Polo-like kinase 4 promotes tumorigenesis and glucose metabolism in glioma by activating AKT1 signaling.

Glioblastoma (GBM) is an extremely aggressive tumor associated with a poor prognosis that impacts the central nervous system. Increasing evidence suggests an inherent association between glucose metabolism dysregulation and the aggression of GBM. Polo-like kinase 4 (PLK4), a highly conserved serine/threonine protein kinase, was found to relate to glioma progression and unfavorable prognosis. As revealed by the integration of proteomics and phosphoproteomics, PLK4 was found to be involved in governing metabolic processes and the PI3K/AKT/mTOR pathway. For the first time, this study supports evidence demonstrating that PLK4 activated PI3K/AKT/mTOR signaling through direct binding to AKT1 and subsequent phosphorylating AKT1 at S124, T308, and S473 to promote tumorigenesis and glucose metabolism in glioma. In addition, PLK4-mediated phosphorylation of AKT1 S124 significantly augmented the phosphorylation of AKT1 S473. Therefore, PLK4 exerted an influence on glucose metabolism by stimulating PI3K/AKT/mTOR signaling. Additionally, the expression of PLK4 protein exhibited a positive correlation with AKT1 phosphorylation in glioma patient tissues. These findings highlight the pivotal role of PLK4-mediated phosphorylation of AKT1 in glioma tumorigenesis and dysregulation of glucose metabolism.

Long non­coding RNAs in gallbladder cancer: From mechanisms to therapeutic opportunities (Review).

Due to the lack of specific symptoms, characteristic diagnostic markers and effective comprehensive treatment, gallbladder cancer (GBC) is currently considered one of the most malignant abdominal tumors. With the rapid development of biological technologies, long non­coding RNAs (lncRNAs), once regarded as transcriptional junk, have been demonstrated to participate in almost the whole process of the central dogma of molecular biology. The central dogma deals with the transfer of sequential information at the level of individual residues. LncRNAs have an effect on multiple cancer types. However, evidence of dysregulated lncRNA functions in GBC is limited. In the present review, the regulatory mechanisms of lncRNA function on gene expression were examined, including epigenetic modification, transcriptional regulation and post­translational modulation. These mechanisms are strongly associated with tumor development and metastasis. Next, it was summarized how lncRNAs affect GBC diverse malignant phenotypes through various mechanisms. Moreover, predictions of lncRNA interactions with other functional molecules in malignancies were made using several valuable databases, including crosstalk between lncRNA and DNA, mRNA, microRNA, and protein. Additionally, several potential therapeutic methods targeting pathological lncRNAs in tumors were identified. Finally, perspectives about lncRNA research and applications in GBC were presented in the current study, including viewpoints of coding potential of lncRNAs and feasible usage of micropeptides encoded by lncRNAs; roles of lncRNAs in tumor cell metabolic reprogramming and tumor microenvironment; and function of lncRNAs as possible biomarkers and therapeutic targets for improving GBC diagnosis, treatment and prognosis.

PEX13 is a potential immunotherapeutic indicator and prognostic biomarker for various tumors including PAAD.

The peroxisome serves a significant role in the occurrence and development of cancers. Specifically, the peroxisomal biogenesis factor 13 (PEX13) is crucial to the occurrence of peroxisomes. However, the biological function of PEX13 in cancers remains unclear. To address this, various portals and databases such as The Cancer Genome Atlas Program, The Genotype-Tissue Expression project, the Gene Expression Profiling Interactive Analysis 2, cBioPortal, the Genomic Identification of Significant Targets In Cancer 2.0, Tumor Immune Estimation Resource 2, SangerBox, LinkedOmics, DAVID and STRING were applied to extract and analyze PEX13 data in tumors. The correlations between PEX13 and prognosis, genetic alterations, PEX13-related gene enrichment analysis, weighted gene co-expression network analysis (WGCNA), protein interaction, long non-coding (lnc)RNA/circular (circ)RNA-micro (mi)RNA network and tumor immunity were explored in various tumors. The lncRNA-miRNA-PEX13 and circRNA-miRNA-PEX13 regulatory networks were identified via miRabel, miRDB, TargetScan and ENCORI portals and Cytoscape tool. In vitro assays were applied to verify the biological functions of PEX13 in pancreatic adenocarcinoma (PAAD) cells. The findings revealed that PEX13 is upregulated in various tumors and high PEX13 mRNA expression is associated with poor prognosis in patients with multiple cancers. Genetic alterations in PEX13 such as amplification, mutation and deep deletion have been found in multiple cancers. PEX13-related genes were associated with T cell receptor, signaling pathway and hippo signaling pathway through 'biological process' subontology of Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses. Through WGCNA analysis, it was discovered that PEX13 hub genes were mainly enriched in the Rap1, ErbB and AMPK signaling pathways in PAAD. Immune analysis showed that PEX13 was significantly related to tumor infiltration immune cells, immune checkpoint genes, microsatellite instability, TMB and tumor purity in a variety of tumors. Cell Counting Kit-8, wound healing, Transwell and colony formation assays displayed that PEX13 knockdown could suppress PAAD cell proliferation, migration, invasion, and colony formation in vitro, respectively. Overall, PEX13 is a potential predictor of immunotherapeutic and prognostic biomarkers in various malignant tumors, including ACC, KICH, LGG, LIHC and PAAD.

