Galectin-4, a novel predictor for lymph node metastasis in lung adenocarcinoma.
ABSTRACT: Metastasis is still a major issue in cancer, and the discovery of biomarkers predicting metastatic capacity is essential for the development of better therapeutic strategies for treating lung adenocarcinoma. By using a proteomic approach, we aimed to identify novel predictors for lymph node metastasis in lung adenocarcinoma. Two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis showed 6 spots differentially expressed between lymph node metastasis-positive and lymph node metastasis-negative groups in a discovery set. Subsequent mass spectrometry showed that 2 of these spots were derived from galectin-4, and western blot analysis confirmed the overexpression of galectin-4 in metastatic samples. The predictive value of galectin-4 was confirmed by immunohistochemical analysis for a validation set consisting of 707 surgically resected specimens of lung adenocarcinomas (stages I to IV). We observed that 148 lung adenocarcinomas (20.9%) expressed galectin-4, which was significantly associated with variables of disease progression such as tumor size (p<0.0001), pleural invasion (p = 0.0071), venous invasion (p = 0.0178), nodal status (p = 0.0007), and TNM stage (p<0.0001). By the multivariate analysis, Galectin-4 expression was revealed as one of the independent predictor for lymph node metastasis, together with solid predominant and micropapillary histologic pattern. Furthermore, galectin-4 expression was revealed to be an independent predictor for lymph node metastasis and an adverse survival factor in patients with lung adenocarcinoma of acinar predominant type. Galectin-4 plays an important role in metastatic process of lung adenocarcinoma. Immunohistochemical testing for galectin-4 expression may be useful together with the detection of specific histology to predict the metastatic potential of lung adenocarcinoma.
Project description:Lymph node metastasis is still an important issue in metastatic process of lung adenocarcinoma. C-C chemokine receptor 7 (CCR7) has been proved to be closely associated with the metastasis of lung adenocarcinoma, and the mechanism is poorly understood. In order to investigate the relationship between CCR7 and lymph node metastasis in lung adenocarcinoma, and to explore the role of CCR7 in treating lung adenocarcinoma, 40 clinical specimens were collected to define the relationship between CCR7 and lymph node metastasis in lung adenocarcinoma by immunohistochemistry. The siRNA was used to suppress CCR7 expression in A549 cells. The scratch test, transwell test, qRT-PCR, western blot, flow cytometry and immunofluorescence were used to investigate the lymph node metastasis-related function of CCR7 in vitro. The athymic mice subcutaneous injection was used to research lung adenocarcinoma formation in vivo. Clinical case studies show that higher expression of CCR7 in lung adenocarcinoma tissues was associated with a higher lymph node metastasis. Inhibition of expression of CCR7 can reduce the migration and invasion and suppress the expression of VEGF-C, VEGF-D and VEGF-R3 in vitro and in vivo. Moreover, CCR7 silence also suppressed WNT and p-ERK pathways in vitro. All the results indicate that CCR7 can promote lymph node metastasis in lung adenocarcinoma by regulating VEGF-C/D-R3 pathway. Thus CCR7 is proposed to be a potential prediction for poor prognosis of lung adenocarcinoma, and a therapeutic target for lymph node metastasis.
Project description:Previous studies demonstrated that a subpopulation of cancer cells, which are CD133 positive (CD133+) feature higher invasive and metastatic abilities, are called cancer stem cells (CSCs). By using tumor cells derived from patients with lung adenocarcinoma, we found that galectin-1 is highly overexpressed in the CD133+ cancer cells as compared to the normal cancer cells (CD133-) from the same patients. We overexpressed galectin-1 in CD133- cancer cells and downregulated it in CSCs. We found that overexpression of galectin-1 promoted invasiveness of CD133- cells, while knockdown of galectin-1 suppressed proliferation, colony formation and invasiveness of CSCs. Furthermore, tumor growth was significantly inhibited in CSCs xenografts with knockdown of galectin-1 as compared to CSCs treated with scramble siRNAs. Biochemical studies revealed that galectin-1 knockdown led to the suppression of COX-2/PGE2 and AKT/mTOR pathways, indicating galectin-1 might control the phenotypes of CSCs by regulating these signaling pathways. Finally, a retrospective study revealed that galectin-1 levels in blood circulation negatively correlates with overall survival and positively correlates with lymph node metastasis of the patients. Taken together, these findings suggested that galectin-1 plays a major role on the tumorigenesis and invasiveness of CD133+ cancer cells and might serve as a potential therapeutic target for treatment of human patients with lung adenocarcinoma.
