INTRODUCTION
Glioma, a common primary neoplasm, is a cancerous tumor that develops from neural stromal cells. It comprises around 80% of all tumors affecting the central nervous system and represents 40-50% of malignancies found in the brain [20,29]. Gliomas are diagnosed annually around 300000 times worldwide and account for approximately 2.5% of cancer-related deaths [26]. Currently, glioma is primarily categorized into diffuse glioma and non-diffuse glioma, with the majority of gliomas falling under the diffuse glioma classification [13]. The classification of tumors in the central nervous system by the World Health Organization (WHO) includes gliomas, which can be categorized as primary or secondary forms. These gliomas are further classified based on WHO grades, which range from 1 to 4. These include astrocytomas, oligodendrogliomas, glioblastoma (GBM), and various other subtypes [15]. Glioma, is currently managed using different methods including surgical removal, radiation therapy, chemotherapy, biological treatment, and tumor-treating fields. While significant advancements have been made in immunotherapy for tumor treatment, its application in glioma treatment has not yielded significant breakthroughs. Due to the aggressive characteristics and high frequency of occurrence linked with glioma, the present survival rate for glioma patients after 5 years remains extremely poor, below 5% [7]. The possible explanation for this phenomenon is due to the absence of genes related to immunity and reduced immune cell presence in the glioma microenvironment [8,17]. Hence, the presence of efficient biomarkers becomes imperative in order to achieve precise diagnosis and prognostic evaluation, alongside identifying potential therapeutic targets.
Survivin, a regulatory protein known as Baculovirus inhibitory of apoptosis repeat-containing 5 (BIRC5), is a crucial member of the inhibitor of apoptosis (IAP) family. Its principal function is to impede caspase activation, ultimately obstructing apoptosis, commonly referred to as programmed cell death [23]. Survivin is a compact protein made up of 142 amino acids, with a molecular weight of around 16 kilodaltons. This multifunctional protein belongs to a class of apoptosis-inhibitory proteins that were identified in 1997 [1]. It is an eukaryotic protein, which is evolutionarily conserved, exhibits significant expression levels in both embryonic tissues and tumors. BIRC5 expression is closely linked to several important biological processes crucial for tumor development and advancement [14]. BIRC5 is typically found at low levels in healthy tissues, but is significantly increased in cancerous tumors [30,32]. Recent studies have demonstrated that suppressing BIRC5 promotes apoptosis and leads to cell cycle arrest, effectively hindering the proliferation of laryngeal squamous cell carcinoma cells [28]. Nonetheless, limited research exists on BIRC5’s role in various illnesses, and the precise mechanism by which BIRC5 contributes to the initiation and advancement of glioma remains unknown.
This study analyzed the role of BIRC5 gene in glioma through The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) database. Through a comprehensive array of bioinformatics analyses, we clarified the expression and critical importance of the BIRC5 gene in the context of glioma. The present investigation presents a theoretical groundwork for identifying tumor biomarkers and prospective targets for therapy, thus furnishing valuable perceptions. Based on our investigation, it is evident that the prognostic prediction of glioma patients can be facilitated through the utilization of BIRC5 as a valuable biomarker.
MATERIALS AND METHODS
This study was approved by the Ethical Committee of The First Affiliated Hospital of Dali University (ethics number : DFY20210123001).
Data collection
The messenger RNA (mRNA) expression and clinical information data for glioma (GMB and LGG) were collected from the TCGA database (https://portal.gdc.cancer.gov/). Additionally, gene expression profiles for GSE4290 were retrieved from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), which included 157 glioma samples and 23 non-tumor samples.
Logistic regression and Cox analysis
In the study of glioma, the association between clinical features and this type of tumor was examined using Wilcoxon signed-rank test and logistic regression analysis. The prognosis of glioma was determined by conducting Univariate Cox regression analysis to identify factors impacting it. Additionally, a multivariate Cox regression analysis was performed to evaluate the influence of various clinical characteristics on the prognosis of glioma.