LncRNA HOXA11-AS promotes glioma malignant phenotypes and reduces its sensitivity to ROS via Tpl2-MEK1/2-ERK1/2 pathway.

Our previous studies showed that dysregulation of the long noncoding RNA (lncRNA) HOXA11-AS plays an important role in the development of glioma. However, the molecular mechanism of HOXA11-AS in glioma remains largely unknown. In this study, we explore the molecular mechanisms underlying abnormal expression and biological function of HOXA11-AS for identifying novel therapeutic targets in glioma. The expression of HOXA11-AS, and the relationship between HOXA11-AS and the prognosis of glioma patients were analyzed using databases and glioma samples. Transcriptomics, proteomics, RIP, ChIRP, luciferase, and ChIP assays were used to explore its upstream and downstream targets in glioma. The role of HOXA11-AS in regulating the sensitivity of glioma cells to reactive oxygen species (ROS) was also investigated in vitro and in vivo. We found that HOXA11-AS was significantly upregulated in glioma, and was correlated with the poor prognosis of glioma patients. Ectopic expression of HOXA11-AS promoted the proliferation, migration, and invasion of glioma cells in vitro and in vivo. Mechanistically, HOXA11-AS acted as a molecular sponge for let-7b-5p in the cytoplasm, antagonizing its ability to repress the expression of CTHRC1, which activates the ß-catenin/c-Myc pathway. In addition, c-Myc was involved in HOXA11-AS dysregulation via binding to its promoter region to form a self-activating loop. HOXA11-AS, functioned as a scaffold in the nucleus, also recruited transcription factor c-Jun to the Tpl2 promoter, which activates the Tpl2-MEK1/2-ERK1/2 pathway to promote ROS resistance in glioma. Importantly, HOXA11-AS knockdown could sensitize glioma cells to ROS. Above, oncogenic HOXA11-AS upregulates CTHRC1 expression as a ceRNA by adsorbing let-7b-5p, which activates c-Myc to regulate itself transcription. HOXA11-AS knockdown promotes ROS sensitivity in glioma cells by regulating the Tpl2-MEK1/2-ERK1/2 axis, demonstrating that HOXA11-AS may be translated to increase ROS sensitivity therapeutically.

Novel insight into the functions of N6­methyladenosine modified lncRNAs in cancers (Review).

Emerging evidence has suggested that N6­methyladenosine (m6A) modification, a typical RNA methylation modification, controls the fate of modified transcripts and is involved in the pathogenesis of various human diseases, such as metabolic disorders, nephropathology, osteoarthritis and malignant tumours. Long noncoding RNAs (lncRNAs), transcripts of >200 nt in length, have also been indicated to be involved in various diseases by participating in processes such as epigenetic modifications, transcriptional alternations and posttranslational regulation. Recent studies revealed that lncRNAs were widely modified by m6A, which has a critical role in various cellular processes that are associated with numerous disorders, particularly human cancers. The present review first examined functions of m6A modification of lncRNAs, including changing the lncRNA structure, mediating transcriptional regulation, affecting mRNA precursor splicing, and regulating lncRNA stability and translation. Furthermore, the regulatory mechanisms of m6A­modified lncRNAs in cancers were summarized and the up­to­date detection methods and prediction tools for identifying m6A sites on lncRNAs were presented. In addition, viewpoints on potential future directions in the field were discussed, including more accurate detection methods, roles of lncRNAs­encoded micropeptides in cancers, the relationship between m6A­modified lncRNAs and the tumour microenvironment, and m6A­modified lncRNAs as potential biomarkers and therapeutic targets in human cancer.

Construction of a Colorectal Cancer Prognostic Risk Model and Screening of Prognostic Risk Genes Using Machine-Learning Algorithms.

