Adjacent Non-cancerous Tissues Research Articles

Molecular evolution in different subtypes ofmultifocal hepatocellular carcinoma.

Multifocal hepatocellular carcinoma (MF-HCC) accounts for > 40% of HCCs, exhibiting a poor prognosis than single primary HCCs. Characterizing molecular features including dynamic changes of mutational signature along with clonal evolution, intrahepatic metastatic timing, and genetic footprint in the preneoplastic stage underlying different subtypes of MF-HCC are important for understanding their molecular evolution and developing a precision management strategy. We conducted whole-exome sequencing in 74 tumor samples from spatially distinct regions in 35 resected lesions and adjacent noncancerous tissues from 11 patients, 15 histologically confirmed preneoplastic lesions, and six samples from peripheral blood mononuclear cells. A previously published MF-HCC cohort (n = 9) was included as an independent validation dataset. We combined well-established approaches to investigate tumor heterogeneity, intrahepatic metastatic timing, and molecular footprints in different subtypes of MF-HCCs. We classified MF-HCCs patients into three subtypes, including intrahepatic metastasis, multicentric occurrence, and mixed intrahepatic metastasis and multicentric occurrence. The dynamic changes in mutational signatures between tumor subclonal expansions demonstrated varied etiologies (e.g., aristolochic acid exposure) underlying the clonal progression in different MF-HCC subtypes. Furthermore, the clonal evolution in intrahepatic metastasis exhibited an early metastatic seeding at 10-4-0.01cm3 in primary tumor volume (below the limits of clinical detection), further validated in an independent cohort. In addition, mutational footprints in the preneoplastic lesions for multicentric occurrence patients revealed common preneoplastic arising clones, evidently being ancestors of different tumor lesions. Our study comprehensively characterized the varied tumor clonal evolutionary history underlying different subtypes of MF-HCC and provided important implications for optimizing personalized clinical management for MF-HCC.

Characterisation of cell lines derived from prostate cancer patients with localised disease.

Prostate cancer is a broad-spectrum disease, spanning from indolent to a highly aggressive lethal malignancy. Prostate cancer cell lines are essential tools to understanding the basic features of this malignancy, as well as in identifying novel therapeutic strategies. However, most cell lines routinely used in prostate cancer research are derived from metastatic disease and may not fully elucidate the molecular events underlying the early stages of cancer development and progression. Thus, there is a need for new cell lines derived from localised disease to better span the disease spectrum. Prostatic tissue from the primary site, and adjacent non-cancerous tissue was obtained from four patients with localised disease undergoing radical prostatectomy. Epithelial cell outgrowths were immortalised with human papillomavirus type 16 (HPV16) E6 and E7 to establish monoclonal cell lines. Chromosomal ploidy was imaged and STR profiles were determined. Cell morphology, colony formation and cell proliferation characteristics were assessed. Androgen receptor (AR) expression and AR-responsiveness to androgen treatment were analysed by immunofluorescence and RT-qPCR, respectively. RNA-seq analysis was performed to identify prostate lineage markers and expression of prostate cancer tumorigenesis-related genes. Two benign cell lines derived from non-cancer cells (AQ0420 and AQ0396) and two tumour tissue derived cancer cell lines (AQ0411 and AQ0415) were immortalised from four patients with localised prostatic adenocarcinoma. The cell lines presented an epithelial morphology and a slow to moderate proliferative rate. None of the cell lines formed anchorage independent colonies or displayed AR-responsiveness. Comparative RNA-seq expression analysis confirmed the prostatic lineage of the four cell lines, with a distinct gene expression profile from that of the metastatic prostate cancer cell lines, PC-3 and LNCaP. Comprehensive characterization of these cell lines may provide new in vitro tools that could bridge the current knowledge gap between benign, early-stage and metastatic disease.

