Podocalyxin as a Prognostic Marker in Gastric Cancer

Background

Podocalyxin-like 1 (PODXL) is a cell-adhesion glycoprotein associated with aggressive tumor phenotype and poor prognosis in several forms of cancer. The aim of this study was to investigate PODXL expression in gastric cancer by use of two different antibodies.

Methods

By tumor-tissue microarrays and immunohistochemistry we evaluated PODXL expression in tumor specimens from 337 patients who underwent surgery for gastric adenocarcinoma at Helsinki University Hospital. We used two different antibodies: HPA2110, which is a polyclonal antibody and an in-house monoclonal antibody called HES9, to investigate the association of PODXL expression with clinicopathologic variables and patient survival.

Results

PODXL staining was positive by the polyclonal antibody in 153 (57.5%) cases and by the monoclonal antibody in 212 (76%). Polyclonal antibody expression was associated with intestinal cancer type (p<0.001). Monoclonal antibody staining was associated with age over 66 (p = 0.001), with intestinal cancer (p<0.001), and with small tumor size (≤ 5 cm; p = 0.024). Both antibodies were associated with high S-phase fraction (p = 0.022; p = 0.010), and high tumor proliferation index (Ki-67; p = 0.003; p = 0.001). PODXL positivity by the polyclonal antibody indicated reduced gastric-cancer-specific 5-year survival of 24.0% (95% CI 16.9–31.1), compared to 43.3% (95% CI 33.7–52.9) for patients with PODXL negativity (p = 0.001). The result remained significant in multivariable analysis (HR = 3.17; 95% CI 1.37–7.34, p = 0.007).

Conclusion

In gastric cancer, PODXL expression by the polyclonal antibody HPA2110 is an independent marker of poor prognosis.     

Cancer Stem Cell-Related Gene Periostin: A Novel Prognostic Marker for Breast Cancer

We investigated the expression status of periostin in breast cancer stem cells and its clinical implications in order to lay a foundation for managing breast cancer. CD44+/CD24−/line- tumor cells (CSC) from clinical specimens were sorted using flow cytometry. Periostin expression status was detected in CSC cells and 1,086 breast cancer specimens by Western blot and immunohistochemistry staining, with the CSC ratio determined by immunofluorescence double staining. The relationship between the periostin protein and clinico-pathological parameters and prognosis was subsequently determined. As a result, CSC cells are more likely to generate new tumors in mice and cell microspheres that are deficient in NOD/SCID compared to the control group. Periostin protein was expressed higher in CSC cells compared to the control cells and was found to be related to CSC chemotherapy resistance. Moreover, periostin expression was found to be related to the CSC ratio in 1,086 breast cancer specimens (P = 0.001). In total, 334 (30.76%) of the 1,086 breast cases showed high periostin expression. After universal and Spearman regression correlation analysis, periostin was observed to be related to histological grade, CSC ratio, lymph node metastasis, tumor size, and triple-negative breast cancer (all P<0.05). Furthermore, periostin was shown to attain a significantly more distant bone metastasis and worse disease-specific survival than those with none or low-expressed periostin protein (P = 0.001). In the Cox regression test, periostin protein was detected as an independent prognostic factor (P = 0.001). In conclusion, periostin was found to be related to the CSC and an independent prognostic factor for breast cancer. It is also perhaps a potential target to breast cancer.     

PTK 7 Is a Transforming Gene and Prognostic Marker for Breast Cancer and Nodal Metastasis Involvement