Survival analysis
Data from the TCGA database was analyzed to evaluate the survival rates of glioma patients with different clinical characteristics. Kaplan-Meier diagrams were created to investigate the correlation between patient survival and the expression of BIRC5. To accomplish this, a log-rank test was conducted using the R survival package. Hazard ratios (HRs) and log-rank p-values with 95% confidence intervals were calculated.
Construction and evaluation of glioma survival prediction nomogram
Using the R package, we plotted the receiver operating characteristic curve (ROC) curve and time-dependent curve of diagnosis, which included pROC and time-ROC. Independent clinicopathological prognostic factors were identified through a Cox regression analysis. Following this, a nomogram was created with the assistance of the R package. This nomogram effectively assesses the likelihood of overall survival (OS) for glioma patients at specific intervals, including 1, 5, and 10 years.
Analysis of functional enrichment
To understand the potential biological processes and pathways linked with BIRC5, we conducted the following analysis. We obtained the top 801 genes linked to BIRC5 in glioma, as well as the top 978 genes associated with glioma survival, from the TCGA database. Enrichment analysis of these genes was performed using Gene Ontology (GO) via the Database for Annotation, Visualization, and Integrated Discovery (DAVID), combined with pathway analysis from the Kyoto Encyclopedia of Genes and Genomes (KEGG), and employing the R package ggplot2. A statistical significance level of p<0.05 was considered for determining corrected results.
Single-sample gene set enrichment analysis (GSEA) immune infiltration analysis
The immune cell infiltration analysis on glioma samples was conducted using the single-sample GSEA method with the GSVA package in R, which examined 24 diverse types of immune cells. To assess the link between glioma and individual immune cell subpopulations, Spearman’s correlation coefficient analysis was performed.
LinkedOmics and TISIDB database analysis
Explore the expression profile of BIRC5 through the LinkedOmics database (http://www.linkedomics.org). We used GSEA in the Link Interpreter module to explore the GO and KEGG pathways of BIRC5 and its co-expressed genes. TISIDB, a vital web platform for identifying connections between tumors and the immune system, serves as a comprehensive repository of knowledge (http://cis.hku.hk/TISIDB/). Our research centered on investigating the correlation between the BIRC5 methylation levels and the chemokine expression, along with chemokine receptor levels, by utilizing the chemokine module.
BIRC5 interactions with chemicals and genes
The study of novel connections in the molecular pathways through which chemicals impact health outcomes is made possible by the Comparative Toxicogenomics Database (CTD; http://ctdbase.org/). We have utilized this valuable resource to investigate the interacting chemicals of BIRC5.
GSEA analysis of BIRC5 genes
The perl language was used to obtain the expression data set, file containing phenotype data and target gene. The GSEA4.0.3 software (Broad Institute, Cambridge, MA, USA) was utilized to import and examine these files, establishing a connection between the target gene and GSEA. The GSEA software, which operates on the JAVA platform.
Tissue specimen collection
In this study, 11 patients with glioma and seven normal brain tissue samples were collected in The First Affiliated Hospital of Dali University. Inclusion criteria : age over 18 years; complete clinical data; postoperative pathology confirmed glioma; haven’t receive radiotherapy or chemotherapy before operation. Exclusion criteria : combined with other tumors; complicated with acute and chronic infection; complicated with serious heart, liver, kidney and other organ diseases; combined with endocrine diseases. Prior to surgery, no additional therapy was given to patients. All tissue samples were preserved in paraffin for subsequent Hematoxylin and Eosin (H&E) staining and immunohistochemical analysis. The committee of The First Affiliated Hospital of Dali University provided approval for the experiments. In addition, informed consent for obtaining tissue samples was secured from all participants involved in the study. Furthermore, all methods utilized in the research were carried out in strict compliance with the applicable guidelines and regulations.
H&E and immunohistochemical staining
Paraffin-embedded tissue samples are sliced on a microtome, and the sections are adhered to slides using warm water before being dried at a constant temperature. Dewaxing is performed with xylene, followed by staining with hematoxylin solution. After washing with ethanol for decolorization, staining with eosin is carried out. The sections are then dehydrated using anhydrous ethanol, made transparent with xylene, and finally mounted with resin before sealing the slides.
