Molecular and bioinformatics analysis of long non-coding RNAs in cervical cancer.

BackgroundIt has been demonstrated by studies globally that RNA binding proteins (RBPs) took part in the development of cervical cancer (CC). Few studies concentrated on the correlation between RBPs and overall survival of CC patients. We retrieved significant DEGs (differently expressed genes, RNA binding proteins) correlated to the process of cervical cancer development.MethodsExpressions level of genes in cervical cancer and normal tissue samples were obtained from GTEx and TCGA database. Differently expressed RNA binding proteins (DEGs) were retrieved by Wilcoxon sum-rank test. ClusterProfiler package worked in R software was used to perform GO and KEGG enrichment analyses. Univariate proportional hazard cox regression and multivariate proportional hazard cox regressions were applied to identify DEGs equipped with prognostic value and other clinical independent risk factors. ROC curve was drawn for comparing the survival predict feasibility of risk score with other risk factors in CC patients. Nomogram was drawn to exhibit the prediction model and validated by C-index and calibration curve. Correlations between differentially expressed RNA binding proteins (DEGs) and other clinical features were investigated by t test or Cruskal Wallis analysis. Correlation between Immune and DEGs in cervical cancer was investigated by ssGSEA.Results347 differentially expressed RBPs (DEGs) were retrieved from cervical cancer tissue and normal tissue samples. GO enrichment analysis showed that these DEGs involved in RNA splicing, catabolic process and metabolism. Cox regression model showed that there were ten DEGs significantly associated with overall survival of cervical cancer patients. WDR43 (HR = 0.423, P = 0.008), RBM38 (HR = 0.533, P < 0.001), RNASEH2A (HR = 0.474, P = 0.002) and HENMT1 (HR = 0.720, P = 0.071) played protective roles in survival among these ten genes. Stage (Stage IV vs Stage I HR = 3.434, P < 0.001) and risk score (HR = 1.214, P < 0.001) were sorted as independent prognostic risk factors based on multivariate cox regression. ROC curve validated that risk score was preferable to predict survival of CC patients than other risk factors. Additionally, we found some of these ten predictor DEGs were correlated significantly in statistic with tumor grade or stage, clinical T stage, clinical N stage, pathology or risk score (all P < 0.05). Part of immune cells and immune functions showed a lower activity in high risk group than low risk group which is stratified by median risk score.ConclusionOur discovery showed that many RNA binding proteins involved in the progress of cervical cancer, which could probably serve as prognostic biomarkers and accelerate the discovery of treatment targets for CC patients.

6036 Background: Diversity of the gut microbiome is associated with response rates for patients receiving immunotherapy. Studies investigating the gut microbiome and outcomes in cancer patients often do not adjust for confounding patient and tumor characteristics. We sought to identify independent gut microbial risk factors in cervical cancer (CC) patients receiving chemoradiation (CRT) and to evaluate their impact on survival. Methods: We analyzed baseline 16S rDNA fecal microbiomes of CC patients receiving standard CRT. Patient and tumor characteristics were analyzed by univariate and multivariate Cox regression models for Recurrence-free survival (RFS) and Overall survival (OS) based on univariate p-value&gt;0.2. Characteristics included age, body mass index (BMI), race, stage, grade, histology, nodal status, and max tumor size. Alpha (within sample) diversity was evaluated using Shannon diversity index (SDI). Kaplan-Meier curves were generated for patients with normal BMI and overweight/obese BMI based on Cox analysis. Results: 55 CC patients were included. Univariate analysis identified older age (Hazard Ratio (HR) of 0.93 (95% CI = 0.87-0.98, p = 0.0096)), SDI (HR of 0.51 (95% CI = 0.23-1.1, p = 0.087)) and BMI (HR of 0.92 (95% CI = 0.84-1, p = 0.096)) as risk factors for RFS. Multivariate survival analyses identified BMI and SDI as independent prognostic factors for RFS with a HR of 0.87 (95% CI = 0.77-0.98, p = 0.02) and 0.36 (95% CI = 0.15-0.84, p = 0.018) respectively. For OS, multivariate survival analyses again identified BMI and SDI as independent prognostic factors with a HR of 0.78 (95% CI = 0.623-0.97, p = 0.025) and 0.19 (95% CI = 0.043-0.83, p = 0.028) respectively. Conclusions: Gut diversity is a significant factor for predicting OS in CC patients undergoing CRT when BMI is accounted for, and may help explain the “obesity paradox” in cancer response. Studies exploring the relationship between gut diversity, CRT, and treatment efficacy are needed to further understand the role of the gut microbiome in treatment outcomes. [Table: see text]