Protein Tyrosin Kinase 7 (PTK7) is upregulated in several human cancers; however, its clinical implication in breast cancer (BC) and lymph node (LN) is still unclear. In order to investigate the function of PTK7 in mediating BC cell motility and invasivity, PTK7 expression in BC cell lines was determined. PTK7 signaling in highly invasive breast cancer cells was inhibited by a dominant-negative PTK7 mutant, an antibody against the extracellular domain of PTK7, and siRNA knockdown of PTK7. This resulted in decreased motility and invasivity of BC cells. We further examined PTK7 expression in BC and LN tissue of 128 BC patients by RT-PCR and its correlation with BC related genes like HER2, HER3, PAI1, MMP1, K19, and CD44. Expression profiling in BC cell lines and primary tumors showed association of PTK7 with ER/PR/HER2-negative (TNBC-triple negative BC) cancer. Oncomine data analysis confirmed this observation and classified PTK7 in a cluster with genes associated with agressive behavior of primary BC. Furthermore PTK7 expression was significantly different with respect to tumor size (ANOVA, p = 0.033) in BC and nodal involvement (ANOVA, p = 0.007) in LN. PTK7 expression in metastatic LN was related to shorter DFS (Cox Regression, p = 0.041). Our observations confirmed the transforming potential of PTK7, as well as its involvement in motility and invasivity of BC cells. PTK7 is highly expressed in TNBC cell lines. It represents a novel prognostic marker for BC patients and has potential therapeutic significance.     

BRG1 Is a Prognostic Marker and Potential Therapeutic Target in Human Breast Cancer

BRG1, a core component of the SWI/SNF chromatin-remodeling complex, has been implicated in cancer development; however, the biological significance of BRG1 in breast cancer remains unknown. We explored the role of BRG1 in human breast cancer pathogenesis. Using tissue microarray and immunohistochemistry, we evaluated BRG1 staining in 437 breast cancer specimens and investigated its role in breast cancer cell proliferation, migration and invasion. Our Kaplan-Meier survival curves showed that high BRG1 expression is inversely correlated with both overall (P = 0.000) and disease-specific (P = 0.000) 5-year patient survival. Furthermore, we found that knockdown of BRG1 by RNA interference markedly inhibits cell proliferation and causes cessation of cell cycle. This reduced cell proliferation is due to G1 phase arrest as cyclin D1 and cyclin E are diminished whereas p27 is upregulated. Moreover, BRG1 depletion induces the expression of TIMP-2 but reduces MMP-2, thereby inhibiting the ability of cells to migrate and to invade. These results highlight the importance of BRG1 in breast cancer pathogenesis and BRG1 may serve as a prognostic marker as well as a potentially selective therapeutic target.     

MiR-942-3p as a Potential Prognostic Marker of Gastric Cancer Associated with AR and MAPK/ERK Signaling Pathway

Gastric cancer is a common tumor with high morbidity and mortality. MicroRNA (miRNA) can regulate gene expression at the translation level and various tumorigenesis processes, playing an important role in tumor occurrence and prognosis. This study aims to screen miRNA associated with gastric cancer prognosis as biomarkers and explore the regulatory genes and related signaling pathways. In this work, R language was used for the standardization and differential analysis of miRNA and mRNA expression profiles. Samples were randomly divided into a testing group and a training group. Subsequently, we built the five miRNAs (has-miR-9-3p, has-miR-135b-3p, has-miR-143-5p, has-miR-942-3p, has-miR-196-3p) prognostic modules, verified and evaluated their prediction ability by the Cox regression analysis. They can be used as an independent factor in the prognosis of gastric cancer. By predicting and analyzing potential biological functions of the miRNA target genes, this study found that the AR gene was not only a hub gene in the PPI network, but also associated with excessive survival of patients. In conclusion, this study demonstrated that hsa-miR-942-3p could be a potential prognostic marker of gastric cancer associated with the AR and MAPK/ERK signaling pathways. The results of this study provide insights into the occurrence and development of gastric cancer.     

MiR-371-5p Facilitates Pancreatic Cancer Cell Proliferation and Decreases Patient Survival

Background

microRNAs (miRNAs) play a critical role in tumorigenesis, either as a tumor suppressor or as an oncogenic miRNA, depending on different tumor types. To date, scientists have obtained a substantial amount of knowledge with regard to miRNAs in pancreatic cancer. However, the expression and function of miR-371-5p in pancreatic cancer has not been clearly elucidated. The aim of this study was to investigate the roles of miR-371-5p in pancreatic cancer and its association with the survival of patients with pancreatic cancer.

