An Immunogenic Cell Death-Related Gene Signature Predicts the Prognosis and Immune Infiltration of Cervical Cancer

Background: Inhibin A (INHBA), a member of the TGF-β superfamily, has been shown to be differentially expressed in various cancer types and is associated with prognosis. However, its role in cervical cancer remains unclear. Methods: We aimed to demonstrate the relationship between INHBA expression and pan-cancer using The Cancer Genome Atlas (TCGA) database. Next, we validated INHBA expression in cervical cancer using the Gene Expression Omnibus (GEO) database, including GSE7803, GSE63514, and GSE9750 datasets. Enrichment analysis of INHBA was performed using the R package “clusterProfiler.” We analyzed the association between immune infiltration level and INHBA expression in cervical cancer using the single-sample gene set enrichment analysis (ssGSEA) method by the R package GSVA. We explored the association between INHBA expression and prognosis using the R package “survival”. Results: Pan-cancer data analysis showed that INHBA expression was elevated in 19 tumor types, including cervical cancer. We further confirmed that INHBA expression was higher in cervical cancer samples from GEO database and cervical cancer cell lines than in normal cervical cells. Survival prognosis analysis indicated that higher INHBA expression was significantly associated with reduced Overall Survival (p = 0.001), disease Specific Survival (p = 0.006), and Progression Free Interval (p = 0.001) in cervical cancer and poorer prognosis in other tumors. GSEA and infiltration analysis showed that INHBA expression was significantly associated with tumor progression and some types of immune infiltrating cells. Conclusion: INHBA was highly expressed in cervical cancer and was significantly associated with poor prognosis. Meanwhile, it was correlated with immune cell infiltration and could be used as a promising prognostic target for cervical cancer.

Up-regulated BCAR4 contributes to proliferation and migration of cervical cancer cells

Cervical cancer (CC) is a prevalent gynecological malignancy with notable heterogeneity. The role of mitochondrial permeability transition (MPT)-driven necrosis, a form of cell death due to mitochondrial dysfunction, in CC progression and prognosis is poorly understood and represents a promising therapeutic target for cancers. This study aimed to create a new prognostic signature linked to MPT-driven necrosis, improving CC prediction and prognosis. This study utilized the GSE63514, TCGA-CESC, CGCI-HTMCP-CC, and GSE197641 transcriptome datasets. Firstly, the GSE63514 dataset was utilized to identify differentially expressed genes (DEGs). Differentially expressed MPT-driven necrosis-related genes (DE-MRGs) were obtained by intersecting DEGs with MRGs. Regression analyses were performed to identify genes significantly associated with prognosis. A prognostic model was established in TCGA-CESC, followed by independent validation and nomogram construction. Additional analyses included immune infiltration, enrichment analysis, and drug susceptibility based on high- and low-risk groups. Finally, cell communication analysis was performed to investigate interactions between key cell types. A total of 156 DE-MRGs were identified. Regression analyses identified three prognostic genes (ICOS, MMP3, and POSTN) to construct a prognostic risk signature. Then, risk score was an independent prognostic factor for CC, and a nomogram demonstrated effective predictive accuracy for CC survival outcomes. The risk signature was linked to immune-associated processes such "Antigen processing and presentation" and immune cell infiltration, especially M0 macrophages and CD8 T cells. Cell communication analysis uncovered a strong interaction between endothelial cells and monocytes. To validate the molecular mechanisms, qRT-PCR, cell proliferation, and wound healing assays were performed. Functional tests showed that MMP3 and POSTN knockdown drastically reduced CC cell growth and migration. This study developed a novel prognostic risk signature based on ICOS, MMP3, and POSTN. MMP3 and POSTN knockdown significantly decrease CC cell growth and migration, highlighting their potential as therapeutic targets.

<b>Background: </b>This study was designed to investigate the roles of RASAL2 in cervical cancer (CC). <b>Methods:</b> Fifty-four CC tissues and 33 adjacent tissues were obtained from CC patients admitted to our hospital between March 2012 and June 2014. Real-time polymerase chain reaction and western blotting were performed to analyze the expression of RASAL2 mRNA and protein in these tissues, CC cell lines, and normal cervical cells. Over-expression and silencing of RASAL2 were induced after transfection, and the migration, invasion, and proliferation of the CC cell lines were examined. <b>Results:</b> RASAL2 mRNA and protein expressions were significantly down-regulated in CC tissues and cell lines than in adjacent tissues and normal cervical cells, respectively. While low RASAL2 expression correlated with advanced stage and metastasis of CC, its over-expression significantly inhibited proliferation and metastasis of CC cells and induced apoptosis. Under <i>in vitro</i> conditions, silencing of RASAL2 expression could significantly increase the proliferation, invasion, and migration of CC cells. <b>Conclusion:</b> RASAL2 functioned as a tumor suppressor in CC, and was down-regulated in CC tissue samples and cell lines. tumor suppressor in CC, and was down-regulated in CC tissue samples and cell lines.

Homeobox C8 (HOXC8) is a transcription factor that has been reported to regulate numerous genes associated with tumor progression. However, its function in cervical cancer (CC) remains to be elucidated. In the present study, the expression level of HOXC8 was examined in CC tissues and cell lines using reverse transcription-quantitative polymerase chain reaction and western blot analysis. Additionally, CC cell lines were transfected with small interfering RNAs (siRNAs) to downregulate the expression of HOX8 and assess cell proliferation using Cell Counting Kit-8. The results demonstrated a significantly increased expression of HOXC8 in CC tissues and cell lines compared with non-tumor tissues, and a normal cervical cell line, respectively. Additionally, the downregulation of HOXC8, which was achieved by siRNA transfection, significantly inhibited the proliferation rate of CC cell lines. Kaplan-Meier curves demonstrated that the increased expression of HOXC8 was associated with poor prognosis of patients with CC. Additionally, univariate and multivariate analysis revealed that HOXC8 was a significant and independent predictor for overall survival of patients with CC. In conclusion, the results of the present study suggest that HOXC8 may be involved in the progression of CC and may serve as a therapeutic target for 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.

Pan-cancer analysis of DCBLD1 and its association with the diagnosis, immunotherapy, and prognosis of cervical cancer.

Aberrant DNA damage repair (DDR) is one of the hallmarks of tumors, and therapeutic approaches targeting this feature are gaining increasing attention. This study aims to develop a signature of DDR-related genes to evaluate the prognosis of cervical cancer (CC). Differentially expressed genes were identified between high and low DDR groups of cells from the single-cell RNA sequencing dataset GSE168652 based on DDR scores. Using the ssGSEA and WGCNA methods, DDR-related differentially expressed genes were identified from different patients within the TCGA-CESC cohort. Using Cox analysis and LASSO regression analysis, a DDR-related gene signature was constructed based on the intersection of two groups of differentially expressed genes and DDR-related genes from WGCNA, and validated in GSE52903. Immune cell infiltration analysis, mutation analysis, survival analysis, drug sensitivity analysis, etc., were performed in different groups which were established based on the DDR gene signature scoring. A key gene affecting prognosis was selected and validated through biological experiments such as wound healing, migration, invasion, and comet assays. A novel DDR-related signature was constructed and the nomogram results showed this signature performed better in predicting prognosis than other clinical features for CC. The high DDR group exhibited poorer prognosis, weaker immune cell infiltration in the immune microenvironment, lower expression of immune checkpoint-related genes, lower gene mutation frequencies and more sensitivity to drugs such as BI.2536, Bleomycin and etc. ITGB1, ZC3H13, and TOMM20 were expressed at higher levels in CaSki and HeLa cells compared to ECT1 cells. Compared with the native CaSki and HeLa cells, the proliferation, migration, invasion and DDR capabilities of CaSki and HeLa cell lines with ITGB1 suppressed expression were significantly decreased. The 7 DDR-related gene signature was an independent and powerful prognostic biomarker that might effectively evaluate the prognosis of CC and provide supplementary information for a more personalized evaluation and precision therapy. ITGB1 was a potential candidate gene that may affect the DDR capacity of CC cells, and its mechanism of action was worth further in-depth study.