Dysregulation of PVT1 and NEAT1 lncRNAs in pituitary adenomas

Pituitary adenomas are slow-growing tumors originated from the anterior part of pituitary gland. These tumors are associated with dysregulation of a number of long non-coding RNAs (lncRNAs). PVT1, TUG1, MALAT1, NEAT1 and GAS5 are among lncRNAs with important roles in the regulation of cell proliferation, cell apoptosis, cell differentiation and cell cycle transition. In the current study, we assessed expression levels of PVT1, TUG1, MALAT1, NEAT1 and GAS5 in the pituitary adenoma samples compared with adjacent non-cancerous samples to find their relevance with this type of tumors and their potential as diagnostic markers in these tumors. Expression of NEAT1 was significantly higher in total adenoma tissues (Expression ratio (95% CI)= 7.06 (2.31–21.4), P value= 0.02) and in non-functioning pituitary adenoma (NFPA) samples (Expression ratio (95% CI)= 8.5 (2.17–33.12), P value= 0.04) compared with corresponding controls. Although both lncRNAs had appropriate sensitivity values for discrimination of NFPAs from adjacent non-cancerous tissues (0.84 and 0.90 for PVT1 and NEAT1, respectively), the calculated AUC values were not adequate for either lncRNAs (0.63 ± 0.04 and 0.58 ± 0.04 for PVT1 and NEAT1, respectively). Therefore, NEAT1 and PVT1 lncRNAs are dysregulated in NFPA. The current study suggests the role of NEAT1 and PVT1 in the pathogenesis of NFPA.

Activation of ACLY by SEC63 deploys metabolic reprogramming to facilitate hepatocellular carcinoma metastasis upon endoplasmic reticulum stress

BackgroundTumor cells display augmented capability to maintain endoplasmic reticulum (ER) homeostasis and hijack ER stress pathway for malignant phenotypes under microenvironmental stimuli. Metabolic reprogramming is a well-known hallmark for tumor cells to provide specific adaptive traits to the microenvironmental alterations. However, it’s unknown how tumor cells orchestrate metabolic reprogramming and tumor progression in response to ER stress. Herein, we aimed to explore the pivotal roles of SEC63-mediated metabolic remodeling in hepatocellular carcinoma (HCC) cell metastasis after ER stress.MethodsThe expression levels of SEC63 in HCC tissues and adjacent non-cancerous tissues were determined by immunohistochemistry and western blot. The regulatory roles of SEC63 in HCC metastasis were investigated both in vitro and in vivo by RNA-sequencing, metabolites detection, immunofluorescence, and transwell migration/invasion analyses. GST pull-down, immunoprecipitation/mass spectrometry and in vivo ubiquitination/phosphorylation assay were conducted to elucidate the underlying molecular mechanisms.ResultsWe identified SEC63 as a new regulator of HCC cell metabolism. Upon ER stress, the phosphorylation of SEC63 at T537 by IRE1α pathway contributed to SEC63 activation. Then, the stability of ACLY was upregulated by SEC63 to increase the supply of acetyl-CoA and lipid biosynthesis, which are beneficial for improving ER capacity. Meanwhile, SEC63 also entered into nucleus for increasing nuclear acetyl-CoA production to upregulate unfolded protein response targets to improve ER homeostasis. Importantly, SEC63 coordinated with ACLY to epigenetically modulate expression of Snail1 in the nucleus. Consequently, SEC63 promoted HCC cell metastasis and these effects were reversed by ACLY inhibition. Clinically, SEC63 expression was significantly upregulated in HCC tissue specimens and was positively correlated with ACLY expression. Importantly, high expression of SEC63 predicted unfavorable prognosis of HCC patients.ConclusionsOur findings revealed that SEC63-mediated metabolic reprogramming plays important roles in keeping ER homeostasis upon stimuli in HCC cells. Meanwhile, SEC63 coordinates with ACLY to upregulate the expression of Snail1, which further promotes HCC metastasis. Metastasis is crucial for helping cancer cells seek new settlements upon microenvironmental stimuli. Taken together, our findings highlight a cancer selective adaption to ER stress as well as reveal the potential roles of the IRE1α-SEC63-ACLY axis in HCC treatment.