A slicer was utilized to slice the paraffin blocks into thin 4 μm sections. These sections were then dewaxed using xylene and rehydrated with a series of graded ethanol solutions. To inhibit endogenous peroxidase activity, the slices were treated with a 3% hydrogen peroxide solution for 20 minutes. Antigen repair was achieved using citrate buffer, followed by the addition of Rabbit anti-human polyclonal antibodies BIRC5 which were allowed to incubate overnight at 4°C. On the subsequent day, the slices were incubated with sheep anti-rabbit secondary antibodies at room temperature for 1 hour. The expression of BIRC5 in glioma and normal brain tissue was examined following staining with diaminobenzidine. The tissues were then subjected to a series of processes, including dehydration, making them transparent, and finally fixing them. Observations were made under a standard light microscope to assess the levels and patterns of BIRC5 expression in the different tissue types. BIRC5 positive evaluation criteria in the table : no coloring is scored as 0 points, light yellow as 1 point, brown as 2 points, tan staining as 3 points; proportion of positive cells scored : <25% as 1 point, 25-50% as 2 points, >50% score 3 points. Multiply the staining intensity score and the proportion of positive cells : 0 to 1 is negative (-), 2 to 4 is positive (+), 5 to 8 is moderately positive (++), and 9 to 12 is strong positive (+++), 2-12 points are considered positive. All immunohistochemistry results were interpreted by two senior pathologists using a double-blind method.
Cell culture
We received the human glioma U251MG cell line as well as the normal astrocytes human astrocytes (HA) cells from Procell Life Science Technology. The two cell lines were cultured in Minimum Essential Medium that was supplemented with 10% fetal bovine serum and 1% cyanin-streptomycin, providing the necessary nutrients and antibiotics to support cell growth. This enriched culture medium was replaced every 48 hours to maintain optimal conditions for cell proliferation. Additionally, cell passage was conducted every 3 to 4 days to prevent over-confluence and ensure continued healthy growth and maintenance of the cell cultures.
Quantitative real‐time polymerase chain reaction (qRT-PCR) and western blot
The levels of BIRC5 mRNA expression were measured in U251MG and HA cells. RNA was extracted from the cells in culture, and approximately 1 μg of total RNA was reverse transcribed into complementary DNA using the Vazyme reverse transcription kit. The Quantitative SYBR Green PCR Kit from Vazyme (CITY, STATE, COUNTRY) was employed to perform qRT-PCR using the SLAN-96S Detection System. The qRT-PCR procedure adhered to a specific series of conditions. Initially, the process began with a step at 95°C for 30 seconds. This was followed by a second step consisting of 40 cycles, where each cycle included holding at 95°C for 10 seconds and then 60°C for 30 seconds. Finally, the procedure concluded with a third step, which involved heating at 95°C for 15 seconds, maintaining at 60°C for 60 seconds, and then heating again at 95°C for 15 seconds. The PCR primers employed in the study were as follows : BIRC5 forward, 5’-CAAGGACCACCGCATC TCTA-3’ and reverse, 5’-TTGGTTTCCTTTGCATGGGGT-3’; glyceraldehyde-3-phosphate dehydrogenase (GAPDH) forward, 5’-CATGT TGCAACCGGGAAGGA-3’ and reverse, 5’-GCCCAATACGACCAAATCAGAGA-3’. mRNA expression levels were calculated and normalized utilizing the 2-ΔΔCt method with reference to GAPDH. Each experiment was carried out three times.
Proteins from the cells were extracted using Radio Immunoprecipitation Assay lysate and analyzed by the BCA method. Following this, the protein samples underwent separation via sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel electrophoresis and were then transferred onto a polyvinylidene fluoride membrane. To start the experiment, a skim milk solution with a concentration of 5% was enclosed and left to incubate at 4°C alongside primary antibodies BIRC5 and GAPDH for a full night. The subsequent morning, the original antibody was washed away, and a secondary antibody was added for a span of 2 hours. Chemiluminescence detection was carried out using the ECL Immobilon Western Kit (KALANG, Shanghai, China), with gray quantization analysis performed using Image J software (National Institutes of Health, Bethesda, MD, USA).