This study is aimed at constructing a prognostic risk model for colorectal cancer (CRC) using machine-learning algorithms to provide accurate staging and screening of credible prognostic risk genes. We extracted CRC data from GSE126092 and GSE156355 of the Gene Expression Omnibus (GEO) database and datasets from TCGA to analyze the differentially expressed genes (DEGs) using bioinformatics analysis. Among the 330 shared DEGs related to CRC prognosis, we divided the analysis period into different phases and applied univariate COX regression, LASSO, and multivariate COX regression analysis. GO analysis and KEGG analysis revealed that the functions of these DEGs were primarily focused on cell cycle, DNA replication, cell mitosis, and other related functions, and this confirmed our results from a biological perspective. Finally, a prognostic risk model for CRC based on the CHGA, CLU, PLK1, AXIN2, NR3C2, IL17RB, GCG, and AJUBA genes was constructed, and the risk score enabled us to predict the prognosis for CRC. To obtain a comprehensive and accurate model, we used both internal and external evaluations, and the model was able to correctly differentiate patients with CRC into a high-risk group with poor prognosis and a low-risk group with good prognosis. The AUC values of the 3-, 5-, and 10-year survival ROC curves were 0.715, 0.721, and 0.777, respectively, according to the internal evaluation, and the AUC values were 0.606, 0.698, and 0.608, respectively, for the external evaluation using GSE39582 from the GEO database. We determined that CLU, PLK1, and IL17RB could be considered to be independent prognostic factors for CRC with significantly different expression (P < 0.05). Using machine-learning methods, a prognostic risk model comprised of eight genes was constructed. Not only does this model provide improved treatment guidance, but it also provides a novel perspective for analyzing survival conditions at a deeper biological level.

Corrigendum: Pan-Cancer Analysis Shows That ALKBH5 Is a Potential Prognostic and Immunotherapeutic Biomarker for Multiple Cancer Types Including Gliomas.

[This corrects the article DOI: 10.3389/fimmu.2022.849592.].

Pan-Cancer Analysis Shows That ALKBH5 Is a Potential Prognostic and Immunotherapeutic Biomarker for Multiple Cancer Types Including Gliomas.

Background: AlkB homolog 5 (ALKBH5) is a N6-methyladenosine (m6A) demethylase associated with the development, growth, and progression of multiple cancer types. However, the biological role of ALKBH5 has not been investigated in pan-cancer datasets. Therefore, in this study, comprehensive bioinformatics analysis of pan-cancer datasets was performed to determine the mechanisms through which ALKBH5 regulates tumorigenesis. Methods: Online websites and databases such as NCBI, UCSC, CCLE, HPA, TIMER2, GEPIA2, cBioPortal, UALCAN, STRING, SangerBox, ImmuCellAl, xCell, and GenePattern were used to extract data of ALKBH5 in multiple cancers. The pan-cancer patient datasets were analyzed to determine the relationship between ALKBH5 expression, genetic alterations, methylation status, and tumor immunity. Targetscan, miRWalk, miRDB, miRabel, LncBase databases and Cytoscape tool were used to identify microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) that regulate expression of ALKBH5 and construct the lncRNA-miRNA-ALKBH5 network. In vitro CCK-8, wound healing, Transwell and M2 macrophage infiltration assays as well as in vivo xenograft animal experiments were performed to determine the biological functions of ALKBH5 in glioma cells. Results: The pan-cancer analysis showed that ALKBH5 was upregulated in several solid tumors. ALKBH5 expression significantly correlated with the prognosis of cancer patients. Genetic alterations including duplications and deep mutations of the ALKBH5 gene were identified in several cancer types. Alterations in the ALKBH5 gene correlated with tumor prognosis. GO and KEGG enrichment analyses showed that ALKBH5-related genes were enriched in the inflammatory, metabolic, and immune signaling pathways in glioma. ALKBH5 expression correlated with the expression of immune checkpoint (ICP) genes, and influenced sensitivity to immunotherapy. We constructed a lncRNA-miRNA network that regulates ALKBH5 expression in tumor development and progression. In vitro and in vivo experiments showed that ALKBH5 promoted proliferation, migration, and invasion of glioma cells and recruited the M2 macrophage to glioma cells. Conclusions: ALKBH5 was overexpressed in multiple cancer types and promoted the development and progression of cancers through several mechanisms including regulation of the tumor-infiltration of immune cells. Our study shows that ALKBH5 is a promising prognostic and immunotherapeutic biomarker in some malignant tumors.

The lncRNA PVT1 regulates autophagy in regulatory T cells to suppress heart transplant rejection in mice by targeting miR-146a.

Regulatory T cells (Tregs) are indispensable for the maintenance of immune tolerance. The purpose of this study was to investigate the effect of the interaction of the lncRNA PVT1 and miR-146a on Treg autophagy and reveal the mechanism to alleviate transplant rejection. PVT1 and miR-146a expression levels were analyzed by qRT-PCR. Bioinformatic analysis and methylation profiling were used to determine the relationship between PVT1 and miR-146a. Altered autophagic status in Tregs was detected by western blotting. The effect of autophagy on Treg function was assessed in cell coculture in vitro and in animal models. Our results showed that PVT1 expression was reduced in Tregs during rejection and negatively correlated with miR-146a expression. Higher PVT1 expression was associated with higher autophagy in Tregs. Further, highly autophagic Tregs had stronger inhibitory effects on CD4+ T cells in vitro, prolonged allograft survival and alleviated rejection in vivo. Mechanistic studies showed that overexpression of PVT1 enhanced TNF receptor-associated factor (TRAF) 6 expression by directly targeting miR-146a. MiR-146a overexpression reversed PVT1-induced Treg autophagy and inhibited PVT1-induced TRAF6 expression. The present study shows a novel regulatory pathway of the autophagy program that comprises PVT1, miR-146a, and TRAF6. Our findings may provide potential targets and new therapeutic strategies for transplant rejection.