Project description:Lymph node metastasis of lung cancer is a serious problem. Therefore, there is a need for a detailed transcriptome study of metastatic lung adenocarcinoma. The lung adenocarcinoma RNA-seq data and the corresponding clinical information available from TCGA were analyzed. Differential expression, gradient changes, and biological pathways were carried out. Potential gene(s) associated with tumor metastasis and survival were validated by Cox regression. A total of 406 and 439 differentially expressed genes were identified for lymph node metastasis and TNM stages, respectively. Of the 296 intersection genes, 112 were associated with nodal metastasis and/or staging. Only 25 of these 112 genes with gradient changes were involved in nodal metastasis, and 13 were involved in staging. Only one gene, RN7SL494P, might be involved in lung adenocarcinoma development and poor outcome. Finally, Cox regression results verified that age, pathology classification, radiotherapy and chemotherapy are all the independent prognostic factors. In particular, RN7SL494P was further verified to be an independent factor affecting lymph node metastasis and patient survival. Furthermore, we verified the RN7SL494P function using simulation data generated by mixing cell lines of the Cancer Cell Line Encyclopedia (CCLE) and obtained consistent results. Our findings suggest a potential clinical application of the RN7SL494P as a promising marker in the evaluation of patients with primary lung adenocarcinoma, not only for predicting nodal metastasis, but also for the prognosis of the outcome.
Project description:BACKGROUND:It is difficult to predict lymph node metastasis in patients with early lung cancer. Pure ground glass opacity (GGO) on computed tomography indicates an early-stage adenocarcinoma that can be removed by limited resection or lobectomy without the need for mediastinal lymph node dissection or sampling, and lung adenocarcinoma with GGO therefore has a good prognosis. We examined the incidence and risk factors of lymph node metastasis in patients with clinical stage IA lung adenocarcinoma. METHODS:We retrospectively analyzed clinical data for 327 patients with stage IA peripheral lung cancer treated in our hospital from March 2014 to December 2018. The patients were divided into four groups according to computed tomography signs. Lobectomy and systematic lymph node dissection were performed in all patients. Correlations between lymph node metastasis and clinical pathological factors were analyzed by logistic regression. RESULTS:Among the 327 patients, 26 (7.95%) had lymph node metastasis. No patients with pure GGO or GGO-dominant types had lymph node metastasis. Logistic regression identified tumor diameter, solid content, plasma carcinoembryonic antigen (CEA) level, pathological type, lymphovascular invasion, and pleural invasion as factors related to the presence of lymph node metastasis. CONCLUSIONS:Tumor diameter, solid component ratio, plasma CEA level, pathological type, vascular tumor thrombus, and pleural invasion are possible independent risk factors for lymph node metastasis in patients with stage IA lung adenocarcinoma. In contrast, lymph node metastasis is rare in patients with pure GGO or GGO-dominant lung adenocarcinoma.
Project description:To identify patients in whom systematic lymph node dissection would be suitable, preoperative diagnosis of the biological invasiveness of lung adenocarcinomas through the classification of these T1aN0M0 lung adenocarcinomas into several subgroups may be warranted. In this retrospective study, we sought to determine predictive factors of lymph node status in clinical stage T1aN0M0 lung adenocarcinomas.We retrospectively reviewed the records of 273 consecutive patients undergone surgical resection of clinical stage T1aN0M0 lung adenocarcinomas at Shanghai Chest Hospital, from January 2011 to December 2012. Preoperative computed tomography findings of all 273 patients were reviewed and their tumors categorized as pure GGO, GGO with minimal solid components (<5 mm), part-solid (solid parts >5 mm), or purely solid. Relevant clinicopathologic features were investigated to identify predictors of hilar or mediastinal lymph node metastasis using univariate or multiple variable analysis.Among the 273 eligible clinical stage T1aN0M0 lung adenocarcinomas examined on thin-section CT, 103 (37.7%) were pure GGO, 118 (43.2%) GGO with minimal solid components, 13 (4.8%) part-solid (solid parts >5 mm, five GGO predominant and eight solid predominant), and 39 (14.3%) pure solid. There were 18 (6.6%) patients with lymph node metastasis. Incidence of N1 and N2 nodal involvement was 11 (6.6%) and seven (2.6%) patients, respectively. All patients with pure GGO and GGO with minimal solid components (<5 mm) tumors were pathologically staged N0. Multivariate analyses showed that the following factors significantly predicted lymph node metastasis for T1a lung adenocarcinomas: symptoms at presentation, GGO status, and abnormal carcinoembryonic antigen (CEA) titer. Multivariate analyses also showed that the following factors significantly predicted lymph node metastasis for pure solid tumors: air bronchogram sign, tumor size, symptoms at presentation, and abnormal CEA titer.The patients of clinical stage T1aN0M0 lung adenocarcinomas with pure GGO and GGO with minimal solid components tumors were pathologically staged N0 and systematic lymph node dissection should be avoided. But systematic lymph node dissection should be performed for pure solid tumors or part-solid, especially in patients with CEA greater than 5 ng/mL or symptoms at presentation, because of the high possibility of lymph node involvement.