Statistical analysis
The data were analyzed using SPSS ver. 19.0 software (IBM, Armonk, NY, USA) through statistical analysis, presenting the results as mean±standard deviation. To evaluate variances between groups, the t-test was utilized. Additionally, the chi-square test was employed to identify distinctions in protein expression across various groups. R software (version 3.6.3; R Rstudio, Boston, MA, USA) was employed for performing statistical analyses. Survival analysis was conducted using the Kaplan-Meier method and Cox regression analysis. Furthermore, to compare curve differences among groups, the Log-Rank test was utilized. A significance level of p≤0.05 was applied to determine statistical significance.
RESULTS
Clinical characteristics of glioma patients.
Table 1 presents the clinical characteristics of 699 glioma patients sourced from the TCGA repository. As illustrated in the Table 1, there are 556 patients aged 60 or younger, 143 patients over the age of 60, 298 women and 401 men. According to WHO classification, there were 224 cases at G2 stage, 245 cases at G3 stage, and 168 cases at G4 stage. For more specific details, refer to Table 1.
BIRC5 gene expression in glioma and clinical sample validation
Utilizing the R package, the data retrieved from the TCGA database were scrutinized to investigate the variations in BIRC5 expression between tumor and normal tissues. The results show that the BIRC5 gene exhibits high expression levels in a variety of tumors. In particular, elevated expression is detected in BLCA, BRCA, CESC, CHOL, COAD, ESCA, and GMB, detailed results are shown in Fig. 1A. The mRNA expression levels of the BIRC5 gene were analyzed in glioma tumor tissues and adjacent normal tissues using data from the TCGA database. Results indicated a significant elevation in BIRC5 expression in glioma tumor specimens compared to the surrounding normal tissues, these outcomes are illustrated in Fig. 1B. When analyzing BIRC5 expression levels based on the WHO classification, the findings indicated that expression at the G3 stage exceeded that at the G2 stage, and the G4 stage exhibited even higher levels compared to both the G2 and G3 stages, these differences were statistically significant, as illustrated in Fig. 1C. When classified according to the 1p/19q codeletion status, the study indicated that there was a notable increase in the expression of BIRC5 in the non-codeletion group as opposed to the codeletion group. Additionally, it was noted that the BIRC5 expression levels were substantially greater in the deceased cohort (deceased group) compared to the surviving cohort (alive group), these findings are illustrated in Fig. 1D and E.
In order to ascertain the levels of expression for BIRC5 in glioma, we conducted an analysis of both the transcription levels of BIRC5 within tumor tissues and in the surrounding non-tumorous tissues. This analysis was performed using clinical data from the GSE4290 dataset, which is available in the GEO database. According to the results, it was noted that there was a significant increase in the expression of BIRC5 in gliomas when compared to the surrounding para-carcinoma tissues in unpaired and paired samples. This discrepancy was further confirmed through a comparison of BIRC5 transcription levels in glioma tissues versus para-carcinoma tissues, the results provide strong evidence supporting the notion that BIRC5 expression is indeed heightened in gliomas, as depicted in Fig. 1F and G.
