miR-195 Targets HDGF to inhibit proliferation and invasion of NSCLC cells

Non-small cell lung cancer (NSCLC) is one of the most common causes of cancer-related death worldwide. MicroRNAs (miRNAs) play critical roles in the development and progression of NSCLC. miR-195 acts as a tumor suppressor in several cancers, however, its role in NSCLC is not well understood. Herein, we found that miR-195 was significantly decreased in both NSCLC tissues and cell lines. Forced expression of miR-195 significantly suppressed proliferation, migration, and invasion of NSCLC cells. Hepatoma-derived growth factor (HDGF) was identified as a target of miR-195 in NSCLC cells. Overexpression of HDGF dramatically abolished the tumor suppressive role of miR-195 in NSCLC cells. Our results demonstrated a tumor suppressive role of miR-195 in NSCLC, and suggested a potential therapeutic target for NSCLC.     

MiR-186 targets ROCK1 to suppress the growth and metastasis of NSCLC cells

MicroRNAs (miRNAs) act as oncogenes or tumor suppressors in human cancers. Increasing evidence shows that deregulation of miRNAs contributes to the development and progression of human non-small cell lung cancer (NSCLC). Here, we identified miR-186 as a tumor suppressor in NSCLC, which was decreased in NSCLC. Overexpression of miR-186 significantly inhibited proliferation, migration, and invasion of NSCLC cells. In addition, Rho-associated protein kinase 1 (ROCK1) was identified as a target of miR-186 in NSCLC cells. Restoration of ROCK1 remarkably reversed the tumor-suppressive effects of miR-186 on cell proliferation, migration, and invasion in NSCLC cells. Furthermore, ROCK1 was inversely correlated with miR-186 expression in NSCLC. Collectively, our data indicate that miR-186 functions as tumor suppressor in NSCLC by targeting ROCK1.     

Prognostic significance of preoperative serum carcinoembryonic antigen in non-small cell lung cancer: a meta-analysis

Observational studies on the prognostic role of preoperative serum carcinoembryonic antigen (CEA) in non-small cell lung cancer (NSCLC) are controversial. Electronic databases updated until June 1, 2014 were searched to find relevant studies. A meta-analysis was conducted with eligible studies which quantitatively evaluated the relationship between preoperative serum CEA level and survival of patients with NSCLC. Survival data were aggregated and quantitatively analyzed. We performed a meta-analysis of 16 studies (n?=?4,296 patients) that evaluated the correlation between preoperative serum CEA level and survival in patients with NSCLC. Combined hazard ratios suggested that preoperative serum CEA overexpression was associated with poor prognosis of overall survival (OS) (hazard ratio (HR)?=?2.28, 95 % confidence interval (CI) 2.24–2.31) in patients with NSCLC. Meanwhile, for p-stage I NSCLC, the HR (95 % CI) was 1.98 (1.73–2.15). In the stratified analysis by patient source, significant risks were found among Asians and non-Asians. However, significant heterogeneity was observed among all studies. In the present meta-analysis, preoperative serum CEA overexpression indicates a poor prognosis for patients with NSCLC.     

Addition of bevacizumab enhances antitumor activity of erlotinib against non-small cell lung cancer xenografts depending on VEGF expression

Purpose

Erlotinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), and bevacizumab, an anti-vascular endothelial growth factor (VEGF) agent, are promising therapies for advanced non-small cell lung cancer (NSCLC). Our study was aimed to determine whether there were conditions under which the addition of bevacizumab would enhance the antitumor activity of erlotinib against NSCLC tumors in vitro and in vivo.

Methods

MTS was for NSCLC cell (PC9, 11–18, H1975, H157, H460 and A549) growth assay in vitro. ELISA was for VEGF protein assay in cells and tumor tissues. Mouse xenograft models were established with H157, H460 and A549 with primary resistance to erlotinib and treated with erlotinib plus bevacizumab or each agent alone. Erlotinib concentrations in tumors were determined by high-performance liquid chromatography.