Project description:The landscape of genetic alterations in lung adenocarcinoma derived from Asian patients is largely uncharacterized. Here we present an integrated genomic and transcriptomic analysis of 335 primary lung adenocarcinomas and 35 corresponding lymph node metastases from Chinese patients. Altogether 13 significantly mutated genes are identified, including the most commonly mutated gene TP53 and novel mutation targets such as RHPN2, GLI3 and MRC2. TP53 mutations are furthermore significantly enriched in tumours from patients harbouring metastases. Genes regulating cytoskeleton remodelling processes are also frequently altered, especially in metastatic samples, of which the high expression level of IQGAP3 is identified as a marker for poor prognosis. Our study represents the first large-scale sequencing effort on lung adenocarcinoma in Asian patients and provides a comprehensive mutational landscape for both primary and metastatic tumours. This may thus form a basis for personalized medical care and shed light on the molecular pathogenesis of metastatic lung adenocarcinoma.
Project description:BACKGROUND:Intralymphatic tumors in the extratumoral area are considered to represent the preceding phase of lymph node metastasis. The aim of this study was to clarify the biological properties of intralymphatic tumors susceptible to the development of lymph node metastasis, with special reference to the expression of cancer initiating/stem cell (CIC/CSC) related markers in cancer cells and the number of infiltrating stromal cells. MATERIAL AND METHODS:Primary lung adenocarcinomas with lymphatic permeation in the extratumoral area were retrospectively examined (n?=?107). We examined the expression levels of CIC/CSC related markers including ALDH1, OCT4, NANOG, SOX2 and Caveolin-1 in the intralymphatic cancer cells to evaluate their relationship to lymph node metastasis. Moreover, the number of infiltrating stromal cells expressing CD34, ?-smooth muscle actin, and CD204 were also evaluated. RESULTS:Among the intralymphatic tissues, low ALDH1 expression in cancer cells, high SOX2 expression in cancer cells, and a high number of CD204(+) macrophages were independent predictive factors for lymph node metastasis (P?=?0.004, P?=?0.008, and P?=?0.028, respectively). Among these factors, only low ALDH1 expression in cancer cells was significantly correlated with the farther spreading of lymph node metastasis (mediastinal lymph node, pathological N2) (P?=?0.046) and the metastatic lymph node ratio (metastatic/resected) (P?=?0.028). On the other hand, in the primary tumors, ALDH1 expression in the cancer cells was not associated with lymph node metastasis. Intralymphatic cancer cells expressing low ALDH1 levels exhibited lower E-cadherin expression levels than cancer cells with high levels of ALDH1 expression (P?=?0.015). CONCLUSIONS:Intralymphatic cancer cells expressing low levels of ALDH1 and infiltrating macrophages expressing CD204 have a critical impact on lymph node metastasis. Our study also highlighted the significance of evaluating the biological properties of intralymphatic tumors for tumor metastasis.
Project description:Background: To investigate the relationship between CXCR4-related circular RNAs (circRNAs) in exosomes and lymph node metastasis of lung adenocarcinoma. Methods: Totally 41 lung adenocarcinoma tissues (21 with lymph node metastasis and 20 without) were collected. Expression of CXCR4 protein was detected by western blotting analysis. A stable PC9/CXCR4-shRNA and PC14/CXCR4-shRNA knockdown lung adenocarcinoma cell lines were established and subjected to functional assays (cell proliferation, colony formation, migration and invasion) for phenotype changes. Exo-hsa-circRNAs (has-circRNAs in exosomes) were detected in vivo and in vitro. The diagnostic value of differentially expressed exo-has-circRNAs was evaluated. Results: Expression levels of CXCR4 were higher in patients with lymph node metastasis than in those without (P = 0.001). Silencing CXCR4 expression in PC9 and PC14 cell lines with short hairpin RNA could effectively abolish colony formation frequency, proliferation rate, migration rate, and the number of invasive cells (all P < 0.001). Exo_circRNA_0056616 was detected in both PC-9/CXCR4-shRNA cells and lung adenocarcinoma plasma at significantly higher levels than in the corresponding control (P < 0.001). When a receiver operating characteristic (ROC) curve for plasma exo-hsa_circRNA_0056616 levels and diagnosis of lymph node metastasis of lung adenocarcinoma was generated, a cutoff value of 0.394 was identified with an area under the curve of 0.812 (95% confidence interval 0.720-0.903), a sensitivity of 0.792, and specificity of 0.810. Conclusions: Taken together, our findings suggested that CXCR4 was higher in the lung adenocarcinoma tissues with lymph node metastasis. Higher plasma levels of exo-hsa_circRNA_0056616 in these patients also suggest that this circRNA represents a potential biomarker for lymph node metastasis predictor in lung adenocarcinoma.
Project description:PURPOSE:We conducted genome-wide miRNA-sequencing (miRNA-seq) in primary cancer tissue from patients of lung adenocarcinoma to identify markers for the presence of lymph node metastasis. EXPERIMENTAL DESIGN:Markers for lymph node metastasis identified by sequencing were validated in a separate cohort using quantitative PCR. After additional validation in the The Cancer Genome Atlas (TCGA) dataset, functional characterization studies were conducted in vitro. RESULTS:MiR-31 was upregulated in lung adenocarcinoma tissues from patients with lymph node metastases compared with those without lymph node metastases. We confirmed miR-31 to be upregulated in lymph node-positive patients in a separate patient cohort (P = 0.009, t test), and to be expressed at higher levels in adenocarcinoma tissue than in matched normal adjacent lung tissues (P < 0.0001, paired t test). MiR-31 was then validated as a marker for lymph node metastasis in an external validation cohort of 233 lung adenocarcinoma cases of the TCGA (P = 0.031, t test). In vitro functional assays showed that miR-31 increases cell migration, invasion, and proliferation in an ERK1/2 signaling-dependent manner. Notably, miR-31 was a significant predictor of survival in a multivariate cox regression model even when controlling for cancer staging. Exploratory in silico analysis showed that low expression of miR-31 is associated with excellent survival for T2N0 patients. CONCLUSIONS:We applied miRNA-seq to study microRNomes in lung adenocarcinoma tissue samples for the first time and potentially identified a miRNA predicting the presence of lymph node metastasis and survival outcomes in patients of lung adenocarcinoma.
Project description:Non-small cell lung cancer (NSCLC), including adenocarcinoma and squamous cell carcinoma, is the leading cause of death from lung malignancies and has a poor prognosis due to metastasis. Suppressor of cytokine signalling-2 (SOCS2), a feedback inhibitor of cytokine signalling, has been shown to be involved in growth control. Here, we show that SOCS2 were significantly downregulated in tumour foci in NSCLC patients. The expression levels of SOCS2 significantly correlated with clinical stage, lymph node metastasis, histological subtype and survival time. In particular, the decreased expression of SOCS2 significantly associated with advanced pathological stage, lymph node metastasis and shorter overall survival in lung adenocarcinoma patients. In vivo animal results showed that overexpressed SOCS2 attenuated the metastatic characteristics of lung adenocarcinoma, including by inhibiting the epithelial-mesenchymal transition (EMT). Further functional studies indicated that insulin-like growth factor 1 (IGF1)-driven migratory and invasive behaviours of lung adenocarcinoma cells can be partially suppressed by exogenous SOCS2 expression. Investigations into the mechanism of action revealed that SOCS2 inhibits EMT by inactivating signal transducer and activator of transcription 3 (STAT3) and STAT5 via the competitive binding of SOCS2 to the STAT binding sites on IGF1R. Altogether, our results reveal an important role for SOCS2 dysregulation in the pathogenicity of lung adenocarcinoma, suggest its potential use as a biomarker for diagnosing lung adenocarcinoma, and paves the way to develop novel therapy targets as the axis of SOCS2-IGF1R-STAT in lung adenocarcinoma.