Integrated clustering signature of genomic heterogeneity, stemness and tumor microenvironment predicts glioma prognosis and immunotherapy response.

BACKGROUND: Glioma is the most frequent primary tumor of the central nervous system. The high heterogeneity of glioma tumors enables them to adapt to challenging environments, leading to resistance to treatment. Therefore, to detect the driving factors and improve the prognosis of glioma, it is essential to have a comprehensive understanding of the genomic heterogeneity, stemness, and immune microenvironment of glioma. METHODS: We classified gliomas into various subtypes based on stemness, genomic heterogeneity, and immune microenvironment consensus clustering analysis. We identified risk hub genes linked to heterogeneous characteristics using WGCNA, LASSO, and multivariate Cox regression analysis and utilized them to create an effective risk model. RESULTS: We thoroughly investigated the genomic heterogeneity, stemness, and immune microenvironment of glioma and identified the risk hub genes RAB42, SH2D4A, and GDF15 based on the TCGA dataset. We developed a risk model utilizing these genes that can reliably predict the prognosis of glioma patients. The risk signature showed a positive correlation with T cell exhaustion and increased infiltration of immunosuppressive cells, and a negative correlation with the response to immunotherapy. Moreover, we discovered that SH2D4A, one of the risk hub genes, could stimulate the migration and proliferation of glioma cells. CONCLUSIONS: This study identified risk hub genes and established a risk model by analyzing the genomic heterogeneity, stemness, and immune microenvironment of glioma. Our findings will facilitate the diagnosis and prediction of glioma prognosis and may lead to potential treatment strategies for glioma.

Application of Induced Pluripotent Stem Cells in Moyamoya Disease: Progress and Promises.

Moyamoya disease (MMD) is a chronic steno-occlusion cerebrovascular disease accompanied by the formation of the abnormal vascular network at the base of the brain. The etiology of MMD is not fully clarified. Lack of pathological specimens hinders the research progress. Induced pluripotent stem cells (iPSC) derived from patients with outstanding differentiation potential and infinite proliferation ability could conquer the problem of insufficient samples. The technology of iPSC holds the promise of clarifying the underlying molecular mechanism in the development of MMD. In this review, we summarized the latest progress and difficulties in the research of mechanism and detailed the application of iPSC in MMD, aiming to provide an outlook of iPSC in molecular mechanism and novel therapies of MMD.

RNA N1-methyladenosine regulator-mediated methylation modification patterns and heterogeneous signatures in glioma.

N1-methyladenosine (m1A) is ubiquitous in eukaryotic RNA and regulates mRNA translation. However, little is known about its regulatory role in glioma. Here, we identified 4 m1A modification-related patterns based on m1A regulators in the TCGA (The Cancer Genome Atlas) and CGGA (Chinese Glioma Genome Atlas) cohorts. The differences in survival prognosis between different clusters were striking. In addition, stemness, genomic heterogeneity, tumor microenvironment (TME), and immune cell infiltration were also significantly different between the poor and best prognostic clusters. To reveal the underlying mechanism, differentially expressed genes (DEGs) between the poor and best prognostic clusters were identified, and then were integrated for weighted correlation network analysis (WGCNA). After Univariate Cox-LASSO-Multivariate Cox analyses, DEGs PLEK2 and ABCC3 were screened as the risk-hub genes and were selected to construct an m1A-related signature. Moreover, ABCC3 exacerbated glioma proliferation and was associated with temozolomide (TMZ) resistance. Overall, our study provided new insights into the function and potential therapeutic role of m1A in glioma.

A Novel TAF-Related Signature Based on ECM Remodeling Genes Predicts Glioma Prognosis.