Background: Many patients with advanced cervical cancer (CC) have a poor prognosis and their mortality rank the first among women with malignant tumors. It's essential to explore the molecular mechanism of CC in clinical practice. Long noncoding RNA maternally expressed gene 3 (MEG3) has been reported to downregulate in CC tissues. However, the underlying mechanism of MEG3 in CC remains poorly elaborated. The current study aimed to explore the potential mechanism of MEG3 inducing endoplasmic reticulum stress (ERs)-mediated apoptosis of CC cells. Methods: The expression of MEG3 and miR-7-5p in CC tissues and cell lines was verified by quantitative reverse transcription/polymerase chain reaction (qRT-PCR). The vector of MEG3, miR-7-5p inhibitor, and sh-SCT1 were transfected into CC cell lines, and their expression was tested by qRT-PCR. Flow cytometry was used to detect apoptosis, and ERs-related protein expression was performed by Western blot. The regulatory relationship between MEG3/SCT1 and miR-7-5p was validated by Dual luciferase reporter assay. Results: CC tissues and cell lines showed downregulated MEG3 and STC1, and upregulated miR-7-5p. Overexpression of MEG3 or miR-7-5p inhibition induced ERs-triggered apoptosis of CC cells. In addition, sh-STC1 can reverse the effects of overexpressing MEG3 on CC cell apoptosis. In addition, dual luciferase reporter assay revealed that miR-7-5p can directly target to MEG3 and STC1. Conclusion: MEG3, act as a competing endogenous RNA of miR-7-5p, accelerates ERs-mediated apoptosis of CC cells through regulating SCT1 expression.

ObjectiveRecent studies indicated that transmembrane protein 40 (TMEM40) is associated with several types of cancers but is not clear in cervical cancer (CC). The study aimed to examine the role of TMEM40 in CC and related mechanisms.MethodsThe expression of TMEM40 in CC tissues and cell lines was studied with western blot and real-time quantitative RT-PCR. The effect of TMEM40 on proliferation was evaluated by CCK-8, EdU and colony formation assay. The migration, invasion, cell cycle and apoptosis of CC cells were studied with wound healing, transwell assays and flow cytometry. Tumor growth was evaluated in vivo using a xenogenous subcutaneously implant model.ResultsThe results revealed that the TMEM40 elevation in CC tissues and cell lines was closely correlated with tumor size and lymph node metastasis in clinical patients. Upregulation of TMEM40 with OE-TMEM40 vector promoted the invasion, migration and proliferation, inhibited the apoptosis and led to distinct S cell cycle arrest in CC cell lines. Silencing TMEM40 with shRNA inhibited the invasion, migration and proliferation, promoted apoptosis and led to a G0/G1 cell cycle arrest in CC cell lines. Silence of TMEM40 downregulated the expression of c-MYC, Cyclin D1, matrix metalloproteinase-1 (MMP-1) and matrix metalloproteinase-9 (MMP-9), but in contrast, activated p53 and several apoptosis related proteins such as p53, Caspase-3, Caspase-9 and PARP1. In addition, TMEM40 silencing dramatically decreased tumor growth in mice models.ConclusionThe present study demonstrates that TMEM40 upregulation can be a potential prognostic biomarker and contribute to CC development.

Several studies demonstrated that aberrant lncRNA expression contributes to cervical cancer (CC) development and progression. LINC00152, a novel lncRNA, has been identified as an oncogene involved in various cancers. In the present study, we aim to investigate the expression pattern, clinical significance, potential functional roles, and regulatory mechanism of LINC00152 in CC. The transcription levels of LINC00152, miR-216b-5p, and HOXA1 in CC tissues and cell lines were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). LINC00152 knockdown in CC cells was conducted by transfecting the LINC00152-specific siRNA. The cell proliferation ability was evaluated by the Cell Counting Kit-8 (CCK-8) assay. Cell cycle and apoptosis analysis were assessed by flow cytometry. The target relation among LINC00152, miR-216b-5p, and HOXA1 were measured using the dual-luciferase reporter assay. The protein levels of HOXA1 in CC cells were determined by Western blot. LINC00152 was up-regulated in CC tissues and cell lines. The high expression level of LINC00152 was positively correlated with poor prognosis and histologic grade in CC. The silence of LINC00152 could inhibit the proliferation of CC cells through inducing the cell cycle arrest at G0/G1 phase and promote apoptosis in vitro. Mechanically, we demonstrated that LINC00152 could modulate the proliferation of CC cells through elevating HOXA1 expression level via sponging miR-216b-5p based on bioinformatics analysis and experimental validation. Our findings revealed a novel molecular mechanism underlying LINC00152 modulating CC progression through the miR-216b-5p/HOXA1 pathway, suggesting that LINC00152 might potentially act as an effective diagnostic marker and therapeutic target for cervical cancer.

