed in the apical membrane of type II alveolar epithelium cells (ATII cells) and played a pivotal role during the fetal lung development and embryonic development. ATII cells acting as multifunctional stem cells might transform into NSCLC after undergoing exogenous or endogenous factors. Increasing evidences showed that the genes performing critical roles during embryogenesis were also expressed during the development of cancer. In addition, recent research found the expression of SLC34A2 had a significant difference between the surgical samples of NSCLC and normal tissues, and SLC34A2 was down-regulated in lung adenocarcinoma cell line A549 and up-regulation expression of SLC34A2 could significantly inhibit cell viability and invasion of A549 in vitro. These results suggested SLC34A2 might play an important role in the development of NSCLC. However, the role of SLC34A2 in tumorigenesis and progression of NSCLC remains unknown.ResultsOur study found that SLC34A2 was also significantly down-regulated in 14/15 of examined NSCLC tissues. Moreover, we found that expressions of SLC34A2 were reduced in six NSCLC cell lines for the first time. Our result also revealed a dramatic inhibitory effects of SLC34A2 on cell growth, migration and invasion of several NSCLC cell lines. SLC34A2 also strongly inhibited tumor growth and metastasis ability in A549 subcutaneous tumor model and lung metastasis model, respectively. Further studies found that the suppressive effects of SLC34A2 on tumorigenesis and progression might be associated with the down-regulation of related protein in PI3K/Akt and Ras/Raf/MEK signal pathway.ConclusionsFor the first time, our data indicated that SLC34A2 could exert significantly suppressive effects on tumorigenesis and progression of NSCLC. SLC34A2 might provide new insights for further understanding the early pathogenesis of human NSCLC.

Association between polymorphisms in the XRCC1 gene and the risk of non-small cell lung cancer.

Here, we have reported a case-control study investigating the association between XRCC1 codons Arg194Trp, Arg280His, and Arg399Gln and the development of NSCLC. NSCLC patients (N = 245) and healthy controls (N = 257) were randomly selected from the Huaihe Hospital between March 2012 and August 2014. DNA extracted from the patient and control blood samples were subjected to polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to assess the genotyping of XRCC1 Arg194Trp, Arg280His, and Arg399Gln. Multivariate logistic regression analyses revealed an association between the expression of the AA genotype and A allele genotypes and a significantly increased risk of NSCLC, compared to the GG genotype [95% confidence interval (CI); Odd's ratio (OR) = 2.82 (1.141-5.86) and 1.67 (1.17-2.37), respectively]. The potential association between the A allele of XRCC1 Arg399Gln and the risk of NSCLC was more evident in smokers (95%CI; OR = 1.70; 1.11- 2.63). In conclusion, the XRCC1 Arg399Gln polymorphism was found to be associated with increased risk of NSCLC, especially in tobacco smokers.

Zinc finger protein ZBTB20 promotes cell proliferation in non-small cell lung cancer through repression of FoxO1

In the present study, we found that ZBTB20, a member of the POK (POZ and Krüppel) family of transcriptional repressors, was significantly up-regulated in lung cancer tissues, compared with adjacent normal tissues. Our in vitro studies further found that ZBTB20 overexpression promoted, while its inhibition using small interfering RNA suppressed cell proliferation. Consistently, key regulators in cell-cycle progression, such as Cyclin D1, Cyclin E, P21 and P27, were also regulated by ZBTB20. At the molecular level, we further revealed that FoxO1, a tumor suppressor in multiple human cancers, was transcriptionally repressed by ZBTB20. Therefore, our results highlight an important role for ZBTB20 in controlling NSCLC development, which might be helpful to identify potential therapeutic targets for its treatment.

CRTC2 and PROM1 expression in non-small cell lung cancer: analysis by Western blot and immunohistochemistry.

