Xun Ye

Identification and Validation of Long Noncoding RNA Biomarkers in Human Non–Small-Cell Lung Carcinomas

Introduction: Dysregulation of long noncoding RNAs (lncRNAs) has been regarded as a primary feature of several human cancers. However, the genome-wide expression and functional significance of lncRNAs in non–small-cell lung carcinomas (NSCLC) remains unclear. The aim of this study was to identify novel lncRNAs that may play an important role in contributing to NSCLC pathogenesis. Methods: We performed an integrative analysis of two NSCLC microarray datasets comprising 165 and 90 patients, respectively. The candidate lncRNAs were identified using the GSE19188 dataset, and then confirmed in the GSE18842 dataset. In addition, an independent cohort of 73 clinical samples was analyzed to validate the selected lncRNAs by quantitative real-time polymerase chain reaction analysis. Results: With microarray gene expression analysis, we identified and validated a list of 64 lncRNAs significantly dysregulated in NSCLC tumors compared with normal lung tissues; and a panel of 181 lncRNAs that were specific to histological subtypes of NSCLC (adenocarcinoma, large-cell carcinoma, and squamous cell carcinoma). The quantitative real-time polymerase chain reaction analysis of six selected lncRNAs in clinical samples further confirmed the results of microarray analysis. Conclusions: We have identified and validated multiple novel lncRNAs associated with tumorigenesis and histological differentiation in human NSCLC. These lncRNAs could be further exploited for the development of useful biomarkers in diagnosis, prognosis, and treatment of NSCLC.

Comparison of Hepatic Resection and Radiofrequency Ablation for Small Hepatocellular Carcinoma: A Meta-Analysis of 16,103 Patients

We performed a meta-analysis to evaluate the therapeutic effects of radiofrequency ablation (RFA) and surgical hepatic resection (HR) in the treatment of small hepatocellular carcinoma (HCC). Thirty-one studies were included in the analysis. A total of 16,103 patients were involved: 8,252 treated with RFA and 7,851 with HR. Compared to the RFA group, the 3, 5-year overall and disease-free survival rates in the HR group were significantly higher. On the other hand, complications were significantly fewer and hospital-stay was significantly shorter in the RFA group than in the HR group. In subgroup analyses, the overall and disease-free survival in the HR group were also significantly higher than those in the RFA group for HCCs ≤ 3 cm, whereas there were no significant differences between the two groups for HCCs ≤ 2 cm. Our analysis showed that although HR was associated with higher complication rate and longer hospital-stay, HR is proposed as the first-line treatment rather than RFA for patients with HCCs larger than 2 cm. For patients with HCCs of 2 cm or less, RFA may be an alternative to HR because of their comparable long-term efficacy.

Investigation of Variation in Gene Expression Profiling of Human Blood by Extended Principle Component Analysis

Background Human peripheral blood is a promising material for biomedical research. However, various kinds of biological and technological factors result in a large degree of variation in blood gene expression profiles. Methodology/Principal Findings Human peripheral blood samples were drawn from healthy volunteers and analysed using the Human Genome U133Plus2 Microarray. We applied a novel approach using the Principle Component Analysis and Eigen-R 2 methods to dissect the overall variation of blood gene expression profiles with respect to the interested biological and technological factors. The results indicated that the predominating sources of the variation could be traced to the individual heterogeneity of the relative proportions of different blood cell types (leukocyte subsets and erythrocytes). The physiological factors like age, gender and BMI were demonstrated to be associated with 5.3% to 9.2% of the total variation in the blood gene expression profiles. We investigated the gene expression profiles of samples from the same donors but with different levels of RNA quality. Although the proportion of variation associated to the RNA Integrity Number was mild (2.1%), the significant impact of RNA quality on the expression of individual genes was observed. Conclusions By characterizing the major sources of variation in blood gene expression profiles, such variability can be minimized by modifications to study designs. Increasing sample size, balancing confounding factors between study groups, using rigorous selection criteria for sample quality, and well controlled experimental processes will significantly improve the accuracy and reproducibility of blood transcriptome study.