High levels of BIRC5 serve as an autonomous predictor for the OS of glioma
We used logistic regression analysis to assess how different clinical factors relate to the prognosis of glioma patients. Our findings, as presented in Table 2, indicate a significant association between OS and factors such as grade, isocitrate dehydrogenase (IDH) status, 1p/19q codeletion, primary therapy outcome, and histological type. To clarify the relationship between BIRC5 expression and clinical characteristics in glioma patients, we utilized the TCGA database to assess BIRC5 mRNA expression levels across various clinical categories. In addition, both Univariate and Multivariate COX analyses were employed to assess the relationship between the BIRC5 gene expression and the clinical traits of glioma patients. The significant connection between increased BIRC5 levels and poor OS among glioma patients was unveiled through Univariate Cox analysis, as illustrated in Table 3. To determine the relationship between BIRC5 gene expression and the prognosis of glioma patients, we divided the glioma patients in the TCGA database into two distinct groups. By doing so, we aimed to better understand how variations in BIRC5 expression might be linked to patient outcomes. The first group consisted of the top 50% of samples displaying high expression levels, while the second group comprised the remaining 50% of samples, showing comparatively lower expression levels. The categorization of subjects was determined by the median value of BIRC5 expression. Examination of the survival outcomes from Kaplan-Meier reveals a connection between the levels of BIRC5 expression and the survival rates for both overall and disease-specific cases. Notably, higher levels of BIRC5 expression correlate with a less favorable survival prognosis, detailed results supporting these findings are presented in Fig. 2A and B. The results of the study on subgroup analysis show that increased levels of BIRC5 are associated with negative outcomes in glioma patients in the following situations : WHO grade G2, G3, G4, HR was 4.95 (p<0.001); G3 and G4 stage, HR was 3.39 (p<0.001); histological type astrocytoma and oligoastrocytoma, HR was 3.04 (p<0.001); oligodendroglioma and GBM, HR was 5.36 (p<0.001); 1p/19q codeletion non-codel and codel, HR was 4.53 (p<0.001); primary therapy outcome progressive disease and stable disease, HR was 2.57 (p<0.001); female and male, HR was 4.58 (p<0.001), detailed data are shown in Fig. 2C-I.
Diagnostic value of BIRC5 gene expression in glioma
The significance of diagnosing glioma using the BIRC5 gene will be evaluated by creating ROC curves and performing nomogram analysis with the TCGA database’s BIRC5 gene expression data. The ROC curve presented in Fig. 3A revealed an area under the curve (AUC) of 0.938, suggesting a considerable diagnostic utility. By utilizing the time-dependent survival ROC curve for BIRC5, projections were generated for the survival probabilities of individuals with glioma at 1-year, 5-year, and 10-year time points. For the 1-year prediction, the AUC was 0.781; for the 5-year prediction, it was 0.782; and for the 10-year prediction, the AUC was found to be 0.665. These AUC values indicate that our predictions are deemed appropriate for forecasting the survival of these patients (Fig. 3B). The prediction of BIRC5 expression level using the nomogram demonstrated superior prognostic potential in comparison to IDH status and 1p/19q codeletion (Fig. 3C). Calibration plots of the model show agreement between prognostic predictions and actual observations (Fig. 3D).
Analysis of functional enrichment and genes associated with BIRC5 in glioma
Utilizing the LinkFinder module on the LinkedOmics website, we analyzed the co-expression pattern of BIRC5 in glioma using data from the TCGA database. Fig. 4A illustrates this analysis, where the red color signifies the first 15 genes demonstrating positive correlation with BIRC5, while the green color represents the last 15 genes exhibiting negative correlation with BIRC5. The GO functional enrichment and KEGG pathway of the BIRC5 gene were analyzed through DAVID functional annotation bioinformatics microarray, and the organelle fission, chromosomal region, catalytic activity acting on DNA, cell cycle pathways were mainly enriched, results are shown in Fig. 4B. To detect genes demonstrating identical regulatory tendencies both in patients with high BIRC5 expression and non-survival cases, we intersected 801 BIRC5-related genes with 978 survival-related genes, identified a total of 72 genes which share a correlation with both BIRC5 and glioma survival, as depicted in Fig. 4C. In glioma patients, 72 protein-coding genes may serving as potential genetic biomarkers. Enrichment analysis of GO functional and KEGG pathway was executed on this set of 72 common genes. The results showed a significant increase in differentially expressed genes in important biological processes such as nuclear division, spindle, tubulin binding, as well as the cell cycle pathway, as depicted in Fig. 4D. Gene co-expression correlation analysis in Fig. 4E shows that there are significant positive correlations between most proteins in the network.