Accumulating evidence supports that genetic factors are another risk factors for lung cancer. Previously, we used whole exome sequencing with sanger sequencing to search for genetic-related mutations in one of four individuals from a pedigree with lung cancer history. Then, we used PCR-RFLP and direct-sequence in the sample size of 318 individuals with lung cancer (cases) and 272 controls. Recently, we detected two new genes including CRTC2 (CREB regulated transcription coactivator 2) and PROM1(human prominin-1,CD133). We investigated the CRTC2 mutation and PROM1 mutation of surgically resected NSCLC tissues (n=200). The presence or absence of CRTC2 and PROM1 mutation was analyzed by direct sequencing. The expression of CRTC2 and PROM1 was studied by western blot and immunohistochemical analysis of the lung cancer tissues which had the mutation of the two genes(cases), the samples without mutations(controls) and the normal lung tissue(controls). CRTC2 and PROM1 mutations in 5 NSCLC tissues and 3 NSCLC tissues out of the samples were identified. The positive results were closely correlated with clinicopathological features, such as male gender, adenocarcinoma, smoker status, and older age (≥55). We found that the CRTC2 and PROM1 expression were significantly higher in tissues of NSCLS with mutations than that without mutations and the normal lung tissue. The results imply that the high expression of CRTC2 and PROM1 may play an important role in the development and hereditary of NSCLC.

Alterations of LKB1 and KRAS and risk of brain metastasis: Comprehensive characterization by mutation analysis, copy number, and gene expression in non-small-cell lung carcinoma

BackgroundBrain metastases are one of the most malignant complications of lung cancer and constitute a significant cause of cancer related morbidity and mortality worldwide. Recent years of investigation suggested a role of LKB1 in NSCLC development and progression, in synergy with KRAS alteration. In this study, we systematically analyzed how LKB1 and KRAS alteration, measured by mutation, gene expression (GE) and copy number (CN), are associated with brain metastasis in NSCLC. Materials and methodsPatients treated at University of North Carolina Hospital from 1990 to 2009 with NSCLC provided frozen, surgically extracted tumors for analysis. GE was measured using Agilent 44,000 custom-designed arrays, CN was assessed by Affymetrix GeneChip Human Mapping 250K Sty Array or the Genome-Wide Human SNP Array 6.0 and gene mutation was detected using ABI sequencing. Integrated analysis was conducted to assess the relationship between these genetic markers and brain metastasis. A model was proposed for brain metastasis prediction using these genetic measurements. Results17 of the 174 patients developed brain metastasis. LKB1 wild type tumors had significantly higher LKB1 CN (p<0.001) and GE (p=0.002) than the LKB1 mutant group. KRAS wild type tumors had significantly lower KRAS GE (p<0.001) and lower CN, although the latter failed to be significant (p=0.295). Lower LKB1 CN (p=0.039) and KRAS mutation (p=0.007) were significantly associated with more brain metastasis. The predictive model based on nodal (N) stage, patient age, LKB1 CN and KRAS mutation had a good prediction accuracy, with area under the ROC curve of 0.832 (p<0.001). ConclusionLKB1 CN in combination with KRAS mutation predicted brain metastasis in NSCLC.

RNA interference-mediated knockdown of Aurora-B alters the metastatic behavior of A549 cells via modulation of the phosphoinositide 3-kinase/Akt signaling pathway.

Accumulating evidence has revealed that an elevated expression level of Aurora-B is associated with metastasis in various types of malignant tumor. However, it is currently unclear whether this molecule is involved in non-small lung cancer (NSCLC) metastasis, and the molecular mechanisms associated with Aurora-B and metastasis remain unknown. In the present study, in order to investigate whether Aurora-B is involved in the development and metastasis of NSCLC, the Aurora-B protein expression in NSCLC tissues was detected by immunohistochemistry and its association with metastasis was analyzed. The results revealed that the expression levels of the Aurora-B protein in tissues obtained from NSCLC patients with lymph node metastasis were significantly higher than those without metastatic disease. Furthermore, the effect of Aurora-B inhibition on A549 cell migration and invasion, as well as the activity of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway was evaluated. Aurora-B was inhibited in the A549 cells using short hairpin RNA, and the cell migration and invasion rates were investigated using wound healing and Transwell invasion assays. In addition, the expression of the main proteins in the PI3K/Akt/nuclear factor-κB (NF-κB) signaling pathway, and matrix metalloproteinase (MMP)-2 and -9 were measured by western blot analysis. The results demonstrated that cell migration and invasion were decreased as a result of silencing Aurora-B. Furthermore, the activity of the PI3K/Akt/NF-κB signaling pathway and the expression of MMP-2 and -9 protein were suppressed by silencing Aurora-B. The results of the present study indicate that the knockdown of Aurora-B suppresses A549 cell invasion and migration via the inhibition of the PI3K/Akt signaling pathway in vitro and thus, targeting Aurora-B may present a potential treatment strategy for NSCLC.

Metadherin regulates proliferation and metastasis via actin cytoskeletal remodelling in non-small cell lung cancer.

Background:Metaderin (MTDH) protein is a novel component part of tight junction complex. The aim of this study was to investigate the correlation between MTDH and prognosis of patients and to explore the role of MTDH on NSCLC development and metastasis.Methods:Relative mRNA expression was evaluated by quantitative real-time PCR, and protein expression was detected using immunohistochemistry staining. The role of MTDH in cancer cell proliferation, migration and invasion was studied by modulation of MTDH expression in NSCLC cell lines. These functions of MTDH were further confirmed in vivo.Results:In NSCLC, low MTDH protein expression was correlated with lymph node metastasis, TNM stage and decreased OS (P=0.001, 0.011 and 0.013, respectively). Overexpression of MTDH reduced anchorage-independent and -dependent growth through arresting cell cycle, inhibited migration and invasion in vitro and further suppressed tumorigenesis, tumour growth and metastasis in vivo. Knockdown of MTDH expression increased cell invasiveness. MTDH overexpression reversed pro-metastatic actin cytoskeleton remodelling and inhibited EMT, supporting that MTDH has a key role on cancer proliferation and metastasis.Conclusions:MTDH has an important role in NSCLC proliferation and metastasis and provides potential in predicting metastasis and prognosis for patients with NSCLC.

Downregulation of miR-638 promotes invasion and proliferation by regulating SOX2 and induces EMT in NSCLC.

Aberrant expression of microRNAs has been shown to regulate the biological processes of lung cancer cells. However, the role of miR-638 in the development of NSCLC is still unclear. In this study, low miR-638 and high SOX2 were shown to be associated with tumor size and metastasis of NSCLC patients. Downregulated miR-638 could promote cell invasion and proliferation, while high miR-638 expression reversed the effect. Furthermore, miR-638 could regulate SOX2 by directly binding to its 3'-UTR. Silencing of SOX2 by siRNA partially abolished the enhancement of cell invasion and proliferation induced by downregulated miR-638. Aberrant miR-638 expression could modulate the expression levels of markers of epithelial-to-mesenchymal transition. Our results indicate that miR-638 may play a pivotal role in the development of NSCLC.

Gene Silencing Associated with SWI/SNF Complex Loss during NSCLC Development

Gene Silencing Associated with SWI/SNF Complex Loss during NSCLC Development

Study of the expression and function of Syk and VEGF-C in the development of non-small cell lung cancer

To explore the role of Syk and VEGF-C in the development of NSCLC. Transfered these genes(eukaryotic expression vector pcDNA3.1-VEGF-C and pLNCX-syk)into A549 cells with the liposomes. Tested the expression of VEGF-C mRNA and Syk mRNA with RT-PCR.Investigated the cell invasion assay with transwell chamber in vitro. Analysis the expression of VEGF-C proteins in A549 cells and detect Syk and VEGF-C proteins of 81 NSCLC tissue samples with immunohistochemical staining. Higher expression of VEGF-C was revealed in VEGF-C-construct-transfected A549 cell than that in controls through RT-PCR (P < 0.05) and immunohistochemistry(P < 0.01).RT-PCR also revealed that Syk expression was higher in Syk-construct-transfected cells than in controls (P < 0.05). The cell invading experiments showed that there was more invaded cells in both transfected terms than in controls (P < 0.05). The expression of Syk protein in NSCLC tissue were significantly lower than that in the normal lung tissue (P < 0.05), while the expression of VEGF-C protein in NSCLC tissue were significantly higher than that in the normal lung tissue(P < 0.05). The expression of Syk and VEGF-C has a negative correlationship (r = -1.000, P = 0.019). The expression of Syk and VEGF-C has a negative correlationship in NSCLC tissue, VEGF-C-construct-transfected A549 cells are more aggressive than Syk-construct-transfected cells. And they may cooperated with each other in the development of NSCLC.