Results

Bevacizumab alone did not inhibit NSCLC cell growth in vitro. In primarily erlotinib-resistant NSCLC cells, the levels of VEGF protein were highest in H157 cell followed in order by H460 and A549 cells. In vivo, bevacizumab alone significantly inhibited tumor growth only in xenograft models with high (H157) and/or moderate (H460) levels of VEGF protein. A combination of erlotinib and bevacizumab partially reversed resistance to erlotinib in H157 xenografts (high VEGF level) with increasing intratumoral erlotinib concentrations, but not in H460 (moderate) or A549 (low) xenografts.

Conclusions

These results support that combined with anti-VEGF therapy could enhance antitumor activity of anti-EGFR therapy and/or partially reverse resistance to EGFR TKI, by increasing EGFR TKI concentration in specific tumors that express high levels of VEGF protein.     

miR-195 inhibits the growth and metastasis of NSCLC cells by targeting IGF1R

MicroRNAs (miRNAs) are small, non-coding RNAs which act as oncogenes or tumor suppressors in multiple human cancers. Accumulating evidence reveals that aberrant expression of miRNAs contributes to the development and progression of non-small cell lung cancer (NSCLC). Here, we identified miR-195 as a tumor suppressor in NSCLC cells, whose expression level was dramatically decreased in both NSCLC tissues and cell lines. Ectopic expression of miR-195 suppressed NSCLC cell proliferation and metastasis-related traits in vitro. Insulin-like growth factor 1 receptor (IGF1R) was identified as a direct target of miR-195 in NSCLC cells. Furthermore, restoration of IGF1R remarkably attenuated the tumor suppressive effects of miR-195 on NSCLC cells. Our data suggest that miR-195 may be involved in the carcinogenesis of NSCLC partially by targeting IGF1R.     

Expression of TGFβ-1 and EHD1 correlated with survival of non-small cell Lung cancer

Transforming growth factor-β1 (TGFβ-1) signaling is regulated by endocytotic pathway. To clarify the prognostic value of TGFβ-1 and to verify the involvement of endocytosis in drug resistance, we examined the expression of TGFβ-1 and Eps15 homology domain 1 (EHD1) in non-small cell lung cancer (NSCLC) and its association with tumor characteristics and survival of patients with NSCLC. Expression of TGFβ-1 and EHD1 was evaluated by immunohistochemistry in paraffin sections from 105 NSCLC patients. Overall survival (OS) was analyzed by Kaplan–Meier method, log-rank test, and multivariate Cox proportional hazard regression model. Positive immunostaining of TGFβ-1 and EHD1 was detected in 52.38 and 39.05 % of NSCLC samples, respectively. In non-adjuvant chemotherapy-treated group (P?=?0.006) and epidermal growth factor receptor (EGFR) (+) group (P?=?0.038), patients with TGFβ-1 expression had a longer OS. EHD1 negative expression predicted a longer OS (P?=?0.003), especially in EGFR (+) (P?=?0.006) and adjuvant chemotherapy-treated patients (P?=?0.003). NSCLC patients with concurrent positive TGFβ-1 and negative EHD1 (combined markers) were significantly correlated with better OS (P?=?0.001). American Joint Committee on Cancer (AJCC) status and combined markers were independent prognostic indicators for OS (HR (95 % CI) 1.576 (1.112–2.232), P?=?0.011 and HR 0.349 (0.180–0.673), P?=?0.002, respectively). We identified concordant TGFβ-1 positive and EHD1 negative as a strong favorable prognosis factor in NSCLC. Our results may help us to select and optimize strategies for individualized therapy.     