Methods

The expression of miR-371-5p was examined in pancreatic duct adenocarcinoma (PDAC) and their adjacent normal pancreatic tissues (ANPT) or in pancreatic cancer cell lines by qRT-PCR. The association of miR-371-5p expression with overall survival was determined. The proliferation and apoptosis of SW-1990 and Panc-1 cells, transfected with miR-371-5p mimics or inhibitor, were assessed using MTT assay and flow cytometry, respectively. The tumorigenicity was evaluated via mice xenograft experiments. miR-371-5p promoter interactions were analyzed by chromatin immunoprecipitation assays (ChIP). Protein expression was analyzed by Western blot.

Results

The expression level of miR-371-5p was dramatically upregulated in clinical PDAC tissues compared with ANPT. Patients with high miR-371-5p expression had a significantly shorter survival than those with low miR-371-5p expression. The in vitro and in vivo assays showed that overexpression of miR-371-5p resulted in cell proliferation and increased tumor growth, which was associated with inhibitor of growth 1 (ING1) downregulation. Interestingly, we also found that ING1, in turn, inhibited expression of miR-371-5p in the promoter region.

Conclusions

our study demonstrates a novel ING1-miR-371-5p regulatory feedback loop, which may have a critical role in PDAC. Thus miR-371-5p can prove to be a novel prognostic factor and therapeutic target for pancreatic cancer treatment.     

Galectin-3 as a Marker and Potential Therapeutic Target in Breast Cancer

Galectin-3 has a relatively high level of expression in triple-negative breast cancers and is a potential marker for this disease. However, the clinical and prognostic implications of galectin-3 expression in breast cancer remain unclear. We examined mastectomy specimens from 1086 breast cancer cases and matching, adjacent non-cancerous tissues using immunohistochemistry. Overall, triple-negative breast cancers expressed galectin-3 more strongly than did other breast cancers types (63.59% vs 21.36%, P = 0.001). Galectin-3 expression was not found to be an independent prognostic factor for breast cancer by Cox regression analysis, but was associated with chemotherapeutic resistance. Apoptosis was only weakly induced by arsenic trioxide (ATO) treatment in galectin-3-positive breast cancer cells (MDA-MB-231 and MCF-7), although ATO treatment up-regulated galectin-3 expression. Knockdown of galectin-3 in MDA-MB-231 cells sensitized them to killing by ATO. These findings support a possible role for galectin-3 as a marker for triple-negative breast cancer progression and as a therapeutic target in combination with ATO treatment, although the mechanisms that underlie this synergy require further investigation.     

A Prognostic Gene Signature for Metastasis-Free Survival of Triple Negative Breast Cancer Patients

Although triple negative breast cancers (TNBC) are the most aggressive subtype of breast cancer, they currently lack targeted therapies. Because this classification still includes a heterogeneous collection of tumors, new tools to classify TNBCs are urgently required in order to improve our prognostic capability for high risk patients and predict response to therapy. We previously defined a gene expression signature, RKIP Pathway Metastasis Signature (RPMS), based upon a metastasis-suppressive signaling pathway initiated by Raf Kinase Inhibitory Protein (RKIP). We have now generated a new BACH1 Pathway Metastasis gene signature (BPMS) that utilizes targets of the metastasis regulator BACH1. Specifically, we substituted experimentally validated target genes to generate a new BACH1 metagene, developed an approach to optimize patient tumor stratification, and reduced the number of signature genes to 30. The BPMS significantly and selectively stratified metastasis-free survival in basal-like and, in particular, TNBC patients. In addition, the BPMS further stratified patients identified as having a good or poor prognosis by other signatures including the Mammaprint® and Oncotype® clinical tests. The BPMS is thus complementary to existing signatures and is a prognostic tool for high risk ER-HER2- patients. We also demonstrate the potential clinical applicability of the BPMS as a single sample predictor. Together, these results reveal the potential of this pathway-based BPMS gene signature to identify high risk TNBC patients that can respond effectively to targeted therapy, and highlight BPMS genes as novel drug targets for therapeutic development.     