Increased expression of FHL2 promotes tumorigenesis in cervical cancer and is correlated with poor prognosis

Literature reports that lncRNA KCNQ1OT1 is markedly up-regulated in cervical cancer (CC) tissues and cell lines, and KCNQ1OT1 can promote the proliferation and metastasis of CC cells. This current work was designed to investigate the molecular mechanism underlying the participation of KCNQ1OT1 in CC progression. Herein, RT-qPCR was utilized for determining the levels of KCNQ1OT1, miR-296-5p and HYOU1 in clinical tumor tissue specimens and CC cell lines. Then, starBase predicted the complementary binding sites of KCNQ1OT1 and miR-296-5p or miR-296-5p and HYOU1. Dual-luciferase reporter assay/RIP assay validated the interplays among KCNQ1OT1/miR-296-5p/HYOU1. In addition, CCK-8, wound healing and transwell assays were employed to assess the proliferative, migrative and invasive properties of CC cells. Moreover, nude mice xenograft model was established by subcutaneously injection with SiHa cells in order to validate the precise functions of KCNQ1OT1/miR-296-5p/HYOU1 axis in CC in vivo. Besides, Immunohistochemical staining examined Ki-67 expression in xenograft tumors and western blotting analysis detected expressions of MMP2/9 and Wnt/β-catenin signaling pathway in CC cells and xenograft tumors. Elevated KCNQ1OT1 and HYOU1 as well as reduced miR-296-5p were observed in clinical tumor tissue specimens and CC cell lines. Results revealed that upregulation of miR-296-5p counteracted the enhancing effects of overexpressed KCNQ1OT1 on the proliferative, migrative and invasive abilities of CC cells. Additionally, HYOU1 overexpression abolished the suppressing effects of silenced KCNQ1OT1 on the malignant behaviors of CC cells and tumor growth. To conclude, KCNQ1OT1 could aggravate the malignant behaviors of CC and facilitate tumor growth through modulating miR-296-5p/HYOU1 axis.

The poor prognosis of cervical cancer patients leads to an annual increase in mortality, while microRNAs are involved in various cancers, including cervical cancer. This study aimed to investigate the clinical value and possible effect of miR-29b-2-5p on the progression of cervical cancer. The expression level of miR-29b-2-5p in cervical cancer tissues and cells was analyzed by polymerase chain reaction. The Kaplan-Meier curve was used to evaluate the role of miR-29b-2-5p in cervical cancer prognosis. The independent prognostic factors of cervical cancer were explored by the multivariate Cox regression analysis. The effect of miR-29b-2-5p on the proliferation, migration, and invasion of cervical cancer cells was determined by in vitro cell experiments. A significantly downregulated miR-29b-2-5p expression was observed in cervical cancer tumor tissues and cervical cancer cells compared with the adjacent tumor tissues (tissues of the negative surgical margin) and H8 cells, respectively. Higher miR-29b-2-5p expression correlated with a better 5-year progression-free survival of cervical cancer. MiR-29b-2-5p was also associated with the indicators (tumor size, tumor differentiation, FIGO (International Federation of Gynecology and Obstetrics) stage, and invasion depth) of the progression of cervical cancer tumors. And miR-29b-2-5p, along with tumor size, tumor differentiation, FIGO stage, histology type, and invasion depth, were independent prognostic factors for poor cervical cancer prognosis. MiR-29b-2-5p showed a suppressive effect on the proliferation, migration, and invasion of cervical cancer cells. MiR-29b-2-5p was downregulated in cervical cancer tumor tissues and could serve as an independent prognostic factor for cervical cancer. The overexpressed miR-29b-2-5p could be considered a tumor suppressor to inhibit the progression of cervical cancer.

Adenosine deaminase acting on RNA 1 (ADAR1), a recently described epigenetic modifier, is believed to play a critical oncogenic role in human cancers. However, its functional role and clinical significance in cervical cancer (CC) remain unclear. ADAR1 knockdown was performed to investigate its oncogenic functions in SiHa (HPV16), HeLa (HPV18), and Yumoto (non-HPV) CC cell lines. Cytoplasmic and nuclear ADAR1 expression were examined to clarify their correlation with clinicopathological parameters and prognosis in patients with CC. This resulted in increased apoptosis and necroptosis in HPV16 -type SiHa, HPV18-type HeLa, and non-HPV-type Yumoto CC cell lines. Progression-free survival (PFS) rates of patients exhibiting high cytoplasmic and nuclear ADAR1 expression were poorer than those in the other groups (P = 0.016). Multivariate analysis indicated that the combination of higher cytoplasmic and nuclear ADAR1 expression was an independent predictor of prognosis in patients with CC (P = 0.017). ADAR1 could be a potential therapeutic target for HPV-positive or HPV-negative CC. The combination of cytoplasmic and nuclear ADAR1 comprises a better prognostic factor for CC.