Correlation of BIRC5 expression with immune characteristics
Utilizing the TIMER database, we conducted an analysis of immune cell infiltration within gliomas, aiming to investigate the correlation between BIRC5 expression levels and the tumor’s immune response. The result indicated a positive association between the levels of BIRC5 expression and the presence of Th2 cells, Macrophages, aDc, along with neutrophils. On the other hand, a negative association was found between the expression levels of BIRC5 and both T follicular helper cells (TFH), central memory T cells, plasmacytoid dendritic cell, natural killer CD56bright cells, Tem (Fig. 5A). Further investigation into the correlation between the extent of BIRC5 expression and immune system invasion in glioma revealed a direct link between BIRC5 expression and Th2 lymphocytes (r=0.878, p<0.001), while revealing a negative correlation with TFH (r=-0.357, p<0.001), in Fig. 5B and C. Furthermore, TISIDB was employed to examine the connection between methylation levels of BIRC5 and chemokines as well as chemokine receptors in gliomas. The results shown in the heat map indicated a significant correlation between the methylation status of BIRC5 in glioma and different chemokines as well as chemokine receptors (Fig. 5D and E). The findings suggest that the methylation of BIRC5 could potentially impact tumor immunity. In a follow-up analysis using the TISIDB database, the relationship between BIRC5 methylation and the levels of immunoinhibitors and immunostimulators in various human tumor types was examined (Fig. 5F and G). These findings suggest that BIRC5 in gliomas may have a role in modulating tumor immunity.
Interacting chemicals of BIRC5
The data from the CTD database listed that BIRC5 is associated with many chemicals, we selected the top 10 chemicals, five of which were up-regulated and five were down-regulated, in Table 4.
GSEA enrichment analysis of BIRC5 gene
To gain a deeper understanding of the biological progression and role of BIRC5 in glioma, patients diagnosed with glioma were categorized into two groups based on the degree of BIRC5 expression : a high expression group and a low expression group. In this research, relevant data on BIRC5 expression were obtained through the utilization of perl language, resulting in the acquisition of the BIRC5 expression dataset as well as the phenotypic dataset. These datasets were then imported into the GSEA4.0.3 software (Broad Institute). The analysis results uncover a considerable association between the abundant expression of BIRC5 mRNA and diverse cellular processes, including the P53 signaling pathway, DNA replication, cell cycle, phosphoinositol signaling system, and neuroactive ligand receptor interaction signaling pathway.
Cell verification and section staining
The levels of mRNA for the BIRC5 genes were confirmed in both U251 and HA cells, as depicted in Fig. 6A. The comparison between HA cells and U251 cells revealed a significant increase in the mRNA levels of BIRC5 in the latter, suggesting that BIRC5 might serve as a potential hazard for glioma development. Analysis of BIRC5 protein expression levels in HA and U251 cells demonstrated a notable rise in BIRC5 levels in U251 cells in comparison to the HA group, as depicted in Fig. 6B. After processing the gathered samples and applying staining, results from H&E staining (Fig. 6C) revealed a slight abnormality in the overall structure of brain tissue. Despite the orderly arrangement of tissue neurons, a significant presence of loosely packed and swollen neurons was observed within the tissue. No signs of apparent neuronal pyknosis or necrosis are present in the tissue, and there is an absence of inflammatory cell infiltration in the observed tissue. In glioma tissue, a large number of abnormal hyperplasia of glioma can be seen in the tissue, forming glioma. Glioma tumor cells are single, numerous and closely arranged, and the atypia is not obvious, all were glioma cells, and obvious hyperemia and expansion of blood vessels can be seen in the tissue. The immunohistochemical analysis of BIRC5 protein expression in normal brain tissue and glioma is illustrated in Fig. 6C. The immunohistochemical staining of tumor tissue was all positive, and the immunohistochemical staining of normal brain tissue was negative in seven cases (p<0.001), the difference was statistically significant. This analysis demonstrates a notable rise in BIRC5 levels in glioma when contrasted with normal brain tissue.