MiR-146a regulates regulatory T cells to suppress heart transplant rejection in mice.

Regulatory T cells (Tregs), which characteristically express forkhead box protein 3 (Foxp3), are essential for the induction of immune tolerance. Here, we investigated microRNA-146a (miR-146a), a miRNA that is widely expressed in Tregs and closely related to their homeostasis and function, with the aim of enhancing the function of Tregs by regulating miR-146a and then suppressing transplant rejection. The effect of the absence of miR-146a on Treg function in the presence or absence of rapamycin was detected in both a mouse heart transplantation model and cell co-cultures in vitro. The absence of miR-146a exerted a mild tissue-protective effect by transiently prolonging allograft survival and reducing the infiltration of CD4+ and CD8+ T cells into the allografts. Meanwhile, the absence of miR-146a increased Treg expansion but impaired the ability of Tregs to restrict T helper cell type 1 (Th1) responses. A miR-146a deficiency combined with interferon (IFN)-γ blockade repaired the impaired Treg function, further prolonged allograft survival, and alleviated rejection. Importantly, miR-146a regulated Tregs mainly through the IFN-γ/signal transducer and activator of transcription (STAT) 1 pathway, which is implicated in Treg function to inhibit Th1 responses. Our data suggest miR-146a controls a specific aspect of Treg function, and modulation of miR-146a may enhance Treg efficacy in alleviating heart transplant rejection in mice.

Regulatory T cells induce transplant immune tolerance.

Organ transplantation is the preferred treatment option for end-stage organ failure. Although immunosuppressants are effective for preventing the occurrence of acute rejection, they also cause a series of side effects in transplant recipients. To improve the quality of patient survival, a new therapeutic strategy that has fewer side effects than current immunosuppressive regimens and can induce allograft immune tolerance and effectively prevent transplant rejection is needed. In this context, regulatory T cells (Tregs) are considered to be promising research targets. With the increasing understanding of the immunomodulatory role of Tregs, the use of Treg-based cellular therapies has shifted from prevention/treatment of autoimmune diseases to clinical trials for organ transplantation. This review describes the phenotype and in vitro expansion of Tregs and the mechanisms by which they exert immunomodulatory effects in transplantation immunity, highlights recent clinical trial data on Treg-based cellular therapies in transplantation, and describes future directions and limitations.

Mesenchymal stem cells transfected with sFgl2 inhibit the acute rejection of heart transplantation in mice by regulating macrophage activation.

BACKGROUND: Mesenchymal stem cells (MSCs) have become a promising candidate for cell-based immune therapy for acute rejection (AR) after heart transplantation due to possessing immunomodulatory properties. In this study, we evaluated the efficacy of soluble fibronectin-like protein 2 (sFgl2) overexpressing mesenchymal stem cells (sFgl2-MSCs) in inhibiting AR of heart transplantation in mice by regulating immune tolerance through inducing M2 phenotype macrophage polarization. METHODS AND RESULTS: The sFgl2, a novel immunomodulatory factor secreted by regulatory T cells, was transfected into MSCs to enhance their immunosuppressive functions. After being co-cultured for 72 h, the sFgl2-MSCs inhibited M1 polarization whereas promoted M2 of polarization macrophages through STAT1 and NF-κB pathways in vitro. Besides, the sFgl2-MSCs significantly enhanced the migration and phagocytosis ability of macrophages stimulated with interferon-γ (IFN-γ) and lipopolysaccharide (LPS). Further, the application potential of sFgl2-MSCs in AR treatment was demonstrated by heterotopic cardiac transplantation in mice. The tissue damage and macrophage infiltration were evaluated by H&E and immunohistochemistry staining, and the secretion of inflammatory cytokines was analyzed by ELISA. The results showed that sFgl2-MSCs injected intravenously were able to locate in the graft, promote the M2 polarization of macrophages in vivo, regulate the local and systemic immune response, significantly protect tissues from damaging, and finally prolonged the survival time of mice heart grafts. CONCLUSION: sFgl2-MSCs ameliorate AR of heart transplantation by regulating macrophages, which provides a new idea for the development of anti-AR treatment methods after heart transplantation.