Background: Lactate dehydrogenase (LDH) is a marker of injury and disease as it is expressed extensively in numerous cell types and tissues. Moreover it is released during tissue breakdown, and is elevated in cancerous tissues. However, the clinical significance and prognostic value of LDH as a tumor marker have been subject to considerable discussion. Objective: In this study, clinical serum LDH data from patients with cervical cancer (CC), CC microarray data, and RNA-seq data were integrated to assess the expression of LDH in CC. Methods: A total of 204 patients with newly diagnosed CC and 204 age-matched healthy controls were included to evaluate serum LDH levels in CC and non-cancer samples. External microarrays and RNA-seq datasets were collected for the differential expression analysis of LDH in CC and non-cancer tissue samples. Kaplan-Meier survival curves of the prognostic value of LDH for CC were plotted for RNA-seq data. Functional enrichment analysis was performed for the genes co-expressed with LDH. Results: The data from our in-house clinical cases as well as the data extracted from microarrays and RNA-seq databases demonstrated significant overexpression of LDH in CC samples. Elevated LDH expression levels were associated with poor overall survival in CC patients. The genes co-expressed with LDH were significantly correlated with the biological processes and pathways, associated with nuclear division, the condensed chromosome, protein serine/threonine kinase activity, and the cell cycle. Conclusion: In conclusion, LDH upregulation might serve as a therapeutic and prognostic biomarker for CC.

Objective:Deregulation of long noncoding RNAs (lncRNAs) is involved in the initiation and progression of cancer. LncRNA DLX6-AS1 is regarded as an oncogene in many cancer types. However, the clinical role of serum exosomal lncRNA DLX6-AS1 in cervical cancer (CC) is poorly known. This study aimed to analyze the diagnostic and prognostic value of serum exosomal lncRNA DLX6-AS1 in CC.Methods:A total of 114 patients with CC, 60 patients with CIN (cervical intraepithelial neoplasia), and 110 healthy women were enrolled in this study. Real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to measure the serum exosomal lncRNA DLX6-AS1 levels in all participants.Results:Serum exosomal lncRNA DLX6-AS1 level was significantly elevated in CC patients compared with CIN patients and normal controls. In addition, high serum exosomal lncRNA DLX6-AS1 expression was positively associated with lymph node metastasis, differentiation, FIGO stage, and shortened survival. Patients with high serum exosomal lncRNA DLX6-AS1 expression were more prone to have a relapse. Furthermore, univariate and multivariate analyses suggested that serum exosomal lncRNA DLX6-AS1 was a potential prognostic indicator for overall survival of CC patients.Conclusions:These findings demonstrated that serum lncRNA DLX6-AS1 might serve as a promising marker for the diagnosis and prognosis prediction of CC.

Integrated bioinformatic analysis of miR-15a/16-1 cluster network in cervical cancer

Long non-coding RNA TTN-AS1 promotes cell growth and metastasis in cervical cancer via miR-573/E2F3

Long non-coding RNA NORAD upregulate SIP1 expression to promote cell proliferation and invasion in cervical cancer

ObjectiveTumor microenvironment (TME) and the expression of immune-related genes (IRGs) are closely related to the development of cervical cancer (CC). This study aims to explore some IRGs as prognostic biomarkers for CC patients based on TME.MethodsThe abundance of tumor-infiltrating immune cells in CC samples was assessed using single-sample gene set enrichment analysis (ssGSEA). Thus, two immune-related groups are generated according to the immune status. The differentially expressed genes were discovered based on the grouping. Then, univariate Cox and LASSO regression analyses were performed using the R package glmnet. Five IRG prognostic signatures (HLA-DMA, DMBT1, CXCR6, CX3CL1, and SEMA3A) were established after that. The protein expression of some genes was verified by immunohistochemistry (IHC).ResultsThe signature of the five IRGs was identified to be an independent prognostic indicator for the overall survival in CC patients. A prognostic risk model was also constructed. CC patients were classified into high- and low-risk groups based on the median risk score. The survival time of patients in the high-risk group was shorter than that of those in the low-risk group. The five genes remarkably related to prognosis were screened, among which HLA-DMA, CXCR6, and CX3CL1 were the protective factors, whereas DMBT1 and SEMA3A were the risk factors. GO and KEGG enrichment analyses showed that the biomarkers of the five IRGs were enriched in the receptor-ligand interaction and chemokine signaling pathway. Moreover, CXCR6 expression was significantly correlated with immune cell infiltration among the five hub genes. IHC results demonstrated that the expression of SEMA3A protein level was increased, and CX3CL1 protein level was decreased in cervical cancer tissue.ConclusionImmune-related prognostic biomarkers in CC include HLA-DMA, DMBT1, CXCR6, CX3CL1, and SEMA3A. The risk score for the five genes is more accurate than that for other clinical risk factors in predicting prognosis at 3 and 5 years. The higher the risk score is, the worse the prognosis of CC patients is. Five prognostic biomarkers may participate in regulating TME through chemokine-mediated signaling pathways and receptor-ligand interactions. These findings provide new insights into the immunotherapy of CC.