Expression and clinical significance of CRABP1 and CRABP2 in non-small cell lung cancer

The impairment of retinoic acid (RA)-dependent signaling is a frequent event during carcinogenesis. Cellular retinoic acid-binding proteins (CRABP1 and CRABP2) are important modulators of RA activity. Up to date, the role of these proteins in cancer progression remains poorly investigated. Here, we studied for the first time the simultaneous messenger RNA (mRNA) and protein expression of CRABPs in non-small cell lung cancer (NSCLC) samples. CRABP1 and CRABP2 mRNA levels were elevated in 42 and 56 % of NSCLC samples, respectively. Decrease of CRABP2 mRNA expression was significantly associated with the presence of lymph node metastases. Protein expression of CRABP1 and CRABP2 was detected in 50 and 56 % of tumor samples, respectively. We also found a positive correlation between CRABP1 and CRABP2 expression. Taken together, we demonstrated significant changes in CRABP expression in NSCLC samples. Importantly, the presented data provide the first evidence of potential involvement of CRABP2 in lung cancer metastasis.     

Evaluation of gefitinib efficacy according to body surface area in patients with non-small cell lung cancer harboring an EGFR mutation

Background

Exon 19 deletions and L858R point mutation are the most commonly encountered active epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC), and they predict greater efficacy of gefitinib therapy. The objective of this study was to evaluate whether body surface area (BSA) affects the efficacy of gefitinib in patients with NSCLC harboring an active EGFR mutation.

Methods

We reviewed the medical records of consecutive patients with advanced NSCLC harboring an active EGFR mutation who received gefitinib monotherapy. The median BSA value was used as the cutoff value to evaluate the impact of BSA on the efficacy of gefitinib.

Results

The median BSA of the 103 NSCLC patients harboring an active EGFR mutation was 1.45 m². The overall response rate, progression-free survival (PFS), and median survival time (MST) were 65.0 %, 11.3 months, and 26.2 months, respectively. There were no significant differences in clinical outcomes between the high-BSA group (BSA ≥ 1.45 m²) and low-BSA group (BSA < 1.45 m²), i.e., the response rates was 60.0 % and 69.8 %, respectively (P = 0.20), and their MST was 24.7 and 26.2 months, respectively (P = 0.78). Although BSA was predictive of PFS between high-BSA group and low-BSA group in the univariate analysis (9.0 and 12.2 months, P = 0.04), the multivariate analysis identified only performance status and smoking status as independent predictors of PFS.

Conclusions

The efficacy of gefitinib in patients with NSCLC harboring an EGFR mutation does not differ according to their BSA.     

The Application of Real-Time PCR Technique to Detect Rare Cell Clones with Primary T790M Substitution of EGFR Gene in Metastases of Non-small Cell Lung Cancer to Central Nervous System in Chemotherapy Naive Patients

The time-limited efficacy of reversible EGFR-TKIs in patients with advanced non-small cell lung cancer (NSCLC) with EGFR gene activating mutations is associated with development of treatment resistance after some period of therapy. This resistance predominantly results from secondary mutations located in EGFR gene, especially T790M substitution. There is limited information available concerning the prevalence of primary T790M mutations in patients with metastatic NSCLC tumors before treatment with EGFR-TKIs. The aim of work was to assess the prevalence of de novo T790M mutations in EGFR gene in tissue samples from NSCLC metastatases in central nervous system (CNS) in both chemotherapy and EGFR-TKI naive NSCLC patients. We analyzed DNA samples isolated from paraffin-embedded tissue from CNS metastases for T790M mutations using real-time PCR and TaqMan probe against the T790M mutant sequence. The tissue samples were taken during palliative neurosurgery in 143 NSCLC patients. Amplification of the T790M-specific sequence was detected in 25 patients (17.5 %). The quantity of mutated DNA was less than 1 % in all samples with amplification, and in vast majority (20 patients, 14 % of all samples) it was even less that 0.1 %. In 5 patients (3.5 %) quantity of mutated DNA ranged from 0.1 to 1 % and true positive results of T790M mutation presence in these patients were most possible. Amplification of this sequence was not concurrent with common EGFR mutations and was not associated with sex, smoking status and pathological type of cancer. There is a possibility to detect the primary T790M mutation in brain metastases of NSCLC in EGFR-TKIs naïve patients.     