Plasma Protein Profiling Reveal Osteoprotegerin as a Marker of Prognostic Impact for Colorectal Cancer

BACKGROUND: Due to difficulties in predicting recurrences in colorectal cancer stages II and III, reliable prognostic biomarkers could be a breakthrough for individualized treatment and follow-up. OBJECTIVE: To find potential prognostic protein biomarkers in colorectal cancer, using the proximity extension assays. METHODS: A panel of 92 oncology-related proteins was analyzed with proximity extension assays, in plasma from a cohort of 261 colorectal cancer patients with stage II-IV. The survival analyses were corrected for disease stage and age, and the recurrence analyses were corrected for disease stage. The significance threshold was adjusted for multiple comparisons. RESULTS: The plasma proteins expression levels had a greater prognostic relevance in disease stage III colorectal cancer than in disease stage II, and for overall survival than for time to recurrence. Osteoprotegerin was the only biomarker candidate in the protein panel that had a statistical significant association with overall survival (P = .00029). None of the proteins were statistically significantly associated with time to recurrence. CONCLUSIONS: Of the 92 analyzed plasma proteins, osteoprotegerin showed the strongest prognostic impact in patients with colorectal cancer, and therefore osteoprotegerin is a potential predictive marker, and it also could be a target for treatments.     

Intratumoral CD8+ Cytotoxic Lymphocyte Is a Favorable Prognostic Marker in Node-Negative Breast Cancer

Background

The prognostic effect of tumor infiltrating CD8⁺ cytotoxic lymphocytes (CTLs) in breast cancer is controversial. We analyzed the association between CD8⁺ CTLs and survival of untreated node-negative breast cancer patients.

Material and Methods

CD8⁺ CTLs infiltrate was evaluated by immunostaining in a cohort of 332 node-negative breast cancer patients with a median follow-up of 152 months. The prognostic significance of CD8⁺ CTLs for disease-free survival (DFS) and breast cancer-specific overall survival (OS) was evaluated with Kaplan-Meier survival analysis as well as univariate analysis and multivariate Cox analysis adjusted for age at diagnosis, pT stage, histological grade, estrogen receptor (ER) status, progesterone receptor (PR) status, Ki-67 expression and human epidermal growth factor receptor 2 (HER-2) status.

Results

285 (85.8%) patients showed strong CD8⁺ CTLs infiltrate positive status. Univariate analysis showed that CD8⁺ CTLs had statistically significant association with DFS (P = 0.004, hazard ratio [HR] = 0.454, 95% confidence interval [CI] = 0.265–0.777) and OS (P = 0.014, HR = 0.430, 95% CI = 0.220–0.840) in the entire cohort. The significance of CD8⁺ CTLs was especially strong in ER negative, HER-2 negative and ER, PR, HER-2 triple-negative breast cancers. In Kaplan-Meier analysis, CD8⁺ CTLs had significant effect on prognosis of patients (Log-rank test: P = 0.003 for DFS and P = 0.011 for OS), independent of established clinical factors for DFS (P = 0.002, HR = 0.418, 95% CI = 0.242–0.724) as well as for OS (P = 0.009, HR = 0.401, 95% CI = 0.202–0.797).     

Presence of CHD1L Over-Expression Is Associated with Aggressive Tumor Biology and Is a Novel Prognostic Biomarker for Patient Survival in Human Breast Cancer

Background

The chromodomain helicase/adenosine triphosphatase DNA binding protein 1–like gene (CHD1L) is a recently identified oncogene localized at 1q21. CHD1L protein over-expression in primary hepatocellular carcinoma is correlated with enhanced apoptosis inhibition, reduced chemosensitivity and shortened patient survival. However, CHD1L protein status or mRNA expression in breast cancer and its clinical significance remain obscure.

Material and Methods

In this study, immunohistochemical staining for CHD1L expression was performed on tissue microarrays containing 179 primary invasive breast cancers and 65 matched normal breast tissue specimens. Clinico-pathological features were collected and compared between different CHD1L statuses. Kaplan-Meier curves were applied to estimate disease-free survival (DFS) and overall survival (OS). Cox regression was used to identify independent prognostic factors. Also, quantitative real-time polymerase chain reaction (QRT-PCR) was employed to evaluate the mRNA level expression of CHD1L in six breast cancer cell lines.

Results

Presence of CHD1L over-expression was observed in 87 of the 179 patients (48.6%), which associated with a younger age (P = 0.011), higher grade (P = 0.004), higher Ki-67 index (P = 0.018) and HER2 over-expression/amplification (P = 0.037). After a median follow-up of 55 months, patients with presence of CHD1L over-expression had significantly poorer DFS (82.6% Vs 76.3%, P = 0.035), but not OS (87.0% Vs 94.9%, P = 0.439). In multivariate analysis, CHD1L status (HR = 2.169, [95%CI, 1.029–4.573], P = 0.042), triple negative subtype (HR = 2.809, [95%CI 1.086–7.264], P = 0.033) and HER2 positive subtype (HR = 5.221, [95%CI 1.788–15.240], P = 0.002) were identified as independent prognostic factors for DFS. In vitro study indicated that relative mRNA expression level of CHD1L was higher in breast cancer cell lines, especially in MDA-MB-231 and LM2-4175, when compared to normal breast epithelial cell line.

Conclusions

Presence of CHD1L over-expression is probably associated with aggressive tumor biology in breast cancer. CHD1L status might be a novel prognostic biomarker for patients with breast cancer.     

c-Met as a Prognostic Marker in Gastric Cancer: A Systematic Review and Meta-Analysis

Background

c-Met has been recognized as an important therapeutic target in gastric cancer, but the prognostic property of the c-Met status is still unclear. We aimed to characterize the prognostic effect of c-Met by systematic review and meta-analysis.

Methods

We identified 15 studies assessing survival in gastric cancer by c-Met status. Effect measure of interest was hazard ratio (HR) for survival. Meta-regression was performed to estimate the relationship between HR and disease stage. Random-effects meta-analyses were used to account for heterogeneity.

Results

15 eligible studies provided outcome data stratified by c-Met status in 2210 patients. Meta-analysis of the HRs indicated a significantly poorer Os in patients with high c-Met expression (average HR=2.112, 95%CI: 1.622–2.748). Subgroup analysis showed the prognostic effect of c-Met was identical in protein-level and gene-level based methodology. The same effect was also seen in Asian and Western ethnicity subgroup analysis. Meta-regression showed HR was not associated with disease stage.

Conclusions

Patients with tumors that harbor high c-Met expression are more likely to have a worse Os, with this prognostic effect independent of disease stage. c-Met status should be evaluated in clinical prognosis.     

A Meta-Analysis of SMAD4 Immunohistochemistry as a Prognostic Marker in Colorectal Cancer

AIM: SMAD4 immunohistochemistry is considered a valuable prognostic marker in colorectal cancer, but individual studies have often been small and the results variable. A meta-analysis could potentially clarify these findings. METHODS: In September 2014, a Pubmed and Google Scholar search was conducted to find publications that reported the prognostic value of SMAD4 expression. A meta-analysis was performed to clarify the association between SMAD4 expression and survival outcomes. RESULTS: 137 studies were found, of which 13 were considered eligible. The studies consisted of a total of 3800 patients. Three different endpoints were taken into account, namely, overall survival (OS), disease-free survival (DFS), and cancer-specific survival (CSS). In addition, the studies were divided into univariate and multivariate analyses. The pooled hazard ratios were given as follows: univariate CSS = 1.75 [95% confidence interval (CI): 0.93-3.32; z= 1.69; P= .09]; multivariate CSS = 2.17 (95% CI: 1.56-3.01; z= 4.65; P= .000); univariate DFS = 2.11 (95% CI: 1.36-3.28; z= 3.32; P= .001); multivariate DFS = 2.15 (95% CI: 1.56-3.01; z= 4.65; P= .000); univariate OS and DFS = 2.30 (95% CI: 1.41-3.73; z= 3.36; P= .001); univariate OS = 2.28 (95% CI: 1.30-4.00; z= 2.89; P= .004). CONCLUSION: The results of the presented meta-analyses indicate that SMAD4 expression status using immunohistochemistry is a prognostic marker for patient survival.     