Identification of lncRNA, miRNA and mRNA expression profiles and ceRNA Networks in small cell lung cancer

BackgroundSmall cell lung cancer (SCLC) is a highly lethal malignant tumor. It accounts for approximately 15% of newly diagnosed lung cancers. Long non-coding RNAs (lncRNAs) can regulate gene expression and contribute to tumorigenesis through interactions with microRNAs (miRNAs). However, there are only a few studies reporting the expression profiles of lncRNAs, miRNAs, and mRNAs in SCLC. Also, the role of differentially expressed lncRNAs, miRNAs, and mRNAs in relation to competitive endogenous RNAs (ceRNA) network in SCLC remain unclear.ResultsIn the present study, we first performed next generation sequencing (NGS) with six pairs of SCLC tumors and adjacent non-cancerous tissues obtained from SCLC patients. Overall, 29 lncRNAs, 48 miRNAs, and 510 mRNAs were found to be differentially expressed in SCLC samples (|log2[fold change] |> 1; P < 0.05). Bioinformatics analysis was performed to predict and construct a lncRNA-miRNA-mRNA ceRNA network, which included 9 lncRNAs, 11 miRNAs, and 392 mRNAs. Four up-regulated lncRNAs and related mRNAs in the ceRNA regulatory pathways were selected and validated by quantitative PCR. In addition, we examined the role of the most upregulated lncRNA, TCONS_00020615, in SCLC cells. We found that TCONS_00020615 may regulate SCLC tumorigenesis through the TCONS_00020615–hsa-miR-26b-5p–TPD52 pathway.ConclusionsOur study provided the comprehensive analysis of the expression profiles of lncRNAs, miRNAs, and mRNAs of SCLC tumors and adjacent non-cancerous tissues. We constructed the ceRNA networks which may provide new evidence for the underlying regulatory mechanism of SCLC. We also found that the lncRNA TCONS_00020615 may regulate the carcinogenesis of SCLC.

Exploring the hub genes and mechanisms of Daphne altaica treating esophageal squamous cell carcinoma based on network pharmacology and bioinformatics analysis.

Esophageal squamous cell carcinoma (ESCC), is a frequent digestive tract malignant carcinoma with a high fatality rate. Daphne altaica (D. altaica), a medicinal plant that is frequently employed in Kazakh traditional medicine, and which has traditionally been used to cure cancer and respiratory conditions, but research on the mechanism is lacking. Therefore, we examined and verified the hub genes and mechanism of D. altaica treating ESCC. Active compounds and targets of D. altaica were screened by databases such as TCMSP, and ESCC targets were screened by databases such as GeneCards and constructed the compound-target network and PPI network. Meantime, data sets between tissues and adjacent non-cancerous tissues from GEO database (GSE100942, GPL570) were analyzed to obtain DEGs using the limma package in R. Hub genes were validated using data from the Kaplan-Meier plotter database, TIMER2.0 and GEPIA2 databases. Finally, AutoDock software was used to predict the binding sites through molecular docking. In total, 830 compound targets were obtained from TCMSP and other databases. In addition, 17,710 disease targets were acquired based on GeneCards and other databases. In addition, we constructed the compound-target network and PPI network. Then, 127 DEGs were observed (82 up-regulated and 45 down-regulated genes). Hub genes were screened including TOP2A, NUF2, CDKN2A, BCHE, and NEK2, and had been validated with the help of several publicly available databases. Finally, molecular docking results showed more stable binding between five hub genes and active compounds. In the present study, five hub genes were screened and validated, and potential mechanisms of action were predicted, which could provide a theoretical understanding of the treatment of ESCC with D. altaica.

The neurosurgical benefit of contactless in vivo optical coherence tomography regarding residual tumor detection: A clinical study.

In brain tumor surgery, it is crucial to achieve complete tumor resection while conserving adjacent noncancerous brain tissue. Several groups have demonstrated that optical coherence tomography (OCT) has the potential of identifying tumorous brain tissue. However, there is little evidence on human in vivo application of this technology, especially regarding applicability and accuracy of residual tumor detection (RTD). In this study, we execute a systematic analysis of a microscope integrated OCT-system for this purpose. Multiple 3-dimensional in vivo OCT-scans were taken at protocol-defined sites at the resection edge in 21 brain tumor patients. The system was evaluated for its intraoperative applicability. Tissue biopsies were obtained at these locations, labeled by a neuropathologist and used as ground truth for further analysis. OCT-scans were visually assessed with a qualitative classifier, optical OCT-properties were obtained and two artificial intelligence (AI)-assisted methods were used for automated scan classification. All approaches were investigated for accuracy of RTD and compared to common techniques. Visual OCT-scan classification correlated well with histopathological findings. Classification with measured OCT image-properties achieved a balanced accuracy of 85%. A neuronal network approach for scan feature recognition achieved 82% and an auto-encoder approach 85% balanced accuracy. Overall applicability showed need for improvement. Contactless in vivo OCT scanning has shown to achieve high values of accuracy for RTD, supporting what has well been described for ex vivo OCT brain tumor scanning, complementing current intraoperative techniques and even exceeding them in accuracy, while not yet in applicability.