The intratumoral microbe-host interaction plays crucial role in the development of cancer. The microbiome can influence cancer development by modulating inflammation, immune responses and metabolic pathways. Therefore, we aim to delineate the landscape and role of intratumoral microbiota in cervical cancer (CC). The presence of bacterial community in CC tissues was confirmed by fluorescence in situ hybridization (FISH). Then 16s rRNA and RNA-Seq were used to characterize the composition of intratumoral microbiota. Combined with cervical squamous cell carcinoma (CESC) data from the Tumor Cancer Genome Atlas (TCGA), the clinical signatures of intratumoral microbiota and DEGs were further analyzed. Finally, the effect of the up-regulated Fibrinogen beta chain (FGB) expressed fragment peptide on the biological behavior of cancer was verified in vitro. We found the composition heterogeneity of bacteria in CC tumors. Pseudomonas was most highly enriched in CC tissues and grouped according to the relative abundance level. The clinical characteristics of patients with relatively high abundance of Pseudomonas had the higher levels of fibrinogen and lower levels of white blood cell (WBC) and albumin (ALB) expression. Combining transcriptome data from the two our collective CC and TCGA-CESC cohorts, we found that Pseudomonas abundance was significantly associated with fibrinogen beta peptide expression and worse overall survival in CC patients. In vitro experiment revealed that Pseudomonas could promote the proliferation and migration of cervical cancer cells through overexpression of FGB. We characterized the composition of the intratumoral microbiota in CC tissues and identified the most significantly differentially abundant bacteria between cancerous and non-cancerous tissues. Our findings provide novel insights into the relationship between intratumoral Pseudomonas and the tumorigenesis of CC. A deeper understanding of the tumor microenvironment and its associated microbiota may reveal new potential therapeutic targets and improve clinical outcomes.

Introduction: Cervical cancer (CC) is the fourth most common cancer type and a leading cause of cancer-related deaths in women worldwide. Its underlying molecular mechanisms are unclear. Cancer cell-derived extracellular vesicles (EVs) are involved in cancer development and progression by delivering regulatory factors, including microRNAs and long non-coding RNAs (lncRNAs). Methods: Here, we identified the EV lncRNA expression profiles associated with different developmental stages of CC using next-generation sequencing. EVs from the serum of patients with stages I–III CC and healthy donors were characterized using EV marker immunoblotting and transmission electron microscopy. Results: The EV concentration increases with progression of the disease. Most particles had a 100–250-nm diameter, and their sizes were similar in all groups. We identified many lncRNAs that were uniquely and differentially expressed (DE) in patients with different stages of CC. The pathway analysis results indicated that the upregulated DE EV lncRNAs abundant in stages I and II were associated with cell proliferation and inflammation and cancer progression pathways, respectively. LINC00941, LINC01910, LINC02454, and DSG2-AS1 were highly expressed, suggesting poor overall survival of CC patients. Interestingly, DSG2-AS1 was associated with the human papillomavirus infection pathway through AKT3, DLG1, and COL6A2 genes. Conclusion: This is the first study that reports the levels of EVs and their lncRNA contents change during cancer development, demonstrating the existence of a unique vesicle-mediated cell-to-cell communication network underlying cancer progression.

Cervical cancer (CC) is one of the leading causes of cancer-related deaths among females in Ghana. Despite the magnitude of the public health challenge posed by CC in Ghana, survival data as well as reported incidence and mortality rates are primarily based on studies conducted in the capital city of the country. Even though age at diagnosis is known to affect the overall survival of CC patients, the role of this factor in the prognosis of CC patients in Ghana has not been sufficiently explored. The aim of this study was to determine the 5-year survival rate of Ghanaian woman treated for CC at a large tertiary healthcare facility in Ghana. This research was a single-institution-based quantitative retrospective cohort study conducted among patients with histopathologically confirmed CC. Clinical and socio-demographic data were retrieved from patients' medical records. Data analysis was done using the Statistical Package for the Social Sciences software version 23. Kaplan Meier curves were used to present the survival rates and median survival time. The peak age at diagnosis was between 45 and 80 years with the modal age group of patients between 75 and 80 years. The mean age at diagnosis was 63.3 ± 15.7 years ranging from 27 to 104 years. The overall survival rates at 1, 3 and 5 years were 76.5%, 51.5% and 32.4%, respectively. The median survival time was 65.8 months. Age < 50 years was associated with higher survival estimates than age >50 years. The 5-year overall survival rate of CC patients reported in this study (32.4%) is relatively low compared with countries in the developed world but like previous reports at other healthcare facilities in Ghana as well as in other underdeveloped countries.