Study of Integrated Heterogeneous Data Reveals Prognostic Power of Gene Expression for Breast Cancer Survival

BackgroundStudies show that thousands of genes are associated with prognosis of breast cancer. Towards utilizing available genetic data, efforts have been made to predict outcomes using gene expression data, and a number of commercial products have been developed. These products have the following shortcomings: 1) They use the Cox model for prediction. However, the RSF model has been shown to significantly outperform the Cox model. 2) Testing was not done to see if a complete set of clinical predictors could predict as well as the gene expression signatures.Methodology/FindingsWe address these shortcomings. The METABRIC data set concerns 1981 breast cancer tumors. Features include 21 clinical features, expression levels for 16,384 genes, and survival. We compare the survival prediction performance of the Cox model and the RSF model using the clinical data and the gene expression data to their performance using only the clinical data. We obtain significantly better results when we used both clinical data and gene expression data for 5 year, 10 year, and 15 year survival prediction. When we replace the gene expression data by PAM50 subtype, our results are significant only for 5 year and 15 year prediction. We obtain significantly better results using the RSF model over the Cox model. Finally, our results indicate that gene expression data alone may predict long-term survival.Conclusions/SignificanceOur results indicate that we can obtain improved survival prediction using clinical data and gene expression data compared to prediction using only clinical data. We further conclude that we can obtain improved survival prediction using the RSF model instead of the Cox model. These results are significant because by incorporating more gene expression data with clinical features and using the RSF model, we could develop decision support systems that better utilize heterogeneous information to improve outcome prediction and decision making.     

Prostate Tumor Overexpressed 1 (PTOV1) Is a Novel Prognostic Marker for Nasopharyngeal Carcinoma Progression and Poor Survival Outcomes

Background

Prostate tumor overexpressed 1 (PTOV1) has been reported to contribute to increased cancer proliferation. However, the clinical significance of PTOV1 in the development and progression of nasopharyngeal carcinoma (NPC) is unclear. Our study aimed to investigate the expression pattern of PTOV1 in NPC and its correlation with clinicopathological features of patients.

Methods

Western blotting and real-time PCR were conducted to examine PTOV1 expression levels in NPC cell lines and biopsy tissues compared with normal controls. Immunohistochemistry (IHC) was performed to analyze PTOV1 protein expression in paraffin-embedded tissues from 123 patients. Statistical analyses were applied to evaluate the clinical significance of PTOV1 expression.

Results

PTOV1 mRNA and protein levels were upregulated in NPC cell lines and clinical samples. IHC analyses showed that PTOV1 was highly expressed in 68 (55.3%) of 123 NPC specimens. Statistical analysis revealed that PTOV1 expression was significantly correlated with clinical stage (P < 0.001), T classification (P = 0.042) and N classification (P = 0.001). Patients with a higher PTOV1 expression had shorter overall survival compared with those with a lower PTOV1 expression level, especially in lower N stage patients. Multivariate analyses suggested that PTOV1 expression was an independent prognostic marker for survival in NPC patients.

Conclusions

Our data demonstrated that PTOV1 overexpression is associated with poor survival outcomes of NPC patients, especially in N0-1 patients. Hence, PTOV1 may help to detect early lymph node metastasis of NPC patients and serve as an independent prognostic biomarker for human NPC.