The composition and abundance of immune and stromal cells in the tumor microenvironment (TME) dramatically affect prognosis. Infiltration of immunosuppressive tumor-associated fibroblasts (TAFs) is a hallmark of glioma. However, the mechanisms regulating TAF infiltration and the prognostic value of TAF-related genes in glioma remain unclear. In this study, we analyzed TAF infiltration by Estimating the Proportion of Immune and Cancer cells (EPIC) algorithm based on multiple glioma databases, including Glioblastoma and low-grade glioma merged cohort from The Cancer Genome Atlas (TCGA GBMLGG) cohort, the Chinese Glioma Genome Atlas (CGGA) #325 cohort, and the CGGA #693 cohort. TAF infiltration was increased in glioblastoma (GBM), and elevated TAF infiltration predicted poorer survival in gliomas. Gene enrichment analyses revealed that differentially expressed genes (DEGs) between low-grade glioma (LGG) and GBM were significantly enriched in the extracellular matrix (ECM) remodeling-related signaling, which may contribute to immune escape and resistance to immune checkpoint blockers (ICBs). To identify co-expression modules and candidate hub genes that may be associated with TAF infiltration, we performed weighted correlation network analysis (WGCNA) of DEGs. Afterward, univariate Cox regression, least absolute shrinkage and selection operator (LASSO) regression, and multivariate Cox regression analyses were performed to screen the positive prognostic hub genes. Finally, a high-efficacy prediction signature was constructed based on the expression of S100A4, PLAUR, and EMP3. The signature correlated with the abundance of TAF infiltration in glioma and was an independent risk factor for glioma. In conclusion, our findings suggested that the TAF-related signature was a valuable prognostic biomarker in glioma and provided potential targets for integrative therapy of gliomas.

Alarm Signal S100-Related Signature Is Correlated with Tumor Microenvironment and Predicts Prognosis in Glioma.

Glioma are the most common malignant central nervous system tumor and are characterized by uncontrolled proliferation and resistance to therapy. Dysregulation of S100 proteins may augment tumor initiation, proliferation, and metastasis by modulating immune response. However, the comprehensive function and prognostic value of S100 proteins in glioma remain unclear. Here, we explored the expression profiles of 17 S100 family genes and constructed a high-efficient prediction model for glioma based on CGGA and TCGA datasets. Immune landscape analysis displayed that the distribution of immune scores, ESTIMATE scores, and stromal scores, as well as infiltrating immune cells (macrophages M0/M1/M2, T cell CD4+ naïve, Tregs, monocyte, neutrophil, and NK activated), were significant different between risk-score subgroups. Overall, we demonstrated the value of S100 protein-related signature in the prediction of glioma patients' prognosis and determined its relationship with the tumor microenvironment (TME) in glioma.

Tumor-associated macrophages related signature in glioma.

BACKGROUND: Glioma is the most common malignant primary tumor with a poor prognosis. Infiltration of tumor-associated macrophages (TAMs) is a hallmark of glioma. However, the regulatory mechanism of TAMs and the prognostic value of related signature in glioma remain unclear. METHODS: TAMs were analyzed by EPIC, MCPCOUNTER and XCELL methods in multiple cohorts, including the TCGA merged GBMLGG, CGGA mRNAseq-325, and CGGA mRNAseq-693. Weighted correlation network analysis (WGCNA) were performed to identify candidate hub genes that might be related to TAMs. The prognostic genes were selected by Univariate Cox regression, Kaplan-Meier analysis and the least absolute shrinkage and selection operator (LASSO) multivariate Cox regression algorithm, and were used to construct a high efficacy prediction model. RESULTS: Compared with LGG, TAMs of GBM in the TCGA merged GBMLGG, CGGA mRNAseq-693, and CGGA mRNAseq-325 cohorts were increased, and high TAMs levels predicted poorer overall survival for gliomas. The prediction model constructed by nine prognostic genes was highly efficient. The TAMs related risk-score was an independent risk factor for glioma. Moreover, high risk score was correlated with an increased population of TAMs in glioma, as well as the high immune scores, stromal scores and ESTIMATE scores. CONCLUSIONS: Increased TAMs might be an immune evasion mechanism of glioma. In addition, our findings suggested that TAMs-related signature was a valuable prognostic biomarker in glioma and provided therapeutic targets for glioma.

Thermal Decomposition Kinetics and Compatibility of 3,5-difluoro-2,4,6-trinitroanisole (DFTNAN).

In this paper, the thermal decomposition behavior of 3,5-difluoro-2,4,6-trinitroanisole (DFTNAN) was studied by differential scanning calorimetry (DSC) and thermogravimetry (TG) by using different heating rates (2, 5, 10, 15 °C·min-1). Subsequently, the kinetic and thermodynamic parameters of non-isothermal thermal decomposition of DFTNAN were calculated. The critical temperature of thermal explosion (Tb) and self-accelerating decomposition temperature (TASDT) were determined to be 249.03 °C and 226.33 °C, respectively. The compatibility of DFTNAN with a number of high explosives (cyclo-1,3,5-trimethylene-2,4,6-trinitramine (RDX), 1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX), 2,4,6,8,10,12-hexanitro-2,4,6,8,10,12-hexaaza-tetracyclo-[5.5.0.05,9.03,11]-dodecane (CL-20) and dihydroxylammonium 5,5'-bistetrazole-1,1'-diolate (TKX-50)) was studied at different mass ratios using DSC. The criteria to judge the compatibility between the materials were based on a standardization agreement (STANAG 4147). The thermodynamic study results revealed that DFTNAN possessed superior thermal safety and stability. The experimental of compatibility results indicated that the mass ratios of the high explosives in the DFTNAN/RDX, DFTNAN/HMX and DFTNAN/CL-20 compositions more than 40%, 60% and 70% exhibited good compatibility, whereas DFTNAN/TKX-50 demonstrated poor compatibility.

