Genome Gene Expression Profiling Research Articles

ZEB family is a prognostic biomarker and correlates with anoikis and immune infiltration in kidney renal clear cell carcinoma

BackgroundZinc finger E-box binding homEeobox 1 (ZEB1) and ZEB2 are two anoikis-related transcription factors. The mRNA expressions of these two genes are significantly increased in kidney renal clear cell carcinoma (KIRC), which are associated with poor survival. Meanwhile, the mechanisms and clinical significance of ZEB1 and ZEB2 upregulation in KIRC remain unknown.MethodsThrough the Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database, expression profiles, prognostic value and receiver operating characteristic curves (ROCs) of ZEB1 and ZEB2 were evaluated. The correlations of ZEB1 and ZEB2 with anoikis were further assessed in TCGA-KIRC database. Next, miRTarBase, miRDB, and TargetScan were used to predict microRNAs targeting ZEB1 and ZEB2, and TCGA-KIRC database was utilized to discern differences in microRNAs and establish the association between microRNAs and ZEBs. TCGA, TIMER, TISIDB, and TISCH were used to analyze tumor immune infiltration.ResultsIt was found that ZEB1 and ZEB2 expression were related with histologic grade in KIRC patient. Kaplan-Meier survival analyses showed that KIRC patients with low ZEB1 or ZEB2 levels had a significantly lower survival rate. Meanwhile, ZEB1 and ZEB2 are closely related to anoikis and are regulated by microRNAs. We constructed a risk model using univariate Cox and LASSO regression analyses to identify two microRNAs (hsa-miR-130b-3p and hsa-miR-138-5p). Furthermore, ZEB1 and ZEB2 regulate immune cell invasion in KIRC tumor microenvironments.ConclusionsAnoikis, cytotoxic immune cell infiltration, and patient survival outcomes were correlated with ZEB1 and ZEB2 mRNA upregulation in KIRC. ZEB1 and ZEB2 are regulated by microRNAs.

Abstract NG02: Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers

Abstract NG02: Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers

Genomic and global gene expression profiling in pediatric and young adult acute leukemia with PICALM::MLLT10 Fusion.

MLLT10 fusion is a rare but recurrent genetic driver in acute leukemias. To better understand the genomic landscape of PICALM::MLLT10 (PM) positive acute leukemia, we performed genomic profiling and gene expression profiling in twenty PM-positive patients, including AML (n = 10), T-ALL/LLy (n = 8), Mixed-phenotype acute leukemia (MPAL), T/B (n = 1) and acute undifferentiated leukemia (AUL) (n = 1). Besides confirming the known activation of HOXA, differential gene expression analysis compared to hematopoietic stem cells demonstrated the enrichment of genes associated with cell proliferation-related pathways and relatively high expression of XPO1 in PM-AML and PM-T-ALL/LLy. Our study also suggested PHF6 disruption as a key cooperating event in PICALM::MLLT10-positive leukemias. In addition, we demonstrated differences in gene expression profiles as well as remarkably different spectra of co-occurring mutations between PM-AML and PM-T-ALL/LLy. Alterations affecting TP53 and NF1, hallmarks of PM-AML, are strongly associated with disease progression and relapse, whereas EZH2 alterations are highly enriched in PM-T-ALL/LLy. This comprehensive genomic and transcriptomic profiling provides insights into the pathogenesis and development of PICALM::MLLT10 positive acute leukemia.

Validation of serum cystatin SN detection for diagnosis and poor prognosis of esophageal squamous cell carcinoma.

The identification of effective tumor markers is of paramount importance for the early diagnosis, treatment, and prognosis of esophageal squamous cell carcinoma (ESCC). The present study endeavors to identify efficacious serological markers that can differentiate patients with early-stage ESCC from those with benign esophageal lesions and healthy controls (HC). Cystatin-SN (CST1), an active cysteine protease inhibitor belonging to the Cystatin (CST) superfamily, is implicated in the pathogenesis of inflammation and tumorigenesis. The objective of this investigation is to assess the diagnostic, therapeutic, and prognostic potential of serum CST1 in ESCC. In our prior RNA sequencing and screening endeavors, we have identified ten genes that are up-regulated in relation to esophageal cancer. Subsequently, we have verified the gene CST1 from the transcriptome data of the The Cancer Genome Atlas Program (TCGA) and Gene Expression Profiling Interactive Analysis (GEPIA) database. Following this, we conducted an enzyme-linked immunosorbent assay (ELISA) to ascertain the expression levels of CST1 in serum samples from clinical cohorts. The study revealed a significant elevation in serum CST1 levels among patients with early-stage esophageal squamous cell carcinoma (ESCC) (7.41 ± 4.32 ng/ml) compared to those with esophageal benign lesions (4.67 ± 2.43 ng/ml) (p < 0.0001) and healthy controls (4.87 ± 2.77 ng/ml) (p < 0.0001). The diagnostic sensitivity of CST1 for ESCC was 75.68% (specificity 70.83%, AUC 0.775). Combination of CST1 and SCC-Ag exhibited the AUC up to 0.819. Additionally, serum CST1 levels exhibited a significant decrease at 1-2 weeks post-surgery (4.49 ± 3.31 ng/ml) compared to pre-surgery levels (7.68 ± 3.71 ng/ml) (p<0.0001). Survival analysis demonstrated a strong association between high (844/415-1543 d) or low (1490/645-1710 d) serum CST1 levels at diagnosis and overall survival time (p < 0.001). Furthermore, multivariate regression analysis confirmed CST1 (p=0.024, HR=2.023, 95%CI 1.099-3.725) as an independent prognostic factor. Serum CST1 has the potential to function as a diagnostic indicator for distinguishing early-stage esophageal squamous cell carcinoma (ESCC) from individuals with benign esophageal lesions and healthy individuals. Additionally, it could serve as a prognostic predictor and therapeutic efficacy indicator for patients with ESCC.