PPAR signaling pathway may be an important predictor of breast cancer response to neoadjuvant chemotherapy

PurposeNeoadjuvant chemotherapy for advanced breast cancer may improve the radicality for a subset of patients, but others may suffer from severe adverse drug reactions without any benefit. To predict the responses to chemotherapy, we performed a phase II trial of neoadjuvant chemotherapy using a weekly PCb [paclitaxel (Taxol) plus carboplatin] regimen for stage II/III breast cancer and assessed the correlation between baseline gene expression and the tumor response to treatment.MethodsA total of 61 patients with stage II-III breast cancer were included and administered four cycles of preoperative PCb. We performed a gene expression analysis using Affymetrix HG-U133 Plus 2.0 GeneChip arrays in 31 breast cancer tissues. Differentially expressed genes (DEGs) were identified by the significance analysis of microarrays (SAM) program using a false discovery rate of 0.05. The Functional Annotation Tool in the DAVID Bioinformatics Resources was used to perform the gene functional enrichment analysis. The other 30 patients (15 pCR and 15 non-pCR patients) were available as an independent validation set to test the selected DEGs by quantitative real-time PCR analysis (qRT-PCR).ResultsBy analyzing six pathological complete response (pCR) patients and 25 patients with non-pCR, 300 probes (231 genes) were identified as differentially expressed between pCR and residual disease by the SAM program when the fold change was >2. The gene functional enrichment analysis revealed 15 prominent gene categories that were different between pCR and non-pCR patients, most notably the genes involved in the peroxisome proliferator-activated receptor (PPAR), DNA repair and ER signal pathways and in the immune-related gene cluster. The qRT-PCR analysis results for the genes in the PPAR pathway (LPL, SORBS1, PLTP, SCD5, MMP1 and CSTA) in independent validation set were consistent with the results from the microarray data analysis.ConclusionIn the present study, we identified a number of gene categories pertinent to the therapeutic response. We believe that the PPAR pathway may be an important predictor of genes that are involved in the chemotherapy response.

Identification and validation of a blood-based 18-gene expression signature in colorectal cancer

Purpose: The early detection of colorectal cancer (CRC) is crucial for successful treatment and patient survival. However, compliance with current screening methods remains poor. This study aimed to identify an accurate blood-based gene expression signature for CRC detection. Experimental Design: Gene expression in peripheral blood samples from 216 patients with CRC tumors and 187 controls was investigated in the study. We first conducted a microarray analysis to select candidate genes that were significantly differentially expressed between patients with cancer and controls. A quantitative reverse transcription PCR assay was then used to evaluate the expression of selected genes. A gene expression signature was identified using a training set (n = 200) and then validated using an independent test set (n = 160). Results: We identified an 18-gene signature that discriminated the patients with CRC from controls with 92% accuracy, 91% sensitivity, and 92% specificity. The signature performance was further validated in the independent test set with 86% accuracy, 84% sensitivity, and 88% specificity. The area under the receiver operating characteristics curve was 0.94. The signature was shown to be enriched in genes related to immune functions. Conclusions: This study identified an 18-gene signature that accurately discriminated patients with CRC from controls in peripheral blood samples. Our results prompt the further development of blood-based gene expression biomarkers for the diagnosis and early detection of CRC. Clin Cancer Res; 19(11); 3039–49. ©2013 AACR.

Decrease in Natural Killer cell associated gene expression as a major characteristic of the immune status in the bloodstream of colorectal cancer patients

Early detection and stratification of patients with colorectal cancer (CRC) are major challenges, particularly in the context of the development of new therapies. Several screening strategies are already in place in various countries, but compliance remains a major issue, mainly due to logistics or discomfort for the patients. In this study, we hypothesized that transcriptional signatures associated with leukocytes in peripheral blood can be informative to the identification of CRC patients. Gene expression was studied using RNA extracted from whole blood samples collected in PAXgene tubes and DNA microarrays. Analyzing 119 CRC patients and 101 colonoscopy-negative control (CNC) samples, we observed 327 differentially expressed genes (DEG), mostly associated with immune cell activation and trafficking. Natural Killer (NK) cell signaling and cytotoxicity associated genes appeared to undergo major changes in CRC peripheral blood samples. These changes were more pronounced in the advanced stages of the disease. A summarizing score of the expression of 10 genes related to NK cells interestingly revealed a marked heterogeneity within the CRC Stage IV group, suggesting possible further stratification of the patients. This study shows the potential of transcriptomics in peripheral blood to discover biomarkers and provides new insight on the immune response in colorectal cancer. In addition to preparing a possible alternative to current screening modalities, these results also show that the expression analysis of genes like those related to NK cells should allow the stratification of patients with colorectal cancer, opening the door to personalized medicine.

Identification and validation of a two-gene expression index for subtype classification and prognosis in Diffuse Large B-Cell Lymphoma

The division of diffuse large B-cell lymphoma (DLBCL) into germinal center B-cell-like (GCB) and activated B-cell-like (ABC) subtypes based on gene expression profiling has proved to be a landmark in understanding the pathogenesis of the disease. This study aims to identify a novel biomarker to facilitate the translation of research into clinical practice. Using a training set of 350 patients, we identified a two-gene expression signature, “LIMD1-MYBL1 Index”, which is significantly associated with cell-of-origin subtypes and clinical outcome. This two-gene index was further validated in two additional dataset. Tested against the gold standard method, the LIMD1-MYBL1 Index achieved 81% sensitivity, 89% specificity for ABC group and 81% sensitivity, 87% specificity for GCB group. The ABC group had significantly worse overall survival than the GCB group (hazard ratio = 3.5, P = 0.01). Furthermore, the performance of LIMD1-MYBL1 Index was satisfactory compared with common immunohistochemical algorithms. Thus, the LIMD1-MYBL1 Index had considerable clinical value for DLBCL subtype classification and prognosis. Our results might prompt the further development of this two-gene index to a simple assay amenable to routine clinical practice.

Gene Expression Analysis of Peripheral Blood Cells Reveals Toll-Like Receptor Pathway Deregulation in Colorectal Cancer

Colorectal cancer is the leading cause of cancer-related deaths worldwide. The disease is curable when detected at an early stage. However, the compliance rate with current screening recommendations remains poor. An accurate, minimally invasive blood test that has the potential for greater patient compliance would be a welcome addition to the current methods. Recent data have shown that gene expression profile of peripheral blood cells can reflect disease states and thus have diagnostic value. In this study, genome-wide gene expression profiling of peripheral blood cells from 20 healthy controls and 20 colorectal cancer patients were performed using PAXgene™ technology and Affymetrix GeneChip® microarrays. We identified a list of 1,469 genes that were differentially expressed between the healthy controls and cancer patients. Gene annotation and functional enrichment analysis revealed that those genes are mainly related to immune functions. Particularly, a set of genes belonging to the Toll-Like Receptor pathways were up-regulated in the colorectal cancer patients. These findings provide a new understanding of blood gene expression profile in colorectal cancer. Our result may serve as the basis for further development of blood biomarkers for the diagnosis and treatment of colorectal cancer.

Using peripheral blood mRNA signature to distinguish between breast cancer and benign breast disease in non-conclusive mammography patients

Due to the small volume and high density of breast tissue in Asian women, particularly younger women, mammographic diagnosis is sometimes non-conclusive, with a Breast Imaging Reporting and Data System (BI-RADS) result of 0. No alternative based on blood biomarkers has yet succeeded in discriminating between patients with breast cancer (BC) and those with benign breast disease (BBD) among BI-RADS 0 patients. In our study, 84 BC and 94 BBD patients with mammographic results and confirmed pathologic information were enrolled and categorized into two groups, namely, 79 BC and 73 BBD patients with BI-RADS 1-5 and 5 BC and 21 BBD patients with BI-RADS 0. RNA extracted from peripheral blood samples collected in PAXgeneTM tubes was analyzed after NuGEN WT-OvationTM RNA amplification using Affymetrix GeneChip

92-Gene Molecular Profiling in Identification of Cancer Origin: A Retrospective Study in Chinese Population and Performance within Different Subgroups

Background After cancer diagnosis, therapy for the patient is largely dependent on the tumor origin, especially when a metastatic tumor is being treated. However, cases such as untypical metastasis, poorly differentiated tumors or even a limited number of tumor cells may lead to challenges in identifying the origin. Moreover, approximately 3% to 5% of total solid tumor patients will not have to have their tumor origin identified in their lifetime. The THEROS CancerTYPE ID® is designed for identifying the tumor origin with an objective, rapid and standardized procedure. Methodology and Principal Findings This is a blinded retrospective study to evaluate performance of the THEROS CancerTYPE ID® in a Chinese population. In total, 184 formalin-fixed paraffin-embedded (FFPE) samples of 23 tumor origins were collected from the tissue bank of Fudan University Shanghai Cancer Center (FDUSCC). A standard tumor cell enrichment process was used, and the prediction results were compared with reference diagnosis, which was confirmed by two experienced pathologists at FDUSCC. All of the 184 samples were successfully analyzed, and no tumor specimens were excluded because of sample quality issues. In total, 151 samples were correctly predicted. The agreement rate was 82.1%. A Pearson Chi-square test shows that there is no difference between this study and the previous evaluation test performed by bioTheranostics Inc. No statistically significant decrease was observed in either the metastasis group or tumors with high grades. Conclusions A comparable result with previous work was obtained. Specifically, specimens with a high probability score (>0.85) have a high chance (agreement rate = 95%) of being correctly predicted. No performance difference was observed between primary and metastatic specimens, and no difference was observed among three tumor grades. The use of laser capture micro-dissection (LCM) makes the THEROS CancerTYPE ID® accessible to almost all of the cancer patients with different tumor statuses.