DISCUSSION
In recent times, there has been a noticeable rise in the number of glioma cases, characterized by a low success rate in treatment, high frequency of occurrence, and frequent reoccurrence [10]. The exploration of immunotherapy as a treatment for glioma has been extensively conducted in clinical settings; however, there remains a dearth of dependable molecular markers to evaluate the efficacy of immunotherapy [11]. Consequently, it is crucial to identify prognostic biomarkers associated with immune response and to screen for target genes in the context of tumor immunotherapy. In the current investigation, the analysis employing various methods suggests that BIRC5 can be regarded as a valuable biomarker and an independent gene associated with a risk of developing glioma. As such, it holds potential as a target for immunotherapy.
The present study uncovered the BIRC5 gene as a promising candidate gene for the treatment of glioma. BIRC5, a component of the IAP protein family, exhibits inhibitory effects on programmed cell death, also called apoptosis. It is crucial in controlling cell division and inhibiting the functions of caspase-3 and caspase-7, which in turn hinders the apoptosis of cancer cells and boosts cell growth [4,16]. BIRC5 exhibits elevated levels of expression in different tumor tissues, while its presence is significantly reduced or even absent in normal tissues. Multiple studies have substantiated its involvement in tumor cell malignant transformation and anti-apoptotic physiological processes, further signifying the BIRC5 gene as a crucial target for anti-tumor therapies [24].
In gliomas, the expression levels of BIRC5 in G2, G3, and G4 grades are significantly different according to WHO classification. This investigation presents notable findings concerning the elevated levels of BIRC5 expression in individuals diagnosed with glioma compared to the normal tissue counterpart. The study reveals a significant link between the prognosis of patients and the expression of BIRC5. Observations indicated that elevated BIRC5 expression was predominantly detected in the deceased group as opposed to the surviving subjects. This outcome is consistent with previous research outcomes, therefore reinforcing the validity of these findings.
Using the median expression level of BIRC5, the subjects were divided into two categories : high-expression and low-expression groups. The high expression of BIRC5 is associated with OS according to WHO classification, histological type, 1p/19q codeletion status, outcomes of primary therapy, and gender categories. Furthermore, independent risk factors for glioma can be identified through logistic univariate COX analysis results, indicating that the BIRC5 gene exhibits potential significance. The ROC curve shows that the BIRC5 gene has a high diagnostic value for glioma. The ROC curve, which varies with time, is deemed appropriate for forecasting, while the model’s calibration graph indicates that the prediction of prognosis aligns with the observed outcomes.
In the development and advancement of tumors, inflammation reactions linked to cancer play a crucial role. Within the tumor microenvironment, alterations in the quantity of immune cells involved in infiltrating inflammation may induce oxidative harm and aberrant DNA repair, thereby leading to genetic or epigenetic modifications. Ultimately, these modifications facilitate the promotion of glioma growth, invasion, and metastasis [6,25]. Through an in-depth examination of the relationship between the methylation levels of BIRC5 and the expression of chemokines and chemokine receptors in immune cells, our study uncovered significant insights into the role of BIRC5 in tumor immunity. The research also explored how methylation levels are influenced by the presence of immunosuppressants and immune stimulators, further highlighting BIRC5’s regulatory function in the immune response to tumors. Checkpoint inhibitors for the immune system, a promising strategy in cancer treatment, have made significant progress in improving the outlook for individuals diagnosed with various forms of cancer. The CTD database analyzed chemicals that interact with BIRC5 and found that a variety of chemicals are related to BIRC5. The expression of some chemicals was up-regulated and the expression of some was down-regulated. The experimental validation through qRT-PCR revealed that BIRC5 expression levels were notably elevated in the U251 glioma cell line in comparison to normal astrocytes (HA cells). Additionally, analysis through immunohistochemistry and western blot techniques clearly demonstrated a notable upregulation of BIRC5 in glioma tissues as opposed to normal brain tissues. These findings strongly support the conclusions of earlier studies.
IAP, a highly conserved factor that prevents cell death, primarily functions by repressing caspase function and regulating NF-κB. Apoptosis plays a crucial role in sustaining cellular homeostasis. Typically, the body triggers the apoptosis process to remove faulty DNA or cells experiencing abnormal cell division. However, when cells undergo malignant transformation and the poptosis machinery becomes impaired, the body loses its ability to actively eliminate malignant cells, leading to the development of tumors [2]. BIRC5 plays a role in the stimulation of new blood vessel growth, making it a useful biomarker for the detection, therapy, and prediction of tumors [1].