Prognostic value of CD44 and CD44v6 expression in patients with non-small cell lung cancer: meta-analysis

We sought to clarify the prognostic value of CD44 in survival of patients with non-small cell lung cancer (NSCLC). We performed a meta-analysis of relevant literature to aggregate the available survival results, using studies published in English until March 2014. Eligible studies dealt with CD44, CD44 standard form (CD44s) and CD44 variant 6 (CD44v6), assessment in NSCLC patients on primary lesions and reported survival data according to CD44 and CD44 isoforms expression. We aggregated 10 trials (5 trials for CD44v6, 3 trials for CD44, and 2 trials for CD44s) comprising 1,074 patients, in this meta-analysis. The combined hazard ratio (HR) with CD44v6 and CD44s was 2.39 (95 % confidence interval (CI) 1.69–3.37) and 1.64 (95 % CI 1.06–2.52), respectively. It associated high CD44v6 and CD44s expression with poor survival in NSCLC patients. However, CD44 overexpression did not significantly correlate with survival in patients with NSCLC (HR 1.44; 95 % CI 0.72–2.89). Our meta-analysis shows that CD44v6 and CD44s overexpression indicates poor prognosis for NSCLC patients. However, the high CD44 expression is not significantly correlated with survival for patients with NSCLC.     

A comparative analysis of EGFR mutation status in association with the efficacy of TKI in combination with WBRT/SRS/surgery plus chemotherapy in brain metastasis from non-small cell lung cancer

We proposed to identify the efficacy of an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) using whole brain radiotherapy (WBRT)/stereotactic radiosurgery (SRS)/surgery in brain metastases from patients with non-small cell lung cancer (NSCLC) and clarify the association between treatment outcome and EGFR gene mutation status. A total of 282 patients with NSCLC brain metastases who underwent WBRT/SRS/surgery alone or in combination with TKI were enrolled in our study from 2003–2013. Amplification mutation refractory system technology was used to determine the EGFR mutation status in 109 tissue samples. EGFR mutation detection was performed in 109 patients with tumor tissues. The EGFR positive rate was 50 % (55/109), including 26 exon 19 deletions and 24 L858R mutations. The median follow-up time was 28 months. The median overall survival, median progression-free survival of intracranial disease, and median progression-free survival of extracranial disease was significantly longer for patients with TKI treatment (31.9 vs 17.0 months, P < 0.0001; 19.8 vs 12.0 months, P < 0.0001; and 19.6 vs 12.3 months, P < 0.0001; respectively). In subgroup analysis within the TKI group, patients harboring EGFR mutations had better extracranial disease control (20.4 vs 14.1 months, P = 0.032). Administration of TKI agents with conventional therapy compared with conventional therapy alone might be beneficial for overall survival, progression-free survival of intracranial disease and progression-free survival of extracranial disease in patients with brain metastases from NSCLC independent of EGFR mutations.     

MMP-14 overexpression correlates with poor prognosis in non-small cell lung cancer

Matrix metalloproteinase-14 (MMP-14) has been demonstrated to play an important role in tumor progression. The aim of this study was to analyze the correlation between MMP-14 expression and clinicopathologic features and its prognostic significance in non-small cell lung cancer (NSCLC). Immunohistochemical staining for MMP-14 protein was performed in 104 patients with NSCLC. High levels of MMP-14 protein were positively correlated with the status of clinical stage (I–II vs. III–IV; P?P?P?=?0.014), and differentiated degree (high vs. low or undifferentiated; P?=?0.001). The patients with higher MMP-14 expression of protein had shorter survival time than patients with low MMP-14 expression. Multivariate analysis indicated that the level of MMP-14 expression was an independent prognostic indicator (P?NSCLC. In conclusion, MMP-14 is a potential unfavorable prognostic factor for patients with NSCLC.     

Gefitinib induces cytoplasmic translocation of the CDK inhibitor p27 and its binding to a cleaved intermediate of caspase 8 in non-small cell lung cancer cells

Background

The epidermal growth factor receptor (EGFR) represents one of the first rationally selected molecules for targeted therapy in non-small cell lung cancer (NSCLC). Gefitinib is a reversible and highly selective tyrosine kinase inhibitor that competitively blocks the binding of adenosine triphosphate to its binding site in the tyrosine kinase domain of the EGFR. It has been found that treatment with gefitinib induces cell cycle arrest and apoptosis in NSCLC cells harboring activating EGFR mutations. Despite its clinical relevance, however, the mechanism underlying gefitinib-induced apoptosis has remained largely unknown.

Methods

We used the gefitinib-sensitive NSCLC cell line HCC827, which harbors a deletion in exon 19 of the EGFR gene, to examine the effect of gefitinib on the apoptotic machinery.

Results

We found that gefitinib treatment caused the NSCLC cells to undergo apoptosis following activation of the caspase 8 cascade. Expression of p27, a cyclin-dependent kinase (CDK) inhibitor whose major target is the cyclin E/CDK2 complex, was found to increase during this process, and this increase was accompanied by translocation of p27 from the nucleus to the cytoplasm. Moreover, we found that cytoplasmic p27 bound to a cleaved intermediate (p43/p41) of caspase 8 and that inhibition of cytoplasmic translocation of p27 reduced gefitinib-induced cell death in HCC827 cells.

Conclusion

Based on our results, we conclude that gefitinib-induced apoptosis is mediated by the interaction of p27 and caspase 8 in NSCLC cells carrying an activating EGFR mutation.     

High Sam68 expression predicts poor prognosis in non-small cell lung cancer

Background

The nuclear protein Sam68 has been implicated in the oncogenesis and tumor growth. The aim of this study was to explore the clinical value of Sam68 in patients with non-small cell lung cancer (NSCLC).

Methods

We examined Sam68 expression in 50 NSCLC tissues and matched adjacent noncancerous tissues by quantitative RT-PCR (qRT-PCR) and Western blotting. Furthermore, the Sam68 protein expression was analyzed by immunohistochemistry in 208 NSCLC samples. Kaplan–Meier method and multivariate Cox regression model were used to evaluate the prognostic value of nuclear Sam68 expression in NSCLC for disease survival.

Results

The expression of Sam68 was significantly elevated in NSCLC tissues as compared with adjacent non-cancerous tissues (P < 0.01). The high expression of Sam68 in NSCLC was significantly correlated with lymph node metastasis and tumor TNM stage. Kaplan–Meier survival analysis revealed that high expression of Sam68 correlated with poor prognosis of NSCLC patients (P < 0.01). Multivariate analysis showed that Sam68 expression was an independent prognostic marker for overall survival of NSCLC patients (HR 2.73, 95 % CI 1.549–4.315, P = 0.002).

Conclusion

Our results suggest that high Sam68 expression predicts poor prognosis of NSCLC patients, and Sam68 may be potentially a prognostic biomarker for NSCLC.     

The programmed cell death 6 interacting protein insertion/deletion polymorphism is associated with non-small cell lung cancer risk in a Chinese Han population

It has been proposed that genetic factors contribute to the susceptibility of non-small cell lung cancer (NSCLC). The programmed cell death 6 interacting protein (PDCD6IP) encodes for a protein that has been known to bind to the products of the PDCD6 gene, a required protein in apoptosis. The aim of this study is to investigate the relationship between PDCD6IP insertion/deletion (I/D) polymorphism (rs28381975) and NSCLC risk in a Chinese population. A population-based case–control study was conducted in 449 NSCLC patients and 512 cancer-free controls. The genotype of the PDCD6IP gene was determined by using a polymerase chain reaction assay. The promoter activity was analyzed by luciferase reporter assay in A549 and H1299 cells. Statistically significant difference was observed when the patients and controls were compared according to ID + II versus DD (OR?=?1.72, 95 % CI 1.29–2.31, P?NSCLC risk (OR?=?1.41, 95 % CI 1.18–1.69, P?PDCD6IP I/D polymorphism significantly increased advanced NSCLC risk (OR?=?2.06, 95 % CI 1.30–3.26, P?P?=?0.001). The results from this study suggested that PDCD6IP I/D polymorphism was potentially related to NSCLC susceptibility in Chinese Han population.     

MicroRNAs as novel biomarkers in the diagnosis of non-small cell lung cancer: a meta-analysis based on 20 studies

The detection of microRNAs (miRNAs), particularly those obtained from the bloodstream, is an emerging method for diagnosing human cancers, including non-small cell lung cancer (NSCLC). However, studies on the accuracy of miRNAs detection in diagnosing NSCLC have yield inconsistent conclusions, making it necessary to conduct a meta-analysis to systematically evaluate the diagnostic value of miRNAs in the diagnosis of NSCLC. The Medline, Embase, Chinese National Knowledge Infrastructure (CNKI), and Sinomed electronic databases were searched to identify all related articles evaluating the diagnostic value of miRNAs for NSCLC. A bivariate regression model was used to calculate the pooled diagnostic accuracy estimates. A total of 20 articles were included in this meta-analysis, involving 1,563 NSCLC patients and 1,060 healthy controls. Overall, our bivariate random effects meta-analysis yielded area under curve (AUC) of 0.85 (95 % CI: 0.82–0.88) with sensitivity of 76 % (95 CI: 72–80) and specificity of 80 % (95 % CI: 77–84) for the use of miRNAs in differentiating NSCLC patients from healthy controls. In addition, subgroup and meta-regression analyses revealed that a combination of multiple miRNAs (AUC, sensitivity, and specificity of 0.89, 81, and 84, respectively) had a higher diagnostic accuracy than single miRNA-based assays (AUC, sensitivity, and specificity of 0.81, 73, and 77 %, respectively). Furthermore, a comparison of miRNAs expression patterns between blood and sputum samples provides additional evidence that miRNAs obtained from blood (AUC, sensitivity, and specificity of 0.86, 78, and 80 %, respectively) are more credible diagnostic biomarkers than those from sputum (AUC, sensitivity, and specificity of 0.79, 66, and 79 %, respectively). In summary, the current meta-analysis suggests that the detection of miRNAs may be used in the future as an initial screening test for NSCLC, particularly, the detection of a combination of multiple miRNAs, which is a more comprehensive indicator than individual miRNAs.     

ARMC8α promotes proliferation and invasion of non-small cell lung cancer cells by activating the canonical Wnt signaling pathway

ARMC8 proteins are novel armadillo repeat containing proteins, which are well conserved in eukaryotes and are involved in a variety of processes such as cell migration, proliferation, tissue maintenance, signal transduction, and tumorigenesis. Armadillo repeat proteins include well-known proteins such as β-catenin and p120ctn. Our current knowledge of ARMC8, especially its role in cancer, is limited. In this study, we quantified ARMC8 expression in 112 non-small cell lung cancer (NSCLC) tissues and adjacent non-cancerous tissues, and seven lung cancer cell lines using immunohistochemistry staining and Western blotting. ARMC8 level was significantly higher in NSCLC tissues than in the adjacent normal tissues (67.9 % versus 5.4 %, p?p?=?0.022), lymph node metastasis (p?=?0.001), and poor prognosis (p?NSCLC patients. Cox regression analysis demonstrated that ARMC8 was an independent prognostic factor for NSCLC. Consistent with this, ARMC8α downregulation by siRNA knockdown inhibited growth, colony formation, and invasion in A549 lung cancer cells, while ARMC8α overexpression promoted growth, colony formation, and invasion in H1299 lung cancer cells. In addition, ARMC8α knockdown downregulated canonical Wnt-signaling pathway activity and cyclin D1 and matrix metalloproteinase (MMP)-7 expression. Consistent with this, ARMC8α overexpression upregulated canonical Wnt-signaling pathway activity and cyclin D1 and MMP-7 expression. These results indicate that ARMC8α upregulates cyclin D1 and MMP7 expression by activating the canonical Wnt-signaling pathway and thereby promoting lung cancer cell proliferation and invasion. Therefore, ARMC8 might serve as a novel therapeutic target in NSCLC.