Microglial Polarization: Novel Therapeutic Strategy against Ischemic Stroke.

Ischemic stroke, which is the second highest cause of death and the leading cause of disability, represents ~71% of all strokes globally. Some studies have found that the key elements of the pathobiology of stroke is immunity and inflammation. Microglia are the first line of defense in the nervous system. After stroke, the activated microglia become a double-edged sword, with distinct phenotypic changes to the deleterious M1 types and neuroprotective M2 types. Therefore, ways to promote microglial polarization toward M2 phenotype after stroke have become the focus of attention in recent years. In this review, we discuss the process of microglial polarization, summarize the alternation of signaling pathways and epigenetic regulation that control microglial polarization in ischemic stroke, aiming to find the potential mechanisms by which microglia can be transformed into the M2 polarized phenotype.

Gene Expression-Based Predication of RNA Pseudouridine Modification in Tumor Microenvironment and Prognosis of Glioma Patients.

Aberrant expression of methyltransferases and demethylases may augment tumor initiation, proliferation and metastasis through RNA modification, such as m6A and m5C. However, activity of pseudouridine (Ψ) modification of RNA remains unknown in glioma, the most common malignant intracranial tumor. In this study, we explored the expression profiles of the Ψ synthase genes in glioma and constructed an efficient prediction model for glioma prognosis based on the CGGA and TCGA datasets. In addition, the risk-score signature was positively associated with malignancy of gliomas and the abundance of tumor-infiltrating immune cells such as macrophages M0 and regulatory T cells (Tregs), but negatively associated with the abundance of monocytes, NK cell activation and T cell CD4+ naive. In terms of mechanism, the risk-score signature was positively associated with the expression of inflammatory molecules such as S100A11 and CASP4 in glioma. Overall, this study provided evidence for the activity of RNA Ψ modification in glioma malignancy and local immunity.

N6-methyladenosine methyltransferase METTL3 affects the phenotype of cerebral arteriovenous malformation via modulating Notch signaling pathway.

BACKGROUND: Cerebral arteriovenous malformation (AVM) is a serious life-threatening congenital cerebrovascular disease. Specific anatomical features, such as nidus size, location, and venous drainage, have been validated to affect treatment outcomes. Until recently, molecular biomarkers and corresponding molecular mechanism related to anatomical features and treatment outcomes remain unknown. METHODS: RNA N6-methyladenosine (m6A) Methyltransferase METTL3 was identified as a differentially expressed gene in groups with different lesion sizes by analyzing the transcriptome sequencing (RNA-seq) data. Tube formation and wound healing assays were performed to investigate the effect of METTL3 on angiogenesis. In addition, Methylated RNA Immunoprecipitation Sequencing technology (MeRIP-seq) was performed to screen downstream targets of METTL3 in endothelial cells and to fully clarify the specific underlying molecular mechanisms affecting the phenotype of cerebral AVM. RESULTS: In the current study, we found that the expression level of METTL3 was reduced in the larger pathological tissues of cerebral AVMs. Moreover, knockdown of METTL3 significantly affected angiogenesis of the human endothelial cells. Mechanistically, down-regulation of METTL3 reduced the level of heterodimeric Notch E3 ubiquitin ligase formed by DTX1 and DTX3L, thereby continuously activating the Notch signaling pathway. Ultimately, the up-regulated downstream genes of Notch signaling pathway dramatically affected the angiogenesis of endothelial cells. In addition, we demonstrated that blocking Notch pathway with DAPT could restore the phenotype of METTL3 deficient endothelial cells. CONCLUSIONS: Our findings revealed the mechanism by which m6A modification regulated the angiogenesis and might provide potential biomarkers to predict the outcome of treatment, as well as provide suitable pharmacological targets for preventing the formation and progression of cerebral AVM.