LncRNA SNHG14 promotes the progression of cervical cancer by regulating miR-206/YWHAZ

PurposeFamily with sequence similarity 83 member H (FAM83H) plays key roles in tumorigenesis. However, the specific roles of FAM83H in cervical cancer (CC) have not been well studied.Materials and MethodsThe RNA-seq data of 306 CC tissues and three normal samples downloaded from The Cancer Genome Atlas were used to analyze the expression of FAM83H. The Kaplan-Meier method was used to draw survival curves. Associations between FAM83H expression and clinicopathological factors were analyzed by chi-square test. Cox proportional hazards model was used to analyze prognostic factors. Loss-of-function assays were conducted to discover the biological functions of FAM83H in cell proliferation, colony formation, invasion, and migration. Real-time Quantitative Reverse Transcription PCR (qRT-PCR) and Western blotting were used to measure the expression levels of FAM83H in CC cell lines.ResultsOur results demonstrated that FAM83H is overexpressed in CC tissues and that high FAM83H expression is associated with worse overall survival (OS). High FAM83H expression in CC was associated with clinical stage, pathologic tumor, and pathologic node. Univariate analysis suggested that FAM83H expression was significantly related to the OS of CC patients. Although multivariate analysis showed that FAM83H expression was not an independent prognostic factor for the OS of CC patients, the effects of FAM83H on CC cell growth and motility was significant. Loss-of-function experiments demonstrated that knockdown of FAM83H inhibited proliferation, colony formation, migration, and invasion of CC cells by inactivating PI3K/AKT pathway.ConclusionFAM83H might play a crucial role in CC progression and could act as a novel therapeutic target in CC.

Emerging studies have demonstrated that Prostate transmembrane protein androgen induced 1 (PMEPA1) plays crucial roles in the carcinogenesis of many developing human tumors. However, the clinical significance of PMEPA1 expression in cervical cancer (CC) and its contribution to cancer immunity have not been investigated. In this study, we identified PMEPA1 as a survival-related gene in CC based on TCGA datasets. Univariate and multivariate analysis showed that PMEPA1 expression was an independent predictor for overall survival in CC patients. We could observe a strong negative correlation between PMEPA1 expression and PMEPA1 methylation. Two CpG sites of PMEPA1 were associated with overall survival, and one CpG site of PMEPA1 was associated with progression-free survival. The low level of PMEPA1 methylation was associated with advanced clinical stage of CC patients. KEGG assays revealed the genes associated with PMEPA1 expression were mainly enriched in several tumor-related pathways. Increased PMEPA1 expressions were observed to be positively related to high immune infiltration levels in several immune cells. Finally, the pan-cancer assays revealed that PMEPA1 expression was associated with the overall survival of UVM, PAAD, LUSC, BLCA, CESC, and LUAD. Taken together, PMEPA1 is a prognosis-related biomarker for multiple cancer types, especially CC. PMEPA1 is involved in tumor immunity, suggesting PMEPA1 may be a potential immunotherapeutic target in CC.

Long noncoding RNAs (lncRNAs) are aberrantly expressed in various cancers. The purpose of this study was to determine the association of lncRNA (BLACAT1) with the prognosis of cervical cancer (CC) patients, and to further investigate the potential mechanisms of BLACAT1 function in CC progression. The expressions of BLACAT1 in CC tissues and cells were estimated by quantitative Real-time polymerase chain reaction (qRT-PCR). We compared the expression of BLACAT1 with the clinicopathological characteristics and survival of CC patients. MTT, colony formation, and transwell assay were performed to explore the effects of BLACAT1 expression on growth, migration, and invasion of CC cells. Protein levels of β-catenin and MMP-7 were evaluated by Western blotting. We found that BLACAT1 expression was significantly increased in CC tissues and cells lines. In addition, the expression level of BLACAT1 was positively correlated with distant metastasis (p=0.001), FIGO stage (p=0.010), and histological grade (p=0.012). Moreover, patients with high BLACAT12 expression had shorter overall survival and progression-free survival time than those with low BLACAT1 expression, with the data provided by multivariate analysis suggesting that BLACAT1 expression could serve as an independent prognostic factor in CC patients. Functionally, in vitro assay indicated that down-regulation of BLACAT1 significantly suppressed CC cells proliferation, migration, and invasion. Mechanistically, the results of Western blot showed that the expression of β-catenin and MMP-7 was significantly down-regulated in CC cells transfected with si-BLACAT1. These findings suggested that BLACAT1, as a novel prognostic biomarker, might be an oncogenic lncRNA which promoted proliferation, migration, and invasion by modulating Wnt/β-catenin signaling. Our results enlarged our knowledge in the molecular pathology of CC tumorigenesis.