Abstract C055: Distinct epigenomic regulation of gene expression in Hispanic/Latino children with high-risk B-cell acute lymphoblastic leukemia

Abstract B-Cell Acute Lymphoblastic Leukemia (B-ALL) is the most common childhood malignancy. Hispanic/Latino (H/L) children have a 40% higher death rate than non-Hispanic white (NHW) children after correcting for socioeconomic factors. Recently, we identified a 3.5-fold increased incidence of deletion of one IKZF1 allele (IKZF1-del) and over 15-fold increased incidence of deletion of one IKZF1 allele concomitant with IGH-CRLF2 translocation (IKZF1-del/IGH-CRLF2) in B-ALL of H/L children. This provided a biological rationale for the worse prognosis of B-ALL in H/L children. The very high incidence (29% in all H/L children and 60% in ≥10yr old H/L children) makes IKZF1-del the most frequent genetic alteration that confers adverse prognosis in B-ALL in H/L children. The IKZF1 gene encodes a DNA-binding protein, IKAROS, which regulates transcription of its target genes via chromatin remodeling. The purpose of the study is to analyze the global epigenomic regulation of gene expression in H/L and NHW children with B-ALL and identify biological determinants of altered gene expression that result in worse prognosis for H/L children with B-ALL. /. B-ALL primary cells from 60 H/L and NHW children were analyzed with whole genome sequencing and gene expression profiling using RNA-seq. A subset of B-ALL from each group was analyzed for epigenomic changes by Reduced Representation Bisulfite Sequencing (RRBS), ChIP-seq, CUT&amp;RUN, and CUT&amp;Tag. Results showed that gene expression pattern in B-ALL of H/L children is quite different from B-ALL of NHW children. GSEA and Ingenuity pathway analysis identified oncogenic drivers of B-ALL in H/L children. B-ALL of H/L children have elevated expression of genes that promotestemness, inflammation, and with antiapoptotic activity. RBBS analysis showed a distinct DNA methylation pattern in B-ALL in H/L vs NHW children at promoters of a large set of genes that are involved in cellular signaling, RNA processing, and cellular proliferation. Presence of IKZF1-del results in the alteration of global epigenomic regulation of gene expression with altered enhancer and super-enhancer landscape. Epigenomic alterations due to IKZF1-del resulted in profound redistribution of enhancers and super-enhancers in B-ALL of both H/L vs. NHW children. This was associated with a distinct alteration in gene expression and uncovered that IKAROS regulates different sets of genes in B-ALL in H/L vs. NHW children. These data identified a unique oncogenic signaling that are responsible for aggressive course of B-ALL in H/L children. In conclusion, the multiomics analysis of primary B-ALL cells revealed a distinct global epigenomic regulation of gene expression between B-ALL from H/L and NHW children. This was demonstrated by alterations in DNA methylation and enhancer landscape. Presence of IKZF1-del has a different effect on gene expression in B-ALL of H//L vs NHW children. Results of the study provide a biological rationale for the worse prognosis of B-ALL in H/L children and identified novel therapeutic targets for this disease. Citation Format: Laura Budurlean, Bing He, Yali Ding, Diwakar Bastihalli Tukaramrao, Chingakham Singh, Joseph Schramm, Daniel Bogush, James Broach, Sinisa Dovat. Distinct epigenomic regulation of gene expression in Hispanic/Latino children with high-risk B-cell acute lymphoblastic leukemia [abstract]. In: Proceedings of the 16th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2023 Sep 29-Oct 2;Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2023;32(12 Suppl):Abstract nr C055.

ANXA3, associated with YAP1 regulation, participates in the proliferation and chemoresistance of cervical cancer cells.