Wilms' tumour 1-associating protein inhibits endothelial cell angiogenesis by m6A-dependent epigenetic silencing of desmoplakin in brain arteriovenous malformation.

Brain arteriovenous malformations (AVMs) are congenital vascular abnormality in which arteries and veins connect directly without an intervening capillary bed. So far, the pathogenesis of brain AVMs remains unclear. Here, we found that Wilms' tumour 1-associating protein (WTAP), which has been identified as a key subunit of the m6A methyltransferase complex, was down-regulated in brain AVM lesions. Furthermore, the lack of WTAP could inhibit endothelial cell angiogenesis in vitro. In order to screen for downstream targets of WTAP, we performed RNA transcriptome sequencing (RNA-seq) and Methylated RNA Immunoprecipitation Sequencing technology (MeRIP-seq) using WTAP-deficient and control endothelial cells. Finally, we determined that WTAP regulated Desmoplakin (DSP) expression through m6A modification, thereby affecting angiogenesis of endothelial cells. In addition, an increase in Wilms' tumour 1 (WT1) activity caused by WTAP deficiency resulted in substantial degradation of ß-catenin, which might also inhibit angiogenesis of endothelial cells. Collectively, our findings revealed the critical function of WTAP in angiogenesis and laid a solid foundation for the elucidation of the pathogenesis of brain AVMs.

A class of novel tubulin polymerization inhibitors exert effective anti-tumor activity via mitotic catastrophe.

In current work, a class of novel 4,5-dihydro-1H-pyrazole-1-carboxylate derivatives (E01-E28) were designed, synthesized and evaluated. Among them, the most potent compound E24 exhibited comparable activity against a panel of cancer cells (GI50 ranging 0.05-0.98 µM) and tubulin polymerization inhibition (IC50 = 1.49 µM) with reference drug CA-4(P) (GI50 ranging 0.019-0.32 µM, IC50 = 2.18 µM). The following assays indicated that compound E24 disturbed the dynamics of tubulin catastrophe and rescue, which triggered G2/M arrest, leading to ROS accumulation, cleavage of PARP and apoptosis. Molecular dynamics simulation validated that compound E24 could tightly bind into tubulin heterodimers with ß Lys 254 and ß Cys 241 of tubulin in the docking pose. Metabolic stability and pharmacokinetics parameters were also determined. The half time (t1/2) displayed species differences in three microsomes. The plasma elimination half-life (t1/2), peak plasma concentration (Cmax), mean retention time (MRT), the area under the curve (AUC0-∞) and distribution volume (Vz) of E24 after intravenous administration were 0.90 ± 0.22 h, 594.50 ± 97.23 ng/mL, 1.09 ± 0.22 h, 413.67 ± 105.64 ng/mL*h and 5.03 ± 1.82 L/kg, respectively. In HeLa-xenografts, compound E24 exhibited obvious antitumor efficacy via the suppression of tumor growth without weight loss of body or organ. In brief, compound E24 might be a hopeful candidate with excellent properties for oncotherapy as tubulin polymerization inhibitor.

Identification of the RNA Pseudoknot within the 3' End of the Porcine Reproductive and Respiratory Syndrome Virus Genome as a Pathogen-Associated Molecular Pattern To Activate Antiviral Signaling via RIG-I and Toll-Like Receptor 3.

Once infected by viruses, cells can detect pathogen-associated molecular patterns (PAMPs) on viral nucleic acid by host pattern recognition receptors (PRRs) to initiate the antiviral response. Porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of porcine reproductive and respiratory syndrome (PRRS), characterized by reproductive failure in sows and respiratory diseases in pigs of different ages. To date, the sensing mechanism of PRRSV has not been elucidated. Here, we reported that the pseudoknot region residing in the 3' untranslated regions (UTR) of the PRRSV genome, which has been proposed to regulate RNA synthesis and virus replication, was sensed as nonself by retinoic acid-inducible gene I (RIG-I) and Toll-like receptor 3 (TLR3) and strongly induced type I interferons (IFNs) and interferon-stimulated genes (ISGs) in porcine alveolar macrophages (PAMs). The interaction between the two stem-loops inside the pseudoknot structure was sufficient for IFN induction, since disruption of the pseudoknot interaction powerfully dampened the IFN induction. Furthermore, transfection of the 3' UTR pseudoknot transcripts in PAMs inhibited PRRSV replication in vitro Importantly, the predicted similar structures of other arterivirus members, including equine arteritis virus (EAV), lactate dehydrogenase-elevating virus (LDV), and simian hemorrhagic fever virus (SHFV), also displayed strong IFN induction activities. Together, in this work we identified an innate recognition mechanism by which the PRRSV 3' UTR pseudoknot region served as PAMPs of arteriviruses and activated innate immune signaling to produce IFNs that inhibit virus replication. All of these results provide novel insights into innate immune recognition during virus infection.IMPORTANCE PRRS is the most common viral disease in the pork industry. It is caused by PRRSV, a positive single-stranded RNA virus, whose infection often leads to persistent infection. To date, it is not yet clear how PRRSV is recognized by the host and what is the exact mechanism of IFN induction. Here, we investigated the nature of PAMPs on PRRSV and the associated PRRs. We found that the 3' UTR pseudoknot region of PRRSV, which has been proposed to regulate viral RNA synthesis, could act as PAMPs recognized by RIG-I and TLR3 to induce type I IFN production to suppress PRRSV infection. This report is the first detailed description of pattern recognition for PRRSV, which is important in understanding the antiviral response of arteriviruses, especially PRRSV, and extends our knowledge on virus recognition.

Genomic analysis of a recombinant NADC30-like porcine reproductive and respiratory syndrome virus in china.

Recently, NADC30-like porcine reproductive and respiratory syndrome viruses (PRRSVs), which are genetically similar to the NADC30 strain isolated in the United States of America in 2008, have become prevalent in China. Here, a novel variant PRRSV strain named HNhx was successfully isolated on porcine alveolar macrophages from Henan province and the full-length genome sequence was determined. Phylogenetic analysis indicated that HNhx strain was classified into the NADC30-like PRRSV subgroup, in which all the strains had the unique discontinuous 131-amino acid deletion relative to that of the nonstructural protein 2 (Nsp2) of the VR2332 strain. Genetically, HNhx shared 92.9% nucleotide similarity to NADC30. Furthermore, HNhx strain contained extensive amino acid mutations in GP5. In particular, the S32H, N33D, D34N, and S36G variations resulted in that HNhx lost all the putative N-linked glycosylation sites at amino acid positions 30, 32, 33, 34, and 35. Recombination analysis revealed that HNhx was the result of recombination between the NADC30 strain and the highly pathogenic PRRSV vaccine strain circulating in China in Nsp4 (nt 5261) to Nsp9 (nt 7911). The novel genome data of HNhx will be helpful for understanding the evolution and epidemiology of PRRSV in China.

Molecular epidemiology of porcine reproductive and respiratory syndrome virus in Central China since 2014: The prevalence of NADC30-like PRRSVs.

Porcine reproductive and respiratory syndrome (PRRS), characterized by respiratory disorders in piglets and reproductive failure in sows, is still the great threat of swine industry. Recently, Emergence of the novel NADC30-like PRRS viruses (PRRSVs) has caused widespread outbreaks of PRRS. To investigate the epidemic characteristics of PRRSVs in Central China since 2014, 6372 clinical serum samples were tested by ELISA, 250 tissue samples were tested by RT-PCR, and among these, 30 ORF5 and 17 Nsp2 genes sequences were analyzed. Phylogenetic tree based on ORF5 revealed that, 17 isolates were clustered into subgroup 1, represented by the NADC30. And for the Nsp2, The strains which had a discontinuous 131-amino-acid deletion in Nsp2, called NADC30-like strains, were clustered into subgroup 2. Our data suggested that the NADC30-like PRRSV strains spread quickly and are now circulating and prevalent in Central China as well as the classical HP-PRRSV strains. In addition, amino acid variation analysis of GP5 revealed that the amino acid sequences of NADC30-like PRRSV strains underwent rapid evolution and contained extensive amino acid substitutions in important motifs, such as potential neutralization epitope and the N-glycosylation sites. In summary, our data would provide a large amount of detailed information on molecular variation and genetic diversity of PRRSV in central China.

Complete Genome Sequence of a Mosaic NADC30-Like Porcine Reproductive and Respiratory Syndrome Virus in China.

Here, we report the complete genome of an NADC30-like porcine reproductive and respiratory syndrome virus (PRRSV) strain, HNhx, which was isolated from Henan Province, China, in 2016 and was characterized by recombination with JXA1 strain (an epidemic highly pathogenic PRRSV strain in China) in Nsp4 to Nsp9.