In an effort to shed light on the molecular mechanisms driving glioma onset and development, this study utilized GSEA to examine publicly accessible datasets. The objective was to identify the crucial pathways associated with the BIRC5 gene. The obtained results demonstrated that heightened expression of BIRC5 gene is capable of impeding the P53 signaling pathway, it is noteworthy that the P53 gene mutation is the prevailing genetic alteration observed in glioma [21], This mutation works by regulating cell growth, preserving DNA integrity, and hindering the development of cells with cancerous tendencies. Accordingly, the P53 gene represents the most extensively studied tumor-associated gene. Multiple research investigations propose that the emergence and advancement of diverse tumor forms often align with P53 genetic mutation [18]. The primary function of P53, a tumor suppressor gene, lies in its ability to mend DNA damage. Moreover, P53 triggers programmed cell death in instances of severe and irreparable DNA damage. The wild type P53 has a negative regulatory effect on BIRC5, inhibiting its expression at both the mRNA and protein levels. When a mutation occurs in P53, it no longer has the capability to interact with the BIRC5 promoter, leading to the abnormal BIRC5 expression. This altered expression contributes to tumor growth and progression. The association between BIRC5 and glioma may be closely associated with the p53 pathway.
However, further investigation is warranted to decipher the precise mechanism operative in this association. Past studies have shown that p53 suppresses the transcription of BIRC5 to promote cell apoptosis that is dependent on p53 [9,19,33]. Significantly, the use of YM155, an inhibitor of BIRC5, resulted in a decrease in BIRC5 levels and a rise in p53-up-regulated modulator of apoptosis expression, a key target of p53, ultimately resulting in cell death [31]. In addition, BIRC5 promotes cell division by creating the chromosomal passenger complex with CDCA8 and INCENP, affecting the movement of microtubules in the G2/M phase [22].
Elevated BIRC5 levels detected in gliomas are associated with indicators of malignancy in tumor histology, and serve as a prognostic factor indicating unfavorable patient outcomes [27]. According to study heightened BIRC5 levels in U251 cells result in cellular deterioration, exacerbated DNA damage, and structural chromosomal abnormalities. Moreover, they facilitate cell proliferation while inhibiting apoptosis [5], These findings are consistent with prior research. Various studies have documented abnormal upregulation of BIRC5 expression in diverse tumor tissues, emphasizing its crucial role in malignant tumor progression [12]. IIn connection with lung adenocarcinoma, increased BIRC5 expression has been linked with a heightened risk of distant metastasis and tumor burden [3]. In conclusion, the aforementioned studies collectively demonstrate the association between BIRC5 and diverse tumors. This study opens up new possibilities for glioma treatment by uncovering BIRC5 as a promising gene target in gliomas.
Glioma patients display a poorer prognosis as a result of the significant increase in BIRC5 levels. Therefore, BIRC5 expression acts as a predictive factor in glioma patients. The role of BIRC5 in the onset and advancement of glioma is through the P53 signaling route. Going forward, focusing on suppressing BIRC5 expression may offer a new strategy for glioma treatment.
The current study has limitations due to insufficient clinical samples available for validating the prognostic impact of BIRC5 as well as its potential association with the immune system. Further investigation is needed to explore the connection between the BIRC5 gene and glioma, thus underscoring the necessity for additional research to elucidate the mechanism and importance of BIRC5 expression in this particular form of brain neoplasm.
CONCLUSION
This investigation has uncovered a correlation between the survival duration of individuals diagnosed with glioma who undergo immunotherapy and the presence of BIRC5. The length of the survival period for glioma patients is inversely proportional to the level of BIRC5 expression. BIRC5 potentially assumes a critical function in glioma development via the P53 signaling pathway. Evaluating the expression level of BIRC5 can be utilized as a prognostic indicator to assess glioma patients. Through investigation, this study boosts our comprehension of the participation of BIRC5 in glioma, providing fresh insights into targeted treatments for glioma.
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