DNAJB4 promotes triple-negative breast cancer cell apoptosis via activation of the Hippo signaling pathway

IntroductionTriple-negative breast cancer (TNBC) is currently the most malignant subtype of breast cancer without effective targeted therapies. DNAJB4 (Dnaj heat shock protein family (Hsp40) member B4) is a member of the human heat shock protein family (Hsp40). The clinical significance of DNAJB4 in breast cancer has been reported in our previous study. However, the biological function of DNAJB4 in TNBC cell apoptosis remains unclear to date.MethodsThe expression of DNAJB4 in normal breast cells, breast cancer cells, four-paired TNBC tissues, and adjacent noncancerous tissues was quantified by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot assay. The role of DNAJB4 in TNBC cell apoptosis was investigated using a number of gain- and loss-of-function in vitro and in vivo assays. The underlying molecular mechanisms in TNBC cell apoptosis were elucidated via Western blot assay.ResultsDNAJB4 expression was significantly downregulated in TNBC tissues and cell lines. DNAJB4 knockdown inhibited TNBC cell apoptosis and promoted tumorigenicity in vitro and in vivo, but DNAJB4 overexpression resulted in the opposite. Mechanically, DNAJB4 knockdown inhibited TNBC cell apoptosis through suppression of the Hippo signaling pathway, and the result was reversed after DNAJB4 overexpression.ConclusionsDNAJB4 promotes TNBC cell apoptosis by activating the Hippo signaling pathway. Therefore, DNAJB4 may act as a prognostic biomarker and therapeutic target for TNBC.

Comprehensive proteome, phosphoproteome and kinome characterization of luminal A breast cancer.

Breast cancer is one of the most frequently occurring malignant cancers worldwide. Invasive ductal carcinoma (IDC) and invasive lobular carcinoma (ILC) are the two most common histological subtypes of breast cancer. In this study, we aimed to deeply explore molecular characteristics and the relationship between IDC and ILC subtypes in luminal A subgroup of breast cancer using comprehensive proteomics and phosphoproteomics analysis. Cancer tissues and noncancerous adjacent tissues (NATs) with the luminal A subtype (ER- and PR-positive, HER2-negative) were obtained from paired IDC and ILC patients respectively. Label-free quantitative proteomics and phosphoproteomics methods were used to detect differential proteins and the phosphorylation status between 10 paired breast cancer and NATs. Then, the difference in protein expression and its phosphorylation between IDC and ILC subtypes were explored. Meanwhile, the activation of kinases and their substrates was also revealed by Kinase-Substrate Enrichment Analysis (KSEA). In the luminal A breast cancer, a total of 5,044 high-confidence proteins and 3,808 phosphoproteins were identified from 10 paired tissues. The protein phosphorylation level in ILC tissues was higher than that in IDC tissues. Histone H1.10 was significantly increased in IDC but decreased in ILC, Conversely, complement C4-B and Crk-like protein were significantly decreased in IDC but increased in ILC. Moreover, the increased protein expression of Septin-2, Septin-9, Heterogeneous nuclear ribonucleoprotein A1 and Kinectin but reduce of their phosphorylation could clearly distinguish IDC from ILC. In addition, IDC was primarily related to energy metabolism and MAPK pathway, while ILC was more closely involved in the AMPK and p53/p21 pathways. Furthermore, the kinomes in IDC were primarily significantly activated in the CMGC groups. Our research provides insights into the molecular characterization of IDC and ILC and contributes to discovering novel targets for further drug development and targeted treatment.

RNA sequencing revealed the multi-stage transcriptome transformations during the development of gallbladder cancer associated with chronic inflammation.

Gallbladder cancer (GBC) is a highly malignant tumor with extremely poor prognosis. Previous studies have suggested that the carcinogenesis and progression of GBC is a multi-stage and multi-step process, but most of them focused on the genome changes. And a few studies just compared the transcriptome differences between tumor tissues and adjacent noncancerous tissues. The transcriptome changes, relating to every stage of GBC evolution, have rarely been studied. We selected three cases of normal gallbladder, four cases of gallbladder with chronic inflammation induced by gallstones, five cases of early GBC, and five cases of advanced GBC, using next-generation RNA sequencing to reveal the changes in mRNAs and lncRNAs expression during the evolution of GBC. In-depth analysis of the sequencing data indicated that transcriptome changes from normal gallbladder to gallbladder with chronic inflammation were distinctly related to inflammation, lipid metabolism, and sex hormone metabolism; transcriptome changes from gallbladder with chronic inflammation to early GBC were distinctly related to immune activities and connection between cells; and the transcriptome changes from early GBC to advanced GBC were distinctly related to transmembrane transport of substances and migration of cells. Expression profiles of mRNAs and lncRNAs change significantly during the evolution of GBC, in which lipid-based metabolic abnormalities play an important promotive role, inflammation and immune activities play a key role, and membrane proteins are very highlighted molecular changes.

LncRNA-NEAT1 inhibits the occurrence and development of pancreatic cancer through spongy miR-146b-5p/traf6

ABSTRACT To investigate the inhibitory effect of LNCNA-NEA1 on pancreatic cancer development and progression via spongiosa miR-146b-5p/TRAF6, 60 pancreatic cancer patients diagnosed from December 2017 to December 2019 were selected as a general source of information. Real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR) was used to detect the expression level of NEAT1 in cancerous and adjacent non-cancerous tissues. Cell counting kit-8 (CCK-8) and transwell were used to determine the effect of LNCNA-NEA1 on the proliferation and migration of pancreatic cancer cells (Panc-1). The results of dual luciferase reporter gene assay showed that nea 1 could target and regulate the expression of spongy miR-146b-5p/TRAF6, and reducing the expression of spongy miR-146b-5p/TRAF6 could reverse the inhibitory effects of nea 1-siRNA on proliferation, migration and invasion of pancreatic cancer cells. Therefore, it was concluded that knockdown of nea 1 could inhibit the proliferation, migration and invasion of pancreatic cancer cells by upregulating the level of miR-146b-5p/TRAF6, and the expression of lnc RNA-nea 1 could be used as an indicator for preoperative diagnosis and postoperative prognosis of pancreatic cancer patients.

Immune profiling and prognostic model of pancreatic cancer using quantitative pathology and single-cell RNA sequencing

BackgroundPancreatic ductal adenocarcinoma (PDAC) has a complex tumor immune microenvironment (TIME), the clinical value of which remains elusive. This study aimed to delineate the immune landscape of PDAC and determine the clinical value of immune features in TIME.MethodsUnivariable and multivariable Cox regression analyses were performed to evaluate the clinical value of immune features and establish a new prognostic model. We also conducted single-cell RNA sequencing (scRNA-seq) to further characterize the immune profiles of PDAC and explore cell-to-cell interactions.ResultsThere was a significant difference in the immune profiles between PDAC and adjacent noncancerous tissues. Several novel immune features were captured by quantitative pathological analysis on multiplex immunohistochemistry (mIHC), some of which were significantly correlated with the prognosis of patients with PDAC. A risk score-based prognostic model was established based on these immune features. We also constructed a user-friendly nomogram plot to predict the overall survival (OS) of patients by combining the risk score and clinicopathological features. Both mIHC and scRNA-seq analysis revealed PD-L1 expression was low in PDAC. We found that PD1 + cells were distributed in different T cell subpopulations, and were not enriched in a specific subpopulation. In addition, there were other conserved receptor-ligand pairs (CCL5-SDC1/4) besides the PD1-PD-L1 interaction between PD1 + T cells and PD-L1 + tumor cells.ConclusionOur findings reveal the immune landscape of PDAC and highlight the significant value of the combined application of mIHC and scRNA-seq for uncovering TIME, which might provide new clues for developing immunotherapy combination strategies.

Overexpression and Role of HHLA2, a Novel Immune Checkpoint, in Colorectal Cancer.

The study aimed to investigate correlations between HHLA2 levels and parameters, including microsatellite instability (MSI) status, CD8+ cells, and histopathological features: budding, tumor-infiltrating lymphocytes (TILs), TNM scale, grading, cytokines, chemokines, and cell signaling moleculesin colorectal cancer (CRC). Furthermore, the immune infiltration landscape and HHLA2-related pathways in colorectal cancer using available online datasets were analyzed. The study included 167 patients diagnosed with CRC. Expression of HHLA2 was detected by immunohistochemistry method (IHC) and enzyme-linked immunosorbent assay (ELISA). The IHC was used to evaluate the MSI and CD8+ status. The budding and TILs were measured using a light microscope. The concentrations of cytokines, chemokines, and cell signaling molecules were measured to analyze the data by the Bio-Plex Pro Human cytokine screening panel, 48 cytokine assay, and principal component analysis (PCA). Geneset enrichment analysis (GSEA) was conducted to identify HHLA2-related pathways. The biological function of HHLA2 was predicted by Gene Ontology (GO). Analysis of the immune infiltration landscape of HHLA2 in colorectal cancer was made by the web-based tool Camoip. High HHLA2 expression was detected in CRC tumor tissues compared to the adjacent noncancerous tissues. The percentage of HHLA2-positive tumors was 97%. GSEA and GO showed that HHLA2 upregulation correlated with cancer-related pathways and several biological functions. Tumor-infiltrating lymphocytes score correlated positively with IHC HHLA2 expression level percentage. There was a negative correlation between HHLA2, anti-tumor cytokines and pro-tumor growth factors. This study provides a valuable insight into the role of HHLA2 in CRC. We reveal the role of HHLA2 expression as well as a stimulatory and inhibitory immune checkpoint in colorectal cancer. Further research may verify the therapeutic values of the HHLA2-KIR3DL3/TMIGD2 pathway in colorectal cancer.

ALKBH5-induced circular RNA NRIP1 promotes glycolysis in thyroid cancer cells by targeting PKM2.

Although circular RNAs (circRNAs) are involved in cell proliferation, differentiation, apoptosis, and invasion, the underlying regulatory mechanisms of circRNAs in thyroid cancer have not been fully elucidated. This article aimed to study the role of circRNA regulated by N6-methyladenosine modification in papillary thyroid cancer (PTC). Quantitative real-time PCR, western blotting, and immunohistochemistry were used to investigate the expressions of circRNA nuclear receptor-interacting protein 1 (circNRIP1) in PTC tissues and adjacent noncancerous thyroid tissues. In vitro and in vivo assays were carried out to assess the effects of circNRIP1 on PTC glycolysis and growth. The N6-methyladenosine mechanisms of circNRIP1 were evaluated by methylated RNA immunoprecipitation sequencing, luciferase reporter gene, and RNA stability assays. Results showed that circNRIP1 levels were significantly upregulated in PTC tissues. Furthermore, elevated circNRIP1 levels in PTC patients were correlated with high tumor lymph node metastasis stage and larger tumor sizes. Functionally, circNRIP1 significantly promoted glycolysis, PTC cell proliferation in vitro, and tumorigenesis in vivo. Mechanistically, circNRIP1 acted as a sponge for microRNA (miR)-541-5p and miR-3064-5p and jointly upregulated pyruvate kinase M2 (PKM2) expression. Knockdown of m6 A demethylase α-ketoglutarate-dependent dioxygenase alkB homolog 5 (ALKBH5) significantly enhanced circNRIP1 m6 A modification and upregulated its expression. These results show that ALKBH5 knockdown upregulates circNRIP1, thus promoting glycolysis in PTC cells. Therefore, circNRIP1 can be a prognostic biomarker and therapeutic target for PTC by acting as a sponge for oncogenic miR-541-5p and miR-3064-5p to upregulate PKM2 expression.

Aberrant DNA Methylation Patterns of Deleted in Liver Cancer 1 Isoforms in Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC), a common primary liver cancer, is the third leading cause of death worldwide. DNA methylation changes are common in HCC and have been studied to be associated with hepatocarcinogenesis. In our study, we used the MassARRAY® EpiTYPER technology to investigate the methylation differences of deleted in liver cancer 1 (DLC1) (isoform 1 and 3) promoter between HCC tissues and corresponding adjacent noncancerous tissues and the association between methylation levels and clinicopathological features. In addition, the modified CRISPR-Cas9 system and the DNA methyltransferase inhibitor (DNMTi) were utilized to explore the functional correlation of epigenetic modifications and DLC1 gene regulation. The methylation levels of the DLC1 isoforms in HCC samples were found significantly lower than those in the adjacent noncancerous tissues (all p < 0.0001). Also, we found that the expression of DLC1 could be bidirectionally regulated by the modified CRISPR-Cas9 system and the DNMTi. Moreover, the hypomethylation of DLC1 in HCC samples was connected with the presence of satellite lesions (p = 0.0305) and incomplete tumor capsule (p = 0.0204). Receiver operator characteristic curve analysis demonstrated that the methylation levels of DLC1 could be applied to discriminate HCC patients (area under the curve = 0.728, p < 0.0001). The hypomethylation status was a key regulatory mechanism of DLC1 expression and might serve as a potential biomarker for HCC.

Mitochondrial elongation factor 4 modulates energy metabolism and promotes breast cancer metastasis by orchestration of mitochondrial translation

To cope with the requirements of energy and building blocks for rapid proliferation, cancer cells reprogram their metabolic pathways profoundly, especially in oxygen- and nutrients-deficient tumor microenvironments. However, functional mitochondria and mitochondria-dependent oxidative phosphorylation are still necessary for the tumorigenesis and metastasis of cancer cells. We show here that mitochondrial elongation factor 4 (mtEF4) is commonly upregulated in breast tumors compared to adjacent non-cancerous tissues, and is relevant to tumor progression and poor prognosis. Down regulation of mtEF4 in breast cancer cells impairs the assembly of mitochondrial respiration complexes, decreases mitochondrial respiration, reduces ATP production, attenuates the formation of lamellipodia, and suppresses cell motility in vitro and cancer metastasis in vivo. On the contrary, upregulation of mtEF4 elevates the mitochondrial oxidative phosphorylation, which contributes to the migratory capacities of breast cancer cells. mtEF4 also increases the potential of glycolysis, probably via an AMPK-related mechanism. In summary, we provide direct evidences that the aberrantly upregulated mtEF4 contributes to the metastasis of breast cancer by coordinating metabolic pathways.

Prognostic Value of Liver Kinase B1 (LKB1) in Gastric Cancer-Associated Tumor Microenvironment Immunity

Liver kinase B1 (LKB1) is a tumor suppressor gene, the inactivation of which occurs frequently in different tumor types. However, whether LKB1 is associated with the clinical features of gastric cancer (GC) and regulating tumor immunity is unknown. In this study, we showed that LKB1 is highly expressed in the serum of healthy individuals (n = 176) compared to GC patients (n = 416) and is also associated with clinical outcomes and good survival rates in GC patients. Furthermore, genes associated with immune checkpoints and T cell activation, such as PD-1, PD-L1, CD8A, CD8B, CD28, and GZMM, were shown to be highly expressed in GC subgroups with high LKB1 expression. Compared with fresh gastric cancerous tissues, LKB1 was highly expressed in CD3+CD8+ and CD3+CD8+CD28+ T cells in fresh adjacent non-cancerous tissues. CD3+CD8+ T cells produced an IFN-γ anti-cancer immune response. Furthermore, the proportion of CD3+CD8+ T cells that expressed LKB had a positive correlation with IFN-γ expression. Moreover, GC patients with low LKB1 expression had a poor objective response rate, and worse progression-free survival and overall survival when treated with pembrolizumab. In conclusion, LKB1 may be a potential immune checkpoint in GC patients.

NLRP3 inflammasome activation By 17β-estradiol is a potential therapeutic target in hepatocellular carcinoma treatment

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, and it mostly arises as a consequence of persistent chronic inflammation. Recently, NLRP3 inflammasome has caught the attention of many research groups due to its involvement in different types of cancer. However, its direct role in HCC remains elusive. Our study aimed to evaluate the role of NLRP3 inflammasome and pyroptosis in HCC and to clarify the potential mechanism by which 17β-estradiol (E2) can be used as a protective factor against HCC. NLRP3, caspase-1 (CASP1) as well as gasdermin-D (GSDMD) mRNA expression levels were assessed in human HCC tissues and adjacent non-cancerous liver tissues. Also, HepG2 HCC cells were cultured and treated with E2, followed by detection of the mRNA levels of these three genes. Our results revealed that NLRP3, CASP1, and GSDMD mRNA expressions were significantly lower in HCC tissues than in controls, and this under-expression was closely correlated with advanced HCC stages and grades. In contrast, HepG2 HCC cells displayed significantly higher expression levels of NLRP3 inflammasome components and GSDMD in the two E2-treated groups compared to the untreated group. Also, NLRP3, CASP1, and GSDMD mRNA expression levels were positively correlated with each other. This study confirmed that lack of NLRP3 inflammasome is involved in HCC progression and 17β-estradiol-induced activation of NLRP3 inflammasome may be effective in HCC treatment as it inhibited tumor cell growth and proliferation by triggering CASP1-dependent pyroptosis in HCC cells.

ARHGAP6 inhibits bladder cancer cell viability, migration, and invasion via β-catenin signaling and enhances mitomycin C sensitivity.

The Rho/ROCK pathway regulates diverse cellular processes and contributes to the development and advancement of several types of human cancers. This study investigated the role of specific Rho GTPase-activating proteins (RhoGAP), ARHGAP6, in bladder cancer (BC). In this study, ARHGAP6 expression in BC and its clinical significance were investigated. In vitro and in vivo assays were used to explore the tumor-related function and the underlying molecular mechanism ARHGAP6 of in BC. The mRNA and protein levels of ARHGAP6 significantly reduced in human BC tissues and cell lines compared with corresponding adjacent non-cancerous tissues and normal urothelial cells. In vitro, ARHGAP6 overexpression markedly decreased the viability, migration, and invasion of BC cells. Interestingly, low ARHGAP6 expression in BC strongly correlated with poor patient survival and was highly associated with metastasis and β-catenin signaling. Furthermore, ARHGAP6 expression strongly influenced the sensitivity of BC cells to mitomycin C treatment. Together, our results demonstrate that ARHGAP6 plays critical roles in regulating the proliferation, migration, invasion, and metastasis of BC cells possibly via the modulation of β-catenin and strongly influences the chemosensitivity of BC cells.

Evaluating the local expression pattern of IGF-1R in tumor tissues and the circulating levels of IGF-1, IGFBP-1, and IGFBP-3 in the blood of patients with different primary bone tumors.

The present study tried to provide insights into the expression pattern and diagnostic significance of the IGF-1 axis main mediators in three main primary bone tumor types with different degrees of severity. The real-time qRT-PCR (to analyze IGF-1R gene expression), the immunohistochemistry (to measure IGF-1R protein), and the ELISA assay (to assess the circulating level of IGF-1, IGFBP-1, and IGFBP-3) were applied to confirm this hypothesis. A total number of 180 bone tissues (90 tumors and 90 noncancerous adjacent tissues) and 120 blood samples drained from 90 patients with bone tumors and 30 healthy controls were enrolled in the study. The association of insulin-like growth factor (IGF)-1 axis expression pattern with the patient's clinical pathological characteristics and tumor aggressive features, the diagnostic and predictive values were assessed for all tumor groups. A significantly elevated level of IGF-1R gene and protein was detected in bone tumors compared to the noncancerous bone tissues that were prominent in osteosarcoma and Ewing sarcoma compared to the GCT group. The positive association of the IGF-1R gene and protein level with tumor grade, metastasis, and recurrence was detected in the osteosarcoma and Ewing sarcoma groups. The circulating level of IGF-1, IGFPB-1, and IGFBP-3 were increased in osteosarcoma and Ewing sarcoma and GCT groups that were correlated significantly to the tumor severity. The ability of the IGF-1 axis to discriminate between bone tumors also malignant and benign tumors was considerable. In summary, our data suggested that IGF-1R, IGF-1, IGFBP-1, and IGFBP-3 levels are associated with bone tumor malignancy, metastasis, and recurrence that might serve as biomarkers for osteosarcoma and Ewing sarcoma recurrence.