Transcriptional Misregulation Research Articles

Adaptive boost approach for possible leads of triple-negative breast cancer

Recent research has shown that cyclin dependent kinase 7 (CDK7), cyclin dependent kinase 9 (CDK9) and cyclin T1 (CCNT1) can assist transcriptional misregulation in cancer. We combined traditional virtual screening technology with artificial intelligence models to screen multi-target inhibitors from FDA database for the target proteins. R-square (R2) was chosen to evaluate the accuracy of these artificial intelligent models. For CDK7 inhibitors dataset, Adaptive Boosting-Decision Tree Regression (AdaBoost), Support Vector Machine Regression (SVR) and Ridge Regression (Ridge) achieved good results and all nine basic models had R2 more than 0.5; R2 of test set of former three models reached 0.886, 0.860 and 0.815. For CDK9 inhibitors dataset, AdaBoost, Random Forest (RF) and Ridge achieved good results; R2 of test set of these models reached 0.833, 0.788 and 0.759. It seems Adaptive Boosting and Ridge Regression has better generalization ability than other basic models. Adaptive Boosting use plenty of weaker regressors to combine a stronger regressor, which can help it control overfitting. Ridge Regression adds a penalty term on regularization . With the addition of penalty term, the estimation of regression coefficient is no longer unbiased. Therefore, ridge regression is a method to solve the ill-conditioned matrix problem at the cost of abandoning unbiasedness. In order to evaluate the stability of bonds between protein and possible leads, Molecular Dynamics (MD) simulation were performed to verify whether the possible leads were docked well in the protein binding sites. By analyzing the results of virtual screening, artificial intelligent models and MD experiments, we suggest ZINC3830891 (Glutathione) and ZINC19363537 (Tetraethylene pentamine) are the possible multi-target leads inhibitors for TNBC.

Transcriptome analysis identifies TODL as a novel lncRNA associated with proliferation, differentiation, and tumorigenesis in liposarcoma through FOXM1

Liposarcoma, the most common soft tissue sarcoma, is a group of fat cell mesenchymal tumors with different histological subtypes. The dysregulation of long non-coding RNAs (lncRNAs) has been observed in human cancers including a few studies in sarcoma. However, the global transcriptome analysis and potential role of lncRNAs remain unexplored in liposarcoma. The present investigation uncovers the transcriptomic profile of liposarcoma by RNA sequencing to gain insight into the global transcriptional changes in liposarcoma. Our RNA sequencing analysis has identified that many oncogenic lncRNAs are differentially expressed in different subtypes of liposarcoma including MALAT1, PVT1, SNHG15, LINC00152, and MIR210HG. Importantly, we identified a highly overexpressed, unannotated, and novel lncRNA in dedifferentiated liposarcomas. We have named it TODL, transcript overexpressed in dedifferentiated liposarcoma. TODL lncRNA displayed significantly higher expression in dedifferentiated liposarcoma cell lines and patient samples. Interestingly, functional studies revealed that TODL lncRNA has an oncogenic function in liposarcoma cells by regulating proliferation, cell cycle, apoptosis, differentiation, and tumorigenesis in the murine model. Silencing of TODL lncRNA highlighted the enrichment of several key oncogenic signaling pathways including cell cycle, transcriptional misregulation, FOXM1 network, p53 signaling, PLK1 signaling, FoxO, and signaling Aurora signaling pathways. RNA pull-down assay revealed the binding of TODL lncRNA with FOXM1, an oncogenic transcription factor, and the key regulator of the cell cycle. Silencing of TODL lncRNA also induces adipogenesis in dedifferentiated liposarcomas. Altogether, our finding indicates that TODL could be utilized as a novel, specific diagnostic biomarker, and a pharmacological target for therapeutic development in controlling aggressive and metastatic dedifferentiated liposarcomas.

Identification of key genes involved in calcific aortic valve disease based on integrated bioinformatics analysis.

The calcific aortic valve disease (CAVD) develops as an aortic valve sclerosis and progresses to an advanced form of stenosis. In many biological fields, bioinformatics becomes a fundamental component. The key mechanisms involved in CAVD are discovered with the use of bioinformatics to investigate gene function and pathways. We downloaded the original data (GSE51472) from the Gene Expression Omnibus (GEO) database (http://www.ncbi.nlm.nih.gov/geo/). After standardization, 2978 differentially expressed genes (DEGs) were identified from the data sets GSE51472 containing samples from normal, calcified, and sclerotic aortic valves. Analysis of DEGs based on the series test of clusters (STCs) revealed the two most significant patterns. Based on the result of the STC, the functional enrichment analysis of gene ontology (GO) was conducted to investigate the molecular function (MF), biological process (BP), and cell compound (CC) of the DEGs. With a p value of 0.01, DEGs associated with "chronic inflammation," "T-cell receptor complexes," and "antigen binding" had the highest significance within BP, CC, and MF. DEG enrichment in signaling pathways was analyzed using KEGG pathway enrichment. Using a p < 0.05 level of significance, the most enriched biological pathways related to CAVD were "Chemokine signaling pathway," "Cytokine-cytokine receptor interaction," "Tuberculosis," "PI3K-Akt signaling pathway," and "Transcriptional misregulation in cancer." Finally, the construction of gene co-expression networks and pathway networks illustrated the pathogensis of CAVD. TLR2, CD86, and TYROBP were identified as hub genes for the development of CAVD. Moreover, "MAPK signaling pathway," "Apoptosis," and "Pathways in cancer" were regarded as the core pathways among the samples of normal, sclerotic and calcified aortic valve samples.

MiR-22-3p as a potential biomarker for coronary artery disease based on integrated bioinformatics analysis.

Background: Coronary artery disease (CAD) is a common cardiovascular disease that has attracted attention worldwide due to its high morbidity and mortality. Recent studies have shown that abnormal microRNA (miRNA) expression is effective in CAD diagnoses and processes. However, the potential relationship between miRNAs and CAD remains unclear. Methods: Microarray datasets GSE105449 and GSE28858 were downloaded directly from the Gene Expression Omnibus (GEO) to identify miRNAs involved in CAD. Target gene prediction and enrichment analyses were performed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Results: There were nine differentially expressed miRNAs in CAD patients compared to the controls. A total of 352 genes were predicted and subjected to GO analysis, which showed that differentially expressed genes (DEGs) were mainly associated with axon guidance, neuron projection guidance, neuron-to-neuron synapses, and postsynaptic density. According to the KEGG pathway analysis, the most enriched pathways were those involved in transcriptional misregulation in cancer, growth hormone synthesis, secretion and action, endocrine resistance, axon guidance, and Cushing syndrome. Pathway analysis was mainly involved in the HIPPO and prion disease signaling pathways. Furthermore, a competing endogenous RNA (ceRNA) interaction network centered on miR-22-3p revealed eight related transcription factors in the cardiovascular system. The receiver operating characteristic (ROC) curve analysis suggested that miR-22-3p may be a better CAD predictor. Conclusion: The results indicate that miR-22-3p may function in pathophysiological CAD processes. Our study potentiates miR-22-3p as a specific biomarker for diagnosing CAD.

Natural shikonin and acetyl-shikonin improve intestinal microbial and protein composition to alleviate colitis-associated colorectal cancer

BackgroundColorectal cancer (CRC) and inflammatory bowel disease (IBD) are the most common diseases of human digestive system. Nowadays, the influence of the inflammatory microenvironment on tumorigenesis has become a new direction, and the exploration of relative molecular mechanism will facilitate the discovery and identification of novel potential anti-cancer molecules. MethodsNatural shikonin (SK) and acetyl-shikonin (acetyl-SK) was administered to azoxymethane (AOM)/dextran sodium sulphate (DSS)-induced colitis-associated colorectal cancer (CAC) mice model by gavage to investigate their therapeutic effects. Moreover, fresh feces and colon tissues were collected for determining the function of SK and acetyl-SK on the gut microbes and protein expression, respectively. ResultsBoth SK and acetyl-SK decreased AOM/DSS-induced CAC, and regulated the intestinal flora structure in CAC mouse model. They, especially SK, improved species richness, evenness and diversity of intestinal flora, recovered the upregulated ratio of Firmicutes to Bacteroidota (F/B ratio) which symbolizes gut microbiota dysbiosis. SK and its derivative increased the beneficial bacteria g__norank_f__Muribaculaceae, Lactobacillus, Lachnospiraceae_NK4A136_Group, and reduced those harmful ones including Ileibacterium and Coriobacteriaceae UCG–002. Notably, AOM/DSS caused significant increase in the abundance of Ileibaterium valens and g__norank_f__norank_o__Clostridia_UCG-014, which were not previously reported in studies of colonic inflammation or cancer, and the disorder was reversed by 20 mg/kg of SK. In our current study, the action of SK and acetyl-SK is dose-dependent, and 20 mg/kg SK exhibited the most effective functions, even better than the positive drug mesalazine.Moreover, differential proteomics and ELISA results showed that SK could recover the increase of pro-inflammatory cytokines (including IL-1β, IL-6 and TNF-α), the upregulation of pyruvate kinase isozyme type M2 (PKM2) and some other proteins (mainly concentrated in transcriptional mis-regulation in cancer and IL-17 signaling pathways), and the downregulation of Aldh1b1-Acc3-Maoa and Μgt2b34-Aldh1a1-Aldh1a7 involved in Wnt/β-catenin signaling pathway. ConclusionOur study identified SK and acetyl-SK, especially SK, as potential preventive agents for CAC through regulating both gut microbes and pathways involved in inflammation and cancer such as Wnt/β-catenin signaling pathway.

MicroRNA expression profiles in extracellular vesicles and intracellular of AURKA inhibitor-induced senescent neuroblastoma cells.

BackgroundTherapy-induced senescence plays an important role in the clinical treatment of tumors, which is positively correlated with the treatment response. However, senescent cells reshape tumor microenvironment and increase the risk of cancer recurrence. Regarded as hormones to regulate cell-to-cell communication, microRNA (miRNA) is highly sensitive to environmental stress. To understand the response of tumor cells to chemotherapeutic drugs, miRNAs in chemotherapy-induced senescent cells and their secreted extracellular vehicles (EVs) were detected and the miRNA profiles were analyzed, hoping to provide some thoughts for the evaluation of chemotherapy effect in clinical tumor treatment.MethodsNeuroblastoma cell line IMR32 was treated with low concentration of aurora kinase A (AURKA) inhibitor MLN8237 to establish a cell senescence model. RNAs were extracted from senescent cells and extracellular vesicles, and miRNA spectrum was investigated by small RNA deep sequencing.ResultsThirteen miRNAs including miR-378b and miR-206 were significantly increased, whereas 32 miRNAs including miR-205-5p, miR-378d, and miR-378f were significantly decreased in the senescence cell group. In senescent cells secreted extracellular vesicles, there were 48 of up-regulated miRNAs including miR-205-5p and 9 of down-regulated miRNAs. Bioinformatics analysis revealed that these differentially expressed miRNAs in senescent cells may potentially regulate many common target genes which belong to the metabolic signaling pathway and transcriptional misregulation in cancer.ConclusionsThe expression profiles of miRNA in senescent neuroblastoma cells and extracellular vesicles were altered, and the differentially expressed miRNAs were mostly involved in cellular metabolic pathways. The information decryption can provide reference for clinical interpretation of the phenomenon of therapy-induced senescence.

Identification of aberrantly methylated differentially expressed genes and pro-tumorigenic role of KIF2C in melanoma.

Background: Skin Cutaneous Melanoma (SKCM) is known as an aggressive malignant cancer, which could be directly derived from melanocytic nevi. However, the molecular mechanisms underlying the malignant transformation of melanocytes and melanoma tumor progression still remain unclear. Increasing research showed significant roles of epigenetic modifications, especially DNA methylation, in melanoma. This study focused on the identification and analysis of methylation-regulated differentially expressed genes (MeDEGs) between melanocytic nevus and malignant melanoma in genome-wide profiles. Methods: The gene expression profiling datasets (GSE3189 and GSE114445) and gene methylation profiling datasets (GSE86355 and GSE120878) were downloaded from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) and differentially methylated genes (DMGs) were identified via GEO2R. MeDEGs were obtained by integrating the DEGs and DMGs. Then, a functional enrichment analysis of MeDEGs was performed. STRING and Cytoscape were used to describe the protein-protein interaction (PPI) network. Furthermore, survival analysis was implemented to select the prognostic hub genes. Next, we conducted gene set enrichment analysis (GSEA) of hub genes. To validate, SKCM cell culture and lentivirus infection was performed to reveal the expression and behavior pattern of KIF2C. Patients and specimens were collected and then immunohistochemistry (IHC) staining was conducted. Results: We identified 237 hypomethylated, upregulated genes and 182 hypermethylated, downregulated genes. Hypomethylation-upregulated genes were enriched in biological processes of the oxidation-reduction process, cell proliferation, cell division, phosphorylation, extracellular matrix disassembly and protein sumoylation. Pathway enrichment showed selenocompound metabolism, small cell lung cancer and lysosome. Hypermethylation-downregulated genes were enriched in biological processes of positive regulation of transcription from RNA polymerase II promoter, cell adhesion, cell proliferation, positive regulation of transcription, DNA-templated and angiogenesis. The most significantly enriched pathways involved the transcriptional misregulation in cancer, circadian rhythm, tight junction, protein digestion and absorption and Hippo signaling pathway. After PPI establishment and survival analysis, seven prognostic hub genes were CKS2, DTL, KIF2C, KPNA2, MYBL2, TPX2, and FBL. Moreover, the most involved hallmarks obtained by GSEA were E2F targets, G2M checkpoint and mitotic spindle. Importantly, among the 7 hub genes, we found that down-regulated level of KIF2C expression significantly inhibited the proliferative ability of SKCM cells and suppressed the metastasis capacity of SKCM cells. Conclusions: Our study identified potential aberrantly methylated-differentially expressed genes participating in the process of malignant transformation from nevus to melanoma tissues based on comprehensive genomic profiles. Transcription profiles of CKS2, DTL, KIF2C, KPNA2, MYBL2, TPX2, and FBL provided clues of aberrantly methylation-based biomarkers, which might improve the development of precision medicine. KIF2C plays a pro-tumorigenic role and potentially inhibited the proliferative ability in SKCM.

Bioinformatics Analysis Reveals the Biomarker Value and Potential Mechanism of miR-675-3p in Gastric Cancer.

Background Gastric cancer (GC) is still the main challenge for the social and clinical system. Increasing studies have proved that microRNA dysfunction is closely associated with the GC progression. miR-675-3p has been confirmed as the tumor support in multiple tumor cells, while its role in GC remains unclear. Methods. The clinical data in the TCGA database were excavated for analyzing the role of miR-675-3p in pan-cancer and GC. qRT-PCR was applied to detect the abundances of the genes. The Starbase 2.0 was executed to target the prediction of miR-675-3p. Moreover, the enrichment analysis was performed with the DAVID database. The PPI-network analysis of the targets was performed with Cytoscape. Results miR-675-3p was dramatically upregulated in multiple types of cancer, and elevated miR-675-3p was also found in GC tissues. Moreover, increased miR-675-3p was closely related with the poor survival rates of the patients. The qRT-PCR showed that miR-675-3p was extremely upregulated in GC tissues and cell lines. The enrichment analysis showed that the targets of miR-675-3p were located in the cellular nucleus and associated with the transcriptional misregulation in cancer. The PPI-network showed that three clusters and total of 40 genes were screened as potential hub nodes. Moreover, BRIP1, MYO5B, and PDS5B were related with the prognostic survival of the patients according to the TCGA database and decreased BRIP1, MYO5B, and PDS5B were also found in GC cell lines. Conclusion This study identified miR-675-3p as a potential biomarker in GC development and revealed the potential regulation network of miR-675-3p.

Orelabrutinib Combined With Lenalidomide and Immunochemotherapy for Relapsed/Refractory Primary Central Nervous System Lymphoma: A Retrospective Analysis of Case Series.

BackgroundRelapsed/refractory (r/r) primary central nervous system lymphoma (PCNSL) is an intractable situation without sound treatment. Bruton’s tyrosine kinase (BTK) represents an attractive drug target in PCNSL. Orelabrutinib is a new-generation BTK inhibitor with high cerebrospinal fluid (CSF) concentration. This study aimed to evaluate the efficacy and safety of orelabrutinib-containing combination therapy in patients with r/r PCNSL.MethodsWe retrospectively analyzed r/r PCNSL patients who received combination therapy with rituximab, high-dose methotrexate, temozolomide, orelabrutinib and lenalidomide, and further explored the relationship between the efficacy and genetic characteristics.ResultsA total of fifteen patients were included in this retrospective study. The overall response rate (ORR) was 86.7%, the complete remission (CR) rate was 73.3% and the disease control rate (DCR) was 93.3%. Among 13 responders, 9 patients are still receiving oral orelabrutinib and lenalidomide. The most common adverse event (AEs) was transaminase increase (66.7%). No grade 4 AE or drug-related death was reported. Genomic sequencing showed that patients who responded to orelabrutinib had abnormal NF-κB activation, while those who had no response were mainly enriched with transcriptional misregulation. Patients who had mutations in TLR, BCR, or NF-κB pathway achieved complete or partial response to the orelabrutinib-containing therapy. Moreover, the blood and cerebrospinal fluid circulating tumor DNA (ctDNA) were closely associated with tumor recurrence and treatment response and sustained tumor responses correlated with the clearance of ctDNA.ConclusionOrelabrutinib-containing regimen was effective and well-tolerated in patients with r/r PCNSL. Genome sequencing of tumor samples could help to screen patients who may respond to the orelabrutinib-containing regimen, and liquid biopsy may contribute to tracing tumor burden and monitoring treatment response.

Identification of HOX signatures contributing to oral cancer phenotype

The role of evolutionarily conserved homeobox-containing HOX genes as transcriptional regulators in the developmental specification of organisms is well known. The contribution of HOX genes involvement in oral cancer phenotype has yet to be fully ascertained. TCGA-HNSC HTSeq-counts and clinical data were retrieved from the GDC portal for oral cavity neoplasms. GEO datasets (GSE72627, GSE30784, GSE37991) were accessed and analyzed using GEO2R. Differential HOX gene expression was profiled using the DESeq2 R package with a log2 fold change cut-off (− 1 and + 1) and Benjamini–Hochberg p-adjusted value at ≤ 0.01. Gene set over-representation analysis and semantic analysis associated with the disease ontology was performed using the ClusterProfiler R package, and pathway over-representation analysis was performed using IMPaLa. HOX protein interaction network was constructed using the Pathfind R package. HOX phenotype associations were performed using Mammalian Phenotype Ontology, Human Phenotype Ontology, PhenGenI associations, Jensen tissues, and OMIM entries. Drug connectivity mapping was carried out with Dr. Insight R package. HOXA2 was upregulated in oral dysplasia but silenced during tumor progression. Loss of HOXB2 expression was consistent in the potentially malignant oral lesions as well as in the primary tumor. HOXA7, HOXA10, HOXB7, HOXC6, HOXC10, HOXD10, and HOXD11 were consistently upregulated from premalignancy to malignancy and were notably associated with risk factors. Overrepresentation analysis suggested HOXA10 was involved in the transcriptional misregulation contributing to the oral cancer phenotype. HOX genes subnetwork analysis showed crucial interactions with cell cycle regulators, growth responsive elements, and proto-oncogenes. Phenotype associations specific to the oral region involving HOX genes provide intrinsic cues to tumor development. The 5′ HOX genes were aberrantly upregulated during oral carcinogenesis reflecting their posterior prevalence.

Abstract LB060: Single-cell CRISPR screen of malignant peripheral nerve sheath tumors (MPNST)

Abstract Background: Malignant peripheral nerve sheath tumors (MPNST) are aggressive sarcomas of the peripheral nervous system. Loss of function of the polycomb repressive complex 2 (PRC2) have been identified in up to 80% of MPNST cases, via recurrent genetic mutations in the core components, EED and/or SUZ12. However, the transcriptional mis-regulation incurred by the loss of transcriptional repression in MPNST, through loss of H3K27me3, has yet to be fully deciphered. We hypothesize that PRC2 loss leads to transcriptional regulatory imbalance between key epigenetic regulating complexes in this malignancy, which may contribute to the oncogenesis of this disease. Therefore, this project aims to elucidate the role of the SWI/SNF chromatin remodeling complexes in MPNST, epigenetic regulators known to act antagonistically to PRC2. Method: This project uses a CRISPR knock out screen combined with a single cell RNA sequencing readout to target 44 known components of the SWI/SNF complexes. This technology enables large scale phenotypic assays, which we leverage to decipher the complicated epigenetic regulatory network within these lesions. Resulting genes of interest were further studied via loss-of-function assays coupled with colony formation assay and chromatin immunoprecipitation DNA sequencing (ChIP-seq) experiments. Results: This CRISPR screen highlighted core SWI/SNF components, SMARCA4 and SMARCE1, as genes of interest in MPNST. Knockdown of these genes in MPNST cell lines reduced proliferation, indicating a role for these genes in MPNST growth and viability. Further, investigations via soft agar assay found both SMARCA4 and SMARCE1 knock out to decrease the tumorigenic potential of MPNST cells. The potential dependency of SMARCA4 function in MPNST cells on PRC2 mutation was investigated using a doxycycline-inducible PRC2-restoration MPNST cell line model. ChIP-seq results suggest that loss of PRC2 does not substantially affect the binding of SMARCA4-containing SWI/SNF complexes across the genome. DPF1, a member of the neuronal-specific SWI/SNF chromatin remodeling complex, was further highlighted as a unique regulator of MPNST transcription via the CRISPR single-cell screen. CRISPR knockdown of this gene in MPNST cell lines has been found to decrease proliferation, while soft agar assay experiments highlighted that DPF1 loss reduces cell tumorigenicity. The unique dependency of this gene observed in MPNST but not in any other cancer subtypes highlights the potential of DPF1 to be a novel and unique therapeutic target in MPNST. Conclusion: Thus far, our studies have shown that members of the SWI/SNF complex play key roles in the MPNST cell proliferation and tumorigenicity. The function of these complexes in MPNST, however, may be independent of the transcriptional imbalance caused by PRC2 mutation. Citation Format: Béga Murray, Xiyuan Zhang, Shahroze Abbas, Hilda Jafarah, Jack F. Shern. Single-cell CRISPR screen of malignant peripheral nerve sheath tumors (MPNST) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB060.

Abstract 2953: Subtype-specific and structure variation induced 3D genome alteration in acute myeloid leukemia

Abstract Acute myeloid leukemia (AML) represents a set of heterogeneous myeloid malignancies hallmarked by mutations in epigenetic modifiers, transcription factors, and kinases that can cause epigenetic reshaping. It is unclear to what extent AML mutations drive chromatin 3D structure alteration and contribute to myeloid transformation. We first performed Hi-C and whole-genome sequencing in 25 AML patient samples and seven healthy donor samples, and identified recurrent alterations of A/B compartments, TADs, and chromatin loops that are unique to different subtypes. To investigate how altered chromatin organization contributes to transcriptional misregulation, we performed RNA-Seq, ATAC-Seq and CUT&amp;ag for CTCF, H3K27ac, and H3K27me3 in the same AML samples. We identified extensive and recurrent AML-specific promoter-enhancer and promoter-repressor loops. We performed both CRISPR deletion and interference experiments and validated two repressor loops that downregulated cancer related genes IKZF2 and RTTN. Furthermore, by using our recently developed algorithm, we identified structural variation-induced enhancer-hijacking and repressor-hijacking events in AML samples. We further demonstrated the role of hijacked enhancers in AML cell growth by CRISPR screening, and the role of hijacked repressors by CRISPR de-repression. We performed whole-genome bisulfite sequencing in 20 AML and normal samples, and showed the delicate relationship between DNA methylation, CTCF binding and 3D genome structure. Finally, by treating the AML cells with the DNA hypomethylating agent and performing triple knockdown of DNMT1/3A/3B, we demonstrated the impact of altered DNA methylation on gene expression and 3D genome organization. Overall this study provides an invaluable resource for leukemia studies and also highlighted the role of repressor-loops and hijacked cis-elements in gene regulation and human diseases. Citation Format: Jie Xu, Fan Song, Baozhen Zhang, Huijue Lyu, Mikoto Kobayashi, Ziyu Zhao, Ye Hou, Xiaotao Wang, Yu Luan, Bei Jia, Lena Stasiak, Qixuan Wang, Qi Jin, Qiushi Jin, Yihao Fu, Ross C. Hardison, Sinisa Dovat, Leonidas C. Platanias, Yue Yang, Tomoko Yamada, Aaron D. Viny, Ross L. Levine, David F. Claxton, James R. Broach, Hong Zheng, Feng Yue. Subtype-specific and structure variation induced 3D genome alteration in acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2953.

Identifying the Potential Roles of PBX4 in Human Cancers Based on Integrative Analysis.

PBX4 belongs to the pre-B-cell leukemia homeobox (PBX) transcription factors family and acts as a transcriptional cofactor of HOX proteins participating in several pathophysiological processes. Recent studies have revealed that the dysregulation of PBX4 is closely related to multiple diseases, especially cancers. However, the research on PBX4’s potential roles in 33 cancers from the Cancer Genome Atlas (TCGA) is still insufficient. Therefore, we performed a comprehensive pan-cancer analysis to explore the roles of PBX4with multiple public databases. Our results showed that PBX4 was differentially expressed in 17 types of human cancer and significantly correlated to the pathological stage, tumor grade, and immune and molecular subtypes. We used the Kaplan–Meier plotter and PrognoScan databases to find the significant associations between PBX4 expression and prognostic values of multiple cancers. It was also found that PBX4 expression was statistically related to mutation status, DNA methylation, immune infiltration, drug sensitivity, and immune checkpoint blockade (ICB) therapy. Additionally, we found that PBX4 was involved in different functional states of multiple cancers from the single-cell resolution perspective. Enrichment analysis results showed that PBX4-related genes were enriched in the cell cycle process, MAPK cascade, ncRNA metabolic process, positive regulation of GTPase activity, and regulation of lipase activity and mainly participated in the pathways of cholesterol metabolism, base excision repair, herpes simplex virus 1 infection, transcriptional misregulation in cancer, and Epstein–Barr virus infection. Altogether, our integrative analysis could help in better understanding the potential roles of PBX4 in different human cancers.

TMT-Based Quantitative Proteomic Analysis of Intestinal Organoids Infected by Listeria monocytogenes Strains with Different Virulence.

L. monocytogenes, consisting of 13 serotypes, is an opportunistic food-borne pathogen that causes different host reactions depending on its serotypes. In this study, highly toxic L. monocytogenes 10403s resulted in more severe infections and lower survival rates. Additionally, to investigate the remodeling of the host proteome by strains exhibiting differential toxicity, the cellular protein responses of intestinal organoids were analyzed using tandem mass tag (TMT) labeling and high-performance liquid chromatography–mass spectrometry. The virulent strain 10403s caused 102 up-regulated and 52 down-regulated proteins, while the low virulent strain M7 caused 188 up-regulated and 25 down-regulated proteins. Based on the analysis of gene ontology (GO) and KEGG databases, the expressions of differential proteins in organoids infected by L. monocytogenes 10403s (virulent strain) or M7 (low virulent strain) were involved in regulating essential processes such as the biological metabolism, the energy metabolism, and immune system processes. The results showed that the immune system process, as the primary host defense response to L. monocytogenes, comprised five pathways, including ECM–receptor interaction, the complement and coagulation cascades, HIF-1, ferroptosis, and NOD-like receptor signaling pathways. As for the L. monocytogenes 10403s vs. M7 group, the expression of differential proteins was involved in two pathways: systemic lupus erythematosus and transcriptional mis-regulation in cancer. All in all, these results revealed that L. monocytogenes strains with different toxicity induced similar biological functions and immune responses while having different regulations on differential proteins in the pathway.

Bioinformatics Analysis of Core Genes and Key Pathways in Myelodysplastic Syndrome

To screen differentially expressed gene (DEG) related to myelodysplastic syndrome (MDS) based on Gene Expression Omnibus (GEO) database, and explore the core genes and pathogenesis of MDS by analyzing the biological functions and related signaling pathways of DEG. The expression profiles of GSE4619, GSE19429, GSE58831 including MDS patients and normal controls were downloaded from GEO database. The gene expression analysis tool (GEO2R) of GEO database was used to screen DEG according to | log FC (fold change) |≥1 and P<0.01. David online database was used to annotate gene ontology function (GO). Metascape online database was used to enrich and analyze differential genes in Kyoto Encyclopedia of Genes and Genomes (KEGG). The protein-protein interaction network (PPI) was constructed by using STRING database. CytoHubba and Mcode plug-ins of Cytoscape were used to analyze the key gene clusters and hub genes. R language was used to diagnose hub genes and draw the ROC curve. GSEA enrichment analysis was performed on GSE19429 according to the expression of LEF1. A total of 74 co-DEG were identified, including 14 up-regulated genes and 60 down regulated genes. GO enrichment analysis indicated that BP of down regulated genes was mainly enriched in the transcription and regulation of RNA polymerase II promoter, negative regulation of cell proliferation, and immune response. CC of down regulated genes was mainly enriched in the nucleus, transcription factor complexes, and adhesion spots. MF was mainly enriched in protein binding, DNA binding, and β-catenin binding. KEGG pathway was enriched in primary immunodeficiency, Hippo signaling pathway, cAMP signaling pathway, transcriptional mis-regulation in cancer and hematopoietic cell lineage. BP of up-regulated genes was mainly enriched in type I interferon signaling pathway and viral response. CC was mainly enriched in cytoplasm. MF was mainly enriched in RNA binding. Ten hub genes and three important gene clusters were screened by STRING database and Cytoscape software. The functions of the three key gene clusters were closely related to immune regulation. ROC analysis showed that the hub genes had a good diagnostic significance for MDS. GSEA analysis indicated that LEF1 may affect the normal function of hematopoietic stem cells by regulating inflammatory reaction, which further revealed the pathogenesis of MDS. Bioinformatics can effectively screen the core genes and key signaling pathways of MDS, which provides a new strategy for the diagnosis and treatment of MDS.

Transcriptomic profiling of nonneoplastic cortical tissues reveals epileptogenic mechanisms in dysembryoplastic neuroepithelial tumors.

Low-grade dysembryoplastic neuroepithelial tumors (DNTs) are a frequent cause of drug-refractory epilepsy. Molecular mechanisms underlying seizure generation in these tumors are poorly understood. This study was conducted to identify altered genes in nonneoplastic epileptogenic cortical tissues (ECTs) resected from DNT patients during electrocorticography (ECoG)-guided surgery. RNA sequencing (RNAseq) was used to determine the differentially expressed genes (DEGs) in these high-spiking ECTs compared to non-epileptic controls. A total of 477 DEGs (180 upregulated; 297 downregulated) were observed in the ECTs compared to non-epileptic controls. Gene ontology analysis revealed enrichment of genes belonging to the following Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways: (i) glutamatergic synapse; (ii) nitrogen metabolism; (iii) transcriptional misregulation in cancer; and (iv) protein digestion and absorption. The glutamatergic synapse pathway was enriched by DEGs such as GRM4, SLC1A6, GRIN2C, GRM2, GRM5, GRIN3A, and GRIN2B. Enhanced glutamatergic activity was observed in the pyramidal neurons of ECTs, which could be attributed to altered synaptic transmission in these tissues compared to non-epileptic controls. Besides glutamatergic synapse, altered expression of other genes such as GABRB1 (synapse formation), SLIT2 (axonal growth), and PROKR2 (neuron migration) could be linked to epileptogenesis in ECTs. Also, upregulation of GABRA6 gene in ECTs could underlie benzodiazepine resistance in these patients. Neural cell-type-specific gene set enrichment analysis (GSEA) revealed transcriptome of ECTs to be predominantly contributed by microglia and neurons. This study provides first comprehensive gene expression profiling of nonneoplastic ECTs of DNT patients and identifies genes/pathways potentially linked to epileptogenesis.

Diagnosis and Prediction of Endometrial Carcinoma Using Machine Learning and Artificial Neural Networks Based on Public Databases.

Endometrial carcinoma (EC), a common female reproductive system malignant tumor, affects thousands of people with high morbidity and mortality worldwide. This study was aimed at developing a prediction model for the diagnosis of EC in the general population. First, we obtained datasets GSE63678, GSE106191, and GSE115810 from the Gene Expression Omnibus (GEO) database, dataset GSE17025 from the GEO database, and the RNA sequence of EC from The Cancer Genome Atlas (TCGA) database to constitute the training, test, and validation groups, respectively. Subsequently, the 96 most significantly differentially expressed genes (DEGs) were identified and analyzed for function and pathway enrichment in the training group. Next, we acquired the disease-specific genes by random forest and established an artificial neural network for the diagnosis. Receiver operating characteristic (ROC) curves were utilized to identify the signature across the three groups. Finally, immune infiltration was analyzed to reveal tumor-immune microenvironment (TIME) alterations in EC. The top 96 DEGs (77 down-regulated and 19 up-regulated genes) were primarily enriched in the interleukin-17 signaling pathway, protein digestion and absorption, and transcriptional misregulation in cancer. Subsequently, 14 characterizing genes of EC were identified by random forest. In the training, test, and validation groups, the artificial neural network was constructed with high diagnostic accuracies of 0.882, 0.864, and 0.839, respectively, and areas under the ROC curve (AUCs) of 0.928, 0.921, and 0.782, respectively. Finally, resting and activated mast cells were found to have increased in TIME. We constructed an artificial diagnostic model with excellent reliability for EC and uncovered variations in the immunological ecosystem of EC through integrated bioinformatics approaches, which might be potential diagnostic targets for EC.

Gene Expression Profile Analysis of Human Epidermal Keratinocytes Expressing Human Papillomavirus Type 8 E7.

Background: Human papillomavirus type 8 (HPV8) has been implicated in the progress of non-melanoma skin cancers and their precursor lesions. The HPV8 E7 oncoprotein plays a key role in the tumorigenesis of HPV-associated cutaneous tumors. However, the exact role of HPV8 E7 in human epidermal carcinogenesis has not been fully elucidated. Methods: To investigate the potential carcinogenic effects of HPV8 E7 on epithelial cells, we used RNA-sequencing technology to analyze the gene expression profile of HPV8 E7-overexpressed normal human epidermal keratinocytes (NHEKs). Results: RNA-sequencing revealed 831 differentially expressed genes (DEGs) between HPV8 E7-expressing NHEKs and control cells, among which, 631 genes were significantly upregulated, and 200 were downregulated. Gene ontology annotation enrichment analysis showed that HPV8 E7 mainly affected the expression of genes associated with protein heterodimerization activity, DNA binding, nucleosomes, and nucleosome assembly. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis revealed that overexpression of HPV8 E7 affected the expression of gene clusters associated with viral carcinogenesis and transcriptional misregulation in cancer and necroptosis signaling pathways that reportedly play crucial roles in HPV infection promotion and cancer progression. We also found the DEGs, such as HKDC1 and TNFAIP3, were associated with epigenetic modifications, immune regulation, and metabolic pathways. Conclusion: Our results demonstrate that the pro-carcinogenic effect of HPV8 expression in epithelial cells may be attributed to the regulatory effect of oncogene E7 on gene expression associated with epigenetic modifications and immune and metabolic status-associated gene expression. Although our data are based on an in vitro experiment, it provides the theoretical evidence that the development of squamous cell carcinoma can be caused by HPV.

Development of a Risk Score Model for Osteosarcoma Based on DNA Methylation-Driven Differentially Expressed Genes.

Osteosarcoma (OS) is the commonest malignant bone tumor in adolescent patients, and patients face amputation, tumor metastasis, chemotherapy resistance, and even death. We investigated the potential connection between abnormal methylation differentially expressed genes and the survival rate of osteosarcoma patients. GSE36002 and GSE12865 datasets of GEO database were utilized for abnormal methylation differentially expressed genes, followed by function and pathway enrichment analyses, the protein-protein interaction network in the STRING database, and cluster analysis in the MCODE app of Cytoscape. The RNA-seq and clinical data from the TARGET-OS project of TCGA were used for univariate and least absolute shrinkage and selection operator (LASSO) Cox regression analyses to predict the risk genes of osteosarcoma. 1191 hypermethylation-downregulated genes might function through plasma membrane, negative regulation of transcription from the RNA polymerase II promoter, and pathways, including transcriptional misregulation in cancer. 127 hypomethylation-upregulated genes were enriched in proteolysis, negative regulation of the canonical Wnt signaling pathway, and metabolic signaling pathways. The univariate Cox analysis revealed 638 genes (P < 0.01), including 50 hypermethylation-downregulated genes and 4 hypomethylation-upregulated genes, subsequently based on LASSO Cox regression analysis for 54 aberrant methylation-driven genes, and three genes (COL13A1, MXI1, and TBRG1) were selected to construct the risk score model. The three genes (COL13A1, MXI1, and TBRG1) regulated by DNA methylation were identified to relate with the outcomes of OS patients, which might provide a new insight to the pathological mechanism of osteosarcoma.

Impact of Environmentally Relevant Concentrations of Bisphenol A (BPA) on the Gene Expression Profile in an In Vitro Model of the Normal Human Ovary.

Endocrine-disrupting chemicals (EDCs), including the xenoestrogen Bisphenol A (BPA), can interfere with hormonal signalling. Despite increasing reports of adverse health effects associated with exposure to EDCs, there are limited data on the effect of BPA in normal human ovaries. In this paper, we present a detailed analysis of the transcriptomic landscape in normal Human Epithelial Ovarian Cells (HOSEpiC) treated with BPA (10 and 100 nM). Gene expression profiles were determined using high-throughput RNA sequencing, followed by functional analyses using bioinformatics tools. In total, 272 and 454 differentially expressed genes (DEGs) were identified in 10 and 100 nM BPA-treated HOSEpiCs, respectively, compared to untreated controls. Biological pathways included mRNA surveillance pathways, oocyte meiosis, cellular senescence, and transcriptional misregulation in cancer. BPA exposure has a considerable impact on 10 genes: ANAPC2, AURKA, CDK1, CCNA2, CCNB1, PLK1, BUB1, KIF22, PDE3B, and CCNB3, which are also associated with progesterone-mediated oocyte maturation pathways. Future studies should further explore the effects of BPA and its metabolites in the ovaries in health and disease, making use of validated in vitro and in vivo models to generate data that will address existing knowledge gaps in basic biology, hazard characterisation, and risk assessment associated with the use of xenoestrogens such as BPA.