Cervical cancer, as one of the most common cancers in women, remains a major health threat worldwide. Annexin A3 (ANXA3), a component of the annexin family, is upregulated in numerous cancers, with no explicit role in cervical cancer. This study aims to investigate the function of ANXA3 in cervical cancer. Differential expression genes between the cervical cancer tissues of patients and the controls were analyzed in The Cancer Genome Atlas (TCGA) and Gene Expression Profiling Interactive Analysis (GEPIA) database. Using transfection approaches to either upregulate or downregulate ANXA3, its role in cell proliferation and chemosensitivity of human cervical cancer cell lines (HeLa and C33A) was evaluated. Furthermore, the binding activity between YAP1 and ANXA3 was also explored. Genomics analysis indicated that differential genes were mostly associated with cell cycle progression and DNA replication. ANXA3 was highly expressed in the cervical cancer tissues and closely linked to malignancy degree. Knockdown of ANXA3 in cervical cancer cells inhibited cell cycle progression. A similar result was observed in the reduction of cyclin D, CDK4, cyclin E, and CDK2 in cervical cancer cells with ANXA3 silencing. Cervical cancer cells obtained high sensitivity to cisplatin (DDP) when ANXA3 was downregulated. Conversely, these capabilities were the opposite in cervical cancer cells overexpressing ANXA3. Furthermore, the expression levels of ANXA3 and YAP1 were positively correlated. YAP1 upregulation was positively connected with malignant behaviors, which were reversed by ANXA3 downregulation. In light of our findings, targeting ANXA3 expressed in cervical cancer might contribute to more potential therapeutic strategies.

Molecular subtypes from comprehensive clustering from multi-omics dataset to predict the therapeutic efficacy of immunotherapeutic agent-based treatments in advanced hepatocellular carcinoma.

4112 Background: In advanced hepatocellular carcinoma (HCC), combined treatment using anti-PD-L1 and anti-VEGF antibodies is a major therapeutic strategy in addition to using anti-PD-1 and anti-CTLA4 target agents. To identify clinically relevant molecular biomarkers for immunotherapeutic agents-based treatment in advanced HCC by comprehensive molecular profiling using genomics alterations and transcriptomic data. Methods: Multi-omics data from clinical tumor samples of patients treated with atezolizumab + bevacizumab (94 patients), nivolumab (40 patients), and ipilimumab + nivolumab (32 patients) were obtained from tumor biopsies or surgical resections in a single institution. The targeted panel sequencing was conducted via the Oncomine comprehensive assay (ThermoFisher, USA). QuantSeq mRNA sequencing (Lexogen, USA) was used to generate datasets for the genomic alterations and gene expression profiles. Results: A total of 166 target sequences for genomic alterations and 136 transcriptomic sequences were performed in patients with advanced HCC treated with immunotherapeutic agent-based treatment options. High tumor mutation burden was significantly associated with favorable therapeutic efficacy (disease control rate, P=0.036) and progression-free survival (P=0.037) in patients who had atezolizumab with bevacizumab treatment. The genomic alteration of CTNNB1 mutation correlated with the favorable therapeutic efficacy of nivolumab monotherapy (PFS, P=0.022), contrary to TP53 mutation (PFS, P=0.063) which demonstrated unfavorable outcomes. CD274(PD-L1) mRNA expression was significantly associated with a high drug response following nivolumab monotherapy (objective response rate, P=0.047) and oncologic outcomes (PFS, P=0.021). Interestingly, PLA2G4A, known as a downstream molecule of the VEGF signaling pathway, was highly correlated with unfavorable therapeutic efficacy of the combined atezolizumab with bevacizumab treatment (disease control rate, P=0.002) and related oncologic outcomes (PFS, P=0.003). We also identified the clinically relevant molecular subtypes from unbiased clustering using the ontologic pathway collection. We also used gene expression profiles to predict the therapeutic response and oncologic outcomes from immunotherapeutic agent-based treatment options. Conclusions: We identified clinically relevant molecular biomarkers from comprehensive clustering of a multi-omics dataset to predict the therapeutic efficacy and oncologic outcomes. Further prospective studies with clinical validation are warranted to confirm clinical implications.

Genomic landscape and gene expression profiles of feline oral squamous cell carcinoma.

Feline oral squamous cell carcinoma (FOSCC) is a cancer of the squamous cell lining in the oral cavity and represents up to 80% of all oral cancers in cats, with a poor prognosis. We have used whole exome sequencing (WES) and RNA sequencing of the tumor to discover somatic mutations and gene expression changes that may be associated with FOSCC occurrence. FOSCC offers a potential comparative model to study human head and neck squamous cell carcinoma (HNSCC) due to its similar spontaneous formation, and morphological and histological features. In this first study using WES to identify somatic mutations in feline cancer, we have identified tumor-associated gene mutations in six cats with FOSCC and found some overlap with identified recurrently mutated genes observed in HNSCC. Four samples each had mutations in TP53, a common mutation in all cancers, but each was unique. Mutations in other cellular growth control genes were also found such as KAT2B and ARID1A. Enrichment analysis of FOSCC gene expression profiles suggests a molecular similarity to human OSCC as well, including alterations in epithelial to mesenchymal transition and IL6/JAK/STAT pathways. In this preliminary study, we present exome and transcriptome results that further our understanding of FOSCC.

HP1α promotes the progression of prostate cancer.

Patients who have been diagnosed with prostate cancer (PCa) typically have a dismal outlook and few therapeutic choices available to them, because the precise pathogenesis of the disease is not yet fully understood. The presence of HP1α, also known as the heterochromatin protein 1α, is required for the creation of higher-order chromatin structures. However, little is known about HP1α that serves roles in the pathogenesis of PCa. The primary purpose of our research was to investigate alterations in the levels of HP1α expression and to plan a series of tests to validate the function of HP1α in PCa. Information on HP1α expression in PCa and benign prostatic hyperplasia (BPH) tissues were gathered using the Cancer Genome Atlas (TCGA) and Gene Expression Profiling Interactive Analysis (GEPIA) databases. RT-qPCR, western blotting, and immunohistochemistry (IHC) were used to assess HP1α mRNA and protein expression in several human PCa tissues and cell lines. The CCK8 assay, clone formation assay, and transwell assay were used to examine biological activities including cell proliferation, migration, and invasion. The expression of proteins connected to apoptosis and the epithelial-mesenchymal transition (EMT) was examined using Western blot. The tumorigenic effect of HP1α was also verified by in vivo experiments. HP1α expression was much higher in PCa than in BPH tissues and cells, and was positively correlated with the Gleason score of PCa. In vitro experiments showed that HP1α knockdown could inhibit the ability of proliferation, invasion, and migration of PC3 and LNCaP cells, and promote cell apoptosis and EMT. In vivo experiments showed that HP1α knockdown inhibited tumorigenesis in mice. Our findings indicate that HP1α expression promotes PCa development and might be a novel therapeutic target for the diagnosis or treatment of PCa.

PUS1 May Be a Potential Prognostic Biomarker and Therapeutic Target for Hepatocellular Carcinoma.

The mechanisms of pseudouridine synthase (PUS) are not definite in hepatocellular carcinoma (HCC), the objective of this study is to investigate the effect of PUS genes in HCC. Differentially expressed and prognostic gene of PUS enzymes was identified based on The Cancer Genome Atlas (TCGA), International Cancer Genome Consortium (ICGC) and Gene Expression Profiling Interactive Analysis (GEPIA) databases. For the identified gene, pseudouridine synthase 1 (PUS1), was used for further research. The clinicopathological feature of PUS1 was analyzed by Student's t-test. Prognostic significance was explored by Kaplan-Meier (KM) analysis and Cox proportional hazards regression model. Receiver operating characteristic (ROC) curve was applied to appraise diagnostic and prognostic value. The Database for Annotation, Visualization, and Integrated Discovery (DAVID) and Gene Set Enrichment Analysis (GSEA) were implemented to explore mechanism of PUS1. A Guangxi cohort was applied to verify differential expression. In vitro cell experiments were implemented to investigate the influence for proliferation, reactive oxygen species (ROS) level, migration, and invasion of HCC cells after a knockdown of PUS1. PUS1 was significantly overexpressed in HCC tissues, and patients with high PUS1 were related to unpromising clinicopathological features. Survival analysis revealed high PUS1 expression was associated with a poor overall survival (OS) and 1 year-recurrence free survival (RFS), was an independent risk factor. Meanwhile, ROC curve showed that PUS1 had a diagnostic and prognostic significance to HCC. Functional enrichment analysis implied that PUS1 may be involved in metabolic pathways, mitochondrial function, non-alcoholic fatty liver disease (NAFLD), and some important carcinogenic pathways. Cell assays revealed that knockdown of PUS1 significantly constrained the migration, proliferation, invasion and improved the ROS level of HCC cells. PUS1 may be a prognostic biomarker and a underlying treatment target for HCC.

RASA3 is a candidate gene in sickle cell disease-associated pulmonary hypertension and pulmonary arterial hypertension.

Pulmonary hypertension (PH)is associated with significant morbidity and mortality. RASA3 is a GTPase activatingprotein integral to angiogenesis and endothelial barrier function. In this study, we explore the association of RASA3 genetic variation with PH risk in patients with sickle cell disease (SCD)-associatedPH and pulmonary arterial hypertension (PAH).Cis-expression quantitative trait loci (eQTL)were queried for RASA3 using whole genome genotype arrays and gene expression profiles derived from peripheral blood mononuclear cells (PBMC)of three SCD cohorts. Genome-wide single nucleotide polymorphisms (SNPs)near or in the RASA3 gene that may associate with lung RASA3 expression were identified, reduced to 9 tagging SNPs for RASA3 and associated with markers of PH. Associations between the top RASA3 SNP and PAH severity were corroborated using data from the PAH Biobank and analyzed based on European or African ancestry (EA,AA).We found that PBMC RASA3 expression was lower in patients with SCD-associated PH as defined by echocardiography and right heart catheterization and was associated with higher mortality. One eQTL for RASA3 (rs9525228) was identified, with the risk allele correlating with PH risk, higher tricuspid regurgitant jet velocity and higher pulmonary vascular resistance in patients with SCD-associated PH. rs9525228 associated with markers of precapillary PH and decreased survival in individuals of EA but not AA. In conclusion, RASA3 is a novel candidate gene in SCD-associated PH and PAH, with RASA3 expression appearing to be protective. Further studies are ongoing to delineate the role of RASA3 in PH.

METTL3/YTHDF2 m6A axis promotes the malignant progression of bladder cancer by epigenetically suppressing RRAS.

The present study aimed to explore the potential roles of the methyltransferase‑like 3 (METTL3)‑mediated methylation of RAS related (RRAS) mRNA in the tumorigenesis and development of bladder cancer (BCa). For this purpose, the relative expression levels of METTL3 in BCa specimens and cell lines were measured using reverse transcription‑quantitative PCR (RT‑qPCR) and western blot analysis. The association between the METTL3 expression level and the clinical characteristics of patients with BCa was analyzed using The Cancer Genome Atlas (TCGA) and Gene Expression Profiling Interactive Analysis databases. Cellular experiments were performed to confirm the effects of METTL3 on the proliferative, migratory and invasive capacities of BCa cells. RT‑qPCR, western blot analysis, methylated RNA immunoprecipitation (MeRIP)‑qPCR and dual‑luciferase report assays were utilized to verify the METTL3/RRAS/YTH N6‑methyladenosine (m6A) RNA binding protein 2 (YTHDF2) regulatory axis in BCa. The results revealed that METTL3 expression was markedly increased in BCa specimens and cell lines, and was associated with poor clinical characteristics of patients with BCa. Invitro and invivo assays demonstrated that the silencing of METTL3 markedly suppressed the proliferative, migratory and invasive capacities of BCa cells. MeRIP‑PCR and dual‑luciferase report assays indicated that METTL3 could bind to the m6A sites of RRAS mRNA and suppress the transcriptional activity of RRAS. YTHDF2 could recognize the m6A sites of RRAS and mediate RRAS degradation. On the whole, the findings of the present study reveal the pivotal role of METTL3‑catalyzed m6A modification in BCa tumorigenesis and development. The change could facilitate BCa tumor growth and metastasis by suppressing RRAS expression in an m6A YTHDF2‑dependent manner. Targeting the METTL3/RRAS/YTHDF2 regulatory axis may thus prove to be a promising strategy for the diagnosis and therapy of BCa.

Bioinformatics-based construction of prognosis-related methylation prediction model for pancreatic cancer patients and its application value.

Objective: Pancreatic adenocarcinoma (PAAD) is a highly malignant gastrointestinal tumor with almost similar morbidity and mortality. In this study, based on bioinformatics, we investigated the role of gene methylation in PAAD, evaluated relevant factors affecting patient prognosis, screened potential anti-cancer small molecule drugs, and constructed a prediction model to assess the prognosis of PAAD. Methods: Clinical and genomic data of PAAD were collected from the Tumor Genome Atlas Project (TCGA) database and gene expression profiles were obtained from the GTEX database. Analysis of differentially methylated genes (DMGs) and significantly differentially expressed genes (DEGs) was performed on tumorous samples with KRAS wild-type and normal samples using the "limma" package and combined analysis. We selected factors significantly associated with survival from the significantly differentially methylated and expressed genes (DMEGs), and their fitting into a relatively streamlined prognostic model was validated separately from the internal training and test sets and the external ICGC database to show the robustness of the model. Results: In the TCGA database, 2,630 DMGs were identified, with the largest gap between DMGs in the gene body and TSS200 region. 318 DEGs were screened, and the enrichment analysis of DMGs and DEGs was taken to intersect DMEGs, showing that the DMEGs were mainly related to Olfactory transduction, natural killer cell mediated cytotoxicity pathway, and Cytokine -cytokine receptor interaction. DMEGs were able to distinguish well between PAAD and paraneoplastic tissues. Through techniques such as drug database and molecular docking, we screened a total of 10 potential oncogenic small molecule compounds, among which felbamate was the most likely target drug for PAAD. We constructed a risk model through combining three DMEGs (S100P, LY6D, and WFDC13) with clinical factors significantly associated with prognosis, and confirmed the model robustness using external and internal validation. Conclusion: The classification model based on DMEGs was able to accurately separate normal samples from tumor samples and find potential anti-PAAD drugs by performing gene-drug interactions on DrugBank.

MMP2 is a immunotherapy related biomarker and correlated with cancer-associated fibroblasts infiltrate in melanoma

BackgroundMounting evidence supports that matrix metalloproteinase (MMPs) are highly associated with tumor progression and that targeting MMPs may overcome the barrier of immune suppression. Among these, whether MMP2 functions as an immunosuppressive role in melanoma, remains unclear.MethodsThe Cancer Genome Atlas (TCGA) and Gene Expression Profiling Interactive Analysis 2 (GEPIA2) databases were used to assess the prognosis of MMP2 in melanoma, after which Tumor immune estimation resource (TIMER) was used to explore the relationship between MMP2 expression and cancer associated fibroblasts (CAFs) infiltration. Finally, we evaluated the efficacy of MMP2 inhibitor on CAFs infiltration and immunotherapy using a mouse melanoma model.ResultsIn general, the expression of MMP2, MMP13, MMP16, MMP17 and MMP25 were significantly associated with skin cutaneous melanoma (SKCM) patients prognosis, among which MMP2 low expression benefited patients the most. Especially, the overall survival (OS) of BRAF mutation patients with high MMP2 expression was significantly lower than the MMP2 low expression group, but there was no significant difference in BRAF wild-type patients. KEGG and GO enrichment analysis indicated that MMP2 related genes were mostly associated with extracellular structure organization, collagen-containing extracellular matrix and extracellular matrix structural constituent. Furthermore, in almost all cancers, MMP2 expression was positively correlated with CAFs infiltration. MMP2 inhibitor works synergistically with PD-1 antibody and induces tumor regression in a mouse melanoma model, which is dependent on decreased CAFs infiltration.ConclusionsThis suggests that MMP2 plays a vital role in the regulation of CAFs infiltration, potentially participating in immunotherapy response, and thus representing a valuable target of immunotherapy in melanoma.

Pan-genome and transcriptome analyses provide insights into genomic variation and differential gene expression profiles related to disease resistance and fatty acid biosynthesis in eastern black walnut (Juglans nigra).

Walnut (Juglans) species are used as nut crops worldwide. Eastern black walnut (EBW, Juglans nigra), a diploid, horticultural important woody species is native to much of eastern North America. Although it is highly valued for its wood and nut, there are few resources for understanding EBW genetics. Here, we present a high-quality genome assembly of J. nigra based on Illumina, Pacbio, and Hi-C technologies. The genome size was 540.8Mb, with a scaffold N50 size of 35.1Mb, and 99.0% of the assembly was anchored to 16 chromosomes. Using this genome as a reference, the resequencing of 74 accessions revealed the effective population size of J. nigra declined during the glacial maximum. A single whole-genome duplication event was identified in the J. nigra genome. Large syntenic blocks among J. nigra, Juglans regia, and Juglans microcarpa predominated, but inversions of more than 600kb were identified. By comparing the EBW genome with those of J. regia and J. microcarpa, we detected InDel sizes of 34.9Mb in J. regia and 18.3Mb in J. microcarpa, respectively. Transcriptomic analysis of differentially expressed genes identified five presumed NBS-LRR (NUCLEOTIDE BINDING SITE-LEUCINE-RICH REPEAT) genes were upregulated during the development of walnut husks and shells compared to developing embryos. We also identified candidate genes with essential roles in seed oil synthesis, including FAD (FATTY ACID DESATURASE) and OLE (OLEOSIN). Our work advances the understanding of fatty acid bioaccumulation and disease resistance in nut crops, and also provides an essential resource for conducting genomics-enabled breeding in walnut.

Single-minded homolog 2 as apotential prognostic signature and assessment ofits correlation with immune cell infiltration in pancreatic cancer.

Single-minded homolog 2 (SIM2) has been identified as apotential contributor to thedevelopment ofsolid tumors. Despite this, there is alack ofcomprehensive research regarding its biological role and underlying mechanism within pancreatic cancer (PC), as well as its prognostic impact. This study systematically evaluated theexpression level and clinical significance ofSIM2 in patients with PC using various databases, including TheCancer Genome Atlas, KM Plotter, and gene expression profiling interactive analysis. To investigate therelationship between SIM2 expression and immune cell infiltration, we conducted ESTIMATE and single-sample gene set enrichment analysis (ssGSEA) analyses. Single-minded homolog 2 was up-regulated in patients with PC. Pancreatic cancer patients with higher SIM2 expression had poorer overall survival rates. Gene set enrichment analysis results suggested that SIM2 may have asignificant impact on theprogression ofPC and theregulation ofimmune responses. According to thessGSEA algorithm, SIM2 has anegative correlation with thelevels ofinfiltrating TFH, mast cells, and pDC. Our study demonstrated that SIM2 serves as abiomarker, and is associated with both prognosis and immune infiltration in PC. This provides asolid foundation for future investigations into theprecise role ofSIM2 in thecarcinogenesis and progression ofPC.

Clinicopathological Significances and Prognostic Value of PPFIA4 in Colorectal Cancer.

Purpose: The PPFIA gene family (PPFIA1, PPFIA2, PPFIA3, and PPFIA4) is associated with multiple human diseases, particularly malignant tumors. However, the expression and prognostic value of the PPFIA family in human colorectal cancers (CRCs) have not been reported. Materials and methods: In this study, several databases, including Oncomine, UALCAN, and the cancer cell line encyclopedia, were used to compare differences in PPFIA1, PPFIA2, PPFIA3, and PPFIA4 expression between normal colon samples and CRCs. The expression levels of these four proteins were used to evaluate the survival of patients with CRC, as determined by the Cancer Genome Atlas Program (TCGA) portal and gene expression profiling interactive analysis (GEPIA) databases. Western blotting and reverse transcription-polymerase chain reaction were performed to detect protein and mRNA levels of PPFIA1, PPFIA3, and PPFIA4, respectively. Immunohistochemical (IHC) staining was used to detect the correlation between PPFIA4 expression and the degree of CRC malignancy. Furthermore, potential miRNAs targeting PPFIA4 in CRCs were studied and confirmed. Results: Bioinformatic analysis showed that the mRNA levels of PPFIA1, PPFIA3, and PPFIA4 were higher in CRC tissue samples than in normal colon tissue. Both mRNA and protein expression of PPFIA1, PPFIA3, and PPFIA4 were increased in the CRC cell lines LoVo and Hct116 compared with the normal colon epithelial cell line. Only PPFIA4 was associated with the prognosis of patients with CRC, which was confirmed by TCGA portal and GEPIA. IHC staining confirmed that the expression of PPFIA4 was higher in CRC tissues than in normal colon tissues and also increased in poorly differentiated CRC tissues and lymph node metastatic foci in comparison with well-differentiated CRC tissues and moderately differentiated CRC tissues. Functional annotation enrichment analysis indicated that the top 100 genes co-expressed with PPFIA4 were enriched in the G-protein coupled peptide receptor activity, leukotrience B4 receptor activity, and peroxisome proliferator-activated receptors and hypoxia-inducible factor-1 signaling pathways. In addition, miR-485-5p negatively regulates the expression of PPFIA4. Conclusion: PPFIA4 expression is associated with the development of CRCs and may be a novel potential prognostic marker for human CRCs.

Upregulated TUBG1 expression is correlated with poor prognosis in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) development is a complex pathological process. Tubulin gamma 1 (TUBG1) plays an oncogenic role in several human cancers; however, its functional role in HCC tumorigenesis remains unknown. Herein we first evaluated the gene expression levels of TUBG1 in HCC using data from The Cancer Genome Atlas and Gene Expression Profiling Interactive Analysis databases. We then elucidated the association between TUBG1 gene expression levels and survival rates of patients with HCC. Cell cycle, proliferation, transwell migration, and matrigel invasion assays were used to study the effects of TUBG1 on the malignant phenotypes of HCC cells. Based on the data obtained from the aforementioned databases and our in vitro experiments, TUBG1 was found to be overexpressed in HCC and patients with high TUBG1 expression levels showed a remarkably poor overall survival rate. In addition, the expression of TUBG1 significantly promoted the malignant phenotypes of HCC cells in vitro. Gene ontology term enrichment analysis revealed that co-regulated genes were enriched in biological processes mainly involved in chromosome segregation, chromosomal region, and chromatin binding; moreover, Kyoto Encyclopedia of Genes and Genome pathway analysis showed that they were mainly involved in cell cycle, oocyte meiosis, platinum drug resistance, and the p53 signaling pathway. We report that TUBG1 is an important oncogene in HCC. It promotes HCC progression and may serve as a potential prognostic biomarker for HCC. Future studies are warranted to unveil molecular biological mechanisms underlying TUBG1 carcinogenesis.

LncRNA surfactant associated 1 activates large tumor suppressor kinase 1/Yes-associated protein pathway via modulating hypoxic exosome-delivered miR-4766-5p to inhibit lung adenocarcinoma metastasis.

LncRNA surfactant associated 1 (SFTA1P) exhibits low expression in non-small cell lung cancer (NSCLC) tissues as compared with that in adjacent tissues, and may play a suppressing role in NSCLC. However, the effect and mechanism of SFTA1P on the metastasis of lung adenocarcinoma (LUAD) remain undefined, which are thus investigated in this research. Herein, potential impacts of SFTA1P on LUAD were determined through the Cancer Genome Atlas (TCGA) database and Gene Expression Profiling Interactive Analysis (GEPIA). After knockdown/overexpression of SFTA1P, the metastatic ability of LUAD cells was evaluated by molecular biology experiments (cell counting kit-8 assay, scratch test, Transwell assay and Western blot). The effect of SFTA1P on Yes-associated protein (YAP) nuclear translocation was assessed by Western blot. Hypoxia-induced exosomes were extracted for LUAD metastasis analysis. The targeting relationship of SFTA1P/miR-4766-5p/large tumor suppressor kinase 1 (LATS1) was verified by dual-luciferase reporter assay and molecular biology experiments. Xenograft and lung metastasis models were constructed for in vivo validation. SFTA1P was lowly expressed in LUAD, which was associated with the poor prognosis of patients with LUAD. Up-regulated SFTA1P prevented the metastasis of LUAD cells and the nuclear translocation of YAP. Hypoxia-induced exosomes stimulated LUAD cell metastasis, but inhibited the SFTA1P and LATS1/YAP axes. MiR-4766-5p acted as an intermediate "bridge" for SFTA1P to regulate LATS1. SFTA1P repressed xenograft growth and LUAD cell metastasis. To sum up, SFTA1P activates hypoxic exosome-delivered miR-4766-5p through modulating LATS1/YAP pathway, thereby suppressing LUAD cell metastasis, which may serve as a suitable target for the LUAD therapy.

Genome-Wide Methylation and Haplotype-Resolved Aberrant Somatic Hypermutation Patterns in B-Cell Lymphomas

Introduction Diffuse large B-cell lymphoma (DLBCL) is a genetically, phenotypically, and clinically heterogenous disease. Multiple molecular classification systems using either genomic aberrations or gene expression profiles are used to delineate subgroups with shared biology. The cell of origin (COO) gene expression-based classification system stratifies DLBCLs into germinal center B-cell-like (GCB) and activated B-cell-like subgroups of DLBCL. More recently, the double-hit signature (DHITsig) has been used to further stratify GCB DLBCLs into DHITsig+ and DHITsig-, where the former is characterized by a gene expression profile reminiscent of the GC dark zone and inferior outcomes. Currently, patterns of somatic driver mutations can only partially explain the expression of DHITsig in some tumors. Genetic features such as DNA methylation have been shown to be associated with transcriptional repression and altered chromatin structure. We hypothesized that resolving DNA methylation patterns genome-wide using long read whole genome sequencing (LR-WGS) may reveal epigenetic differences associated with differential gene expression profiles within the DHITsig+ and DHITsig- subgroups and in the context of histologic transformation from follicular lymphoma. Methods Whole genome sequencing data of 10 patient samples and 4 cell line GCB DLBCL samples were generated using both short read (SR-WGS; Illumina) and LR-WGS (Oxford Nanopore Technologies, PromethION). The complementary approach combines somatic variant calling from highly accurate SR data with DNA methylation calling from LR data. GCB samples were selected based on the NanoString DLBCL90 assay and further subdivided into DHITsig+ (N=4) and DHITsig- (N=3). RNA sequencing data of 9 patient samples and 3 GCB DLBCL cell lines were generated and analyzed using Salmon and DESeq2. Simple somatic mutations from SR-WGS data were called using an ensemble of somatic variant callers, which confirmed all cases to belong to the EZB genetic subgroup. Methylated bases were called with Nanopolish followed by a differential methylation analysis using the DSS package in R to identify differences in methylation between both subgroups. Differentially methylated regions (DMRs) were annotated for GC B-cell specific superenhancers, CpG islands, and genes. Using WhatsHap and Nanomethphase, heterozygous single nucleotide polymorphisms (SNPs) from SR and LR data were used to phase LRs into haplotypes. Haplotype analyses were performed to determine the phase of aberrant somatic hypermutation (aSHM) and methylation patterns in the context of proximal translocation breakpoints. Results Most DMRs were found to be associated with regulatory regions such as promoters, superenhancers, and CpG islands. We found methylation differences between DHITsig+ and DHITsig- subgroups in 1430 superenhancers known to be active in GC B-cells. Within these superenhancer regions, DHITsig+ tumors were typically hypomethylated relative to DHITsig-. Several DHITsig classifier genes, including WNK2, TNFSF8, and IL21R, had methylation patterns inverted relative to the typical expression pattern (i.e. low methylation with high expression). Similarly, among 633 differentially methylated promoters, several including TCF3 and WNK2 demonstrated the same inverted methylation pattern. CpG islands showed distinct patterns of methylation in both subgroups that were inversely associated with expression, this also included genes differentially expressed between DHITsig+ and DHITsig- tumors. The DHITsig- group had increased levels of methylation of DHITsig genes in both promoters and gene bodies suggesting a functional role for gene body methylation. Finally, we observed that a high density of mutations near the BCL2 transcription start site consistently phased into the same haplotype as the BCL2-IGH translocation in five tumours, demonstrating the allele specificity of aSHM associated with these events (Panel B). Conclusions These results demonstrate differences in methylation between the subgroups. The association of hypomethylated superenhancers, promoters, and CpG islands with genes over-expressed in DHITsig+ suggests these methylation patterns may contribute to the DHITsig phenotype. These results may aid in improving GCB DLBCL patient classification and reveal factors contributing to its heterogeneity. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal