Cancer-related Signaling Pathways Research Articles

Effect of SIRT7 on inhibiting the epithelial-mesenchymal transformation in pancreatic cancer cells and related mechanism

Objective: To investigate the effect and mechanism of SIRT7 in epithelial mesenchymal transformation (EMT) of pancreatic cancer cells. Methods: The pancreatic cancer cells were divided into siControl, siSIRT7, over-expression SIRT7, siSIRT7+siCOL4A1, and siSIRT7+siSLUG groups using siRNA or plasmid transfection. The proliferation, migration and invasion of pancreatic cancer cells were detected by EdU, wound healing assay and Transwell experiments, respectively. The expression of EMT and cancer stem cell (CSC) markers were detected by quantitative real-time reverse transcription polymerase chain reaction assay (qRT-PCR) and western blot. RNA sequencing (RNA-seq) in SIRT7 knockdown PANC-1 cells was performed to explore the signaling pathways and target genes regulated by SIRT7. Then the target genes directly regulated by SIRT7 were identified with quantitative chromatin immunoprecipitation experiment (q-ChIP) and chromatin immunoprecipitation polymerase chain reaction (ChIP-PCR). The expressions of SIRT7 and target genes were detected by immunohistochemical (IHC) in pancreatic cancer tissues, and the correlation between SIRT7 and target gene expression was analyzed using TCGA dataset. The correlation between expression of SIRT7 or target genes and survival was analyzed on KM-plotter website. Finally, GeneMANIA, STRING and ENCORI were used to predict SIRT7-related proteins and miRNAs. Results: EdU assay showed that the cell proliferation rates in SIRT7-overexpressed PANC-1 [(19.33±0.35)%] and BxPC-3 cells [(17.00±1.89)%] were lower than those in the control group [(31.60±1.37)% and (24.33±0.78)%, respectively, P＜0.05]. The proliferation rates of SIRT7-knockdown PANC-1 [(23.94±1.00)% and (27.08±0.97)%] and BxPC-3 cells [(22.00±1.86)% and (25.96±1.61)%] were higher than those of the siControl group [(11.80±1.86)% and (13.42±1.39)%, respectively, P＜0.05]. In PANC-1 cells, the wound healing assay showed that the relative migration rate of SIRT7-overexpression cells [(76.67±2.74)%] was lower than that of control cells [(100.00±2.13)%, P＜0.05]; the relative migration rate of cells with SIRT7 knockdown [(134.22±4.08)% and (199.82±9.20)%, respectively] was higher than that of siControl group [(102.24±3.13)%, P＜0.05]. Compared with the control group, SIRT7 overexpression decreased the number of migrated BxPC-3 cells (45.66±1.69 vs 28.33±2.62, P＜0.05); while SIRT7 knockdown increased these numbers (65.66±2.86 and 82.00±2.94 versus 33.00±0.81, P＜0.01). Transwell experiment revealed that the number of invaded cells in SIRT7 overexpression groups (16.33±2.05 and 34.66±1.69) was lower than that control groups (54.33±4.64 and 58.66±5.90, P＜0.05); with SIRT7 knockdown, the numbers of invaded PANC-1 (63.66±2.49 and 69.33±3.29) and BxPC-3 cells (134.33±3.09 and 181.66±4.02) were higher than those in control groups (35.33±2.49 and 42.00±0.81, P˂0.05). Also, SIRT7 knockdown decreased the expressions of epithelial markers and increased the expressions of mesenchymal and CSC markers. RNA-seq analysis showed that SIRT7 was involved in regulating a variety of cancer-related signaling pathways, including the pancreatic cancer pathway and the EMT pathway. Furthermore, SIRT7 could directly bind to the promoter regions of target genes, such as COL4A1 and SLUG. SIRT7 was negatively correlated with the expression and function of COL4A1 and SLUG in pancreatic cancer cells. The expressions of SIRT7, COL4A1, SLUG and SOX2 were verified in pancreatic cancer tissues by IHC. Finally, SIRT7 was predicted to be associated with many proteins and miRNAs based on GeneMANIA, STRING, and ENCORI online tools. Conclusions: SIRT7 can inhibit the EMT of pancreatic cancer cells through transcriptionally inhibiting the expression of target genes, such as COL4A1 and SLUG. Thus, SIRT7 may serve as a potential tumor suppressor gene in pancreatic cancer.

Unraveling the role of tissue colonized microbiome in ovarian cancer progression

BackgroundOvarian cancer (OC) is found to be the third most common gynecologic malignancy over the world, having the highest mortality rate among such tumors. Emerging studies underscore the presence of microorganisms within tumor tissues, with certain pathogens intricately linked to disease onset and progression. Disruption of the microbiome frequently precipitates disturbances in host metabolic and immune pathways, thereby fostering the development of cancer. MethodsIn this study, we initiated the investigation by conducting microbial reannotation on the RNA sequencing data derived from ovarian cancer tissues. Subsequently, a comprehensive array of analyses on tissue microbes was executed. These analyses encompassed the assessment of intergroup variations in microbial diversity, differential microbiological analysis, exploration of the association between host gene expression and microbial abundance, as well as an enrichment analysis of functional pathways linked to host genes associated with microbes. ResultsThe analysis results revealed that Proteobacteria, Actinobacteria, Firmicutes, and Bacteroidetes were the main components at phylum level in ovarian tissue. Notably, the microbial composition of ovarian cancer tissue significantly diverged from that of normal ovarian tissue e, exhibiting markedly lower alpha diversity and distinct beta diversity. Besides, pathogenic microorganisms Achromobacter xylosoxidans and Enterobacter hormaechei were enriched in cancer tissue. Host genes associated with these pathogens were enriched in key pathways including “JAK-STAT signaling pathway”, “Transcriptional misregulation in cancer”, and “Th1 and Th2 cell differentiation”, suggesting their role in ovarian cancer progression through microbial dysbiosis and immune response interaction. ConclusionAbundance of pathogenic microorganisms in ovarian cancer tissue could modulate the expression of host genes, consequently impacting cancer-related signaling pathways and fostering cancer progression.

Inhibitory Effects of Soluble Dietary Fiber from Foxtail Millet on Colorectal Cancer by the Restoration of Gut Microbiota.

Colorectal cancer (CRC) is a common malignant tumor that occurs in the colon. Gut microbiota is a complex ecosystem that plays an important role in the pathogenesis of CRC. Our previous studies showed that the soluble dietary fiber of foxtail millet (FMB-SDF) exhibited significant antitumor activity in vitro. The present study evaluated the anticancer potential of FMB-SDF in the azoxymethane (AOM)- and dextran sodium sulfate (DSS)-induced mouse CRC models. The results showed that FMB-SDF could significantly alleviate colon cancer symptoms in mice. Further, we found that FMB-SDF consumption significantly altered gut microbiota diversity and the overall structure and regulated the abundance of some microorganisms in CRC mice. Meanwhile, KEGG pathway enrichment showed that FMB-SDF can also alleviate the occurrence of colon cancer in mice by regulating certain cancer-related signaling pathways. In conclusion, our findings may provide a novel approach for the prevention and biotherapy of CRC.

Abstract PO1-13-05: Genomic characteristics and its therapeutic implications in breast cancer patients with detectable molecular residual disease

Abstract Background: Breast cancer constitutes approximately 30% of cancers in women, with a mortality-to-incidence ratio of 15%. While early and middle-stage breast cancer patients can undergo radical surgical resection, postoperative recurrence remains a significant challenge for clinicians. Molecular residual disease (MRD) is the main cause of postoperative recurrence of breast cancer. However, the baseline tumor genomic characteristics and therapeutic implications of breast cancer patients with detectable MRD are still unknown. Methods: In this study, we enrolled 80 patients with breast cancer who underwent next-generation sequencing (NGS)-based genetic testing of 1,021 cancer-related genes performed on baseline tumor and postoperative plasma from Army Specialty Medical Center between June 2017 and January 2023. The criteria for the patient’s enrollment were: (1) pathological diagnosis as breast cancer, (2) no distal metastasis, (3) radical surgery and R0 resection, (4) 1021 cancer-related gene testing of the tumor, (5) plasma MRD testing performed within 3 months after surgery and before adjuvant therapy. Results: A total of 18 patients in our cohort had detectable MRD. Baseline clinical characteristics found that patients with higher clinical stages were more likely to have detectable MRD. Analysis of single nucleotide variations and small insertions/deletions in baseline tumors showed that somatic mutations in MAP3K1, ATM, FLT1, GNAS, POLD1, SPEN, and WWP2 were significantly enriched in patients with postoperative MRD. Oncogenic signaling pathway analysis revealed that alteration of the Cell cycle pathway was more likely to occur in patients with detectable MRD (p=0.0125). Mutational signature analysis showed that defective DNA mismatch repair and activation-induced cytidine deaminase (AID) mediated somatic hypermutation (SHM) were associated with detectable MRD. Somatic copy number variation analysis found that amplification of CDKN2A, HOXB13, PPM1D, MPL, and VHL was significantly enriched in patients with detectable MRD. According to the OncoKB database, 77.8% (14/18) of patients with detectable MRD had U.S. Food and Drug Administration-approved mutational biomarkers and targeted therapy. Conclusion: For the first time, our study reports genomic characteristics of breast cancer patients with detectable MRD. The tumor biology is probably the main driver of detectable MRD. We also found the vast majority of patients with detectable MRD have the opportunity to access targeted therapy. Keywords: Breast cancer, MRD, Genomic character, Cell cycle pathway, Defective DNA mismatch repair, Activation-induced cytidine deaminase Figure 1. Somatic mutational landscape and clinical actionability of breast cancer patients with detectable MRD A. Oncoplot of top 20 genes altered in patients with detectable MRD. B. Forest plot of different mutant genes between patients with detectable and undetectable MRD. OR means odds ratio. Inf means infinity. ＊ means P-value < 0.05. C. The ten cancer-related signaling pathways were affected by somatic mutations both in patients with detectable and undetectable MRD. ＊ means P-value < 0.05. D&E. The fraction and etiology of each signature in patients with detectable (D) or undetectable (E) MRD. F. The number of patients with detectable MRD in different actionable alterations. gBRCA1 means germline BRCA1 mutation. sBRCA1 means somatic BRCA1 mutation. sBRCA2 means somatic BRCA2 mutation. TMB-H means TMB-High. Citation Format: Xu Yan, Shu Zhang, Lu zhou, Yan Jiang, Jing Xu, Gang Zhang, Lu Shen. Genomic characteristics and its therapeutic implications in breast cancer patients with detectable molecular residual disease [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-13-05.

Network Pharmacology, Molecular Docking, and Molecular Dynamics Simulation Analysis Reveal Insights into the Molecular Mechanism of Cordia myxa in the Treatment of Liver Cancer.

Traditional treatments of cancer have faced various challenges, including toxicity, medication resistance, and financial burdens. On the other hand, bioactive phytochemicals employed in complementary alternative medicine have recently gained interest due to their ability to control a wide range of molecular pathways while being less harmful. As a result, we used a network pharmacology approach to study the possible regulatory mechanisms of active constituents of Cordia myxa for the treatment of liver cancer (LC). Active constituents were retrieved from the IMPPAT database and the literature review, and their targets were retrieved from the STITCH and Swiss Target Prediction databases. LC-related targets were retrieved from expression datasets (GSE39791, GSE76427, GSE22058, GSE87630, and GSE112790) through gene expression omnibus (GEO). The DAVID Gene Ontology (GO) database was used to annotate target proteins, while the Kyoto Encyclopedia and Genome Database (KEGG) was used to analyze signaling pathway enrichment. STRING and Cytoscape were used to create protein-protein interaction networks (PPI), while the degree scoring algorithm of CytoHubba was used to identify hub genes. The GEPIA2 server was used for survival analysis, and PyRx was used for molecular docking analysis. Survival and network analysis revealed that five genes named heat shot protein 90 AA1 (HSP90AA1), estrogen receptor 1 (ESR1), cytochrome P450 3A4 (CYP3A4), cyclin-dependent kinase 1 (CDK1), and matrix metalloproteinase-9 (MMP9) are linked with the survival of LC patients. Finally, we conclude that four extremely active ingredients, namely cosmosiin, rosmarinic acid, quercetin, and rubinin influence the expression of HSP90AA1, which may serve as a potential therapeutic target for LC. These results were further validated by molecular dynamics simulation analysis, which predicted the complexes with highly stable dynamics. The residues of the targeted protein showed a highly stable nature except for the N-terminal domain without affecting the drug binding. An integrated network pharmacology and docking study demonstrated that C. myxa had a promising preventative effect on LC by working on cancer-related signaling pathways.

Network pharmacology, single gene survival analysis and molecular docking to study the mechanism of Sotetsuflavone in the treatment of pancreatic cancer

Pancreatic cancer is a highly lethal cancer with limited treatment options. The number of pancreatic cancer patients is increasing rapidly worldwide. Many natural products have been shown to have anticancer activity in a range of studies. Sotetsuflavone is derived from Cycas revoluta Thunb. and exhibits anticancer activity. The present study incorporates network pharmacology, single gene survival analysis, gene expression analysis and molecular docking to reveal the mechanism of Sotetsuflavone in the treatment of pancreatic cancer. We have acquired 31 hub targets for the treatment of pancreatic cancer by Sotetsuflavone, namely ABCB1, AURKA, CDK1, and so on. Kaplan-Meier survival analyses demonstrated that ABCB1, AURKA, CDK1, HDAC6, MET, and MMP3 are promising hub targets that can be used as biomarkers for pancreatic cancer diagnosis and prognosis. These hub targets are highly expressed in pancreatic cancer tissues compared to normal tissues. The molecular docking results showed a strong binding capacity of Sotetsuflavone to these hub targets. In summary, it is proposed that Sotetsuflavone is a new anticancer drug, which can regulate pancreatic cancer-related signalling pathways by inhibiting the activities of ABCB1, AURKA, CDK1, HDAC6, MET, and MMP3, which are hub targets with up-regulated expression in pancreatic cancer tissues, in order to treat pancreatic cancer. However, it also requires a series of in vivo and in vitro studies to ensure its safety and efficacy.

High N6-methyladenosine-activated TCEAL8 mRNA is a novel pancreatic cancer marker.

N6-methyladenosine (m6A) is an RNA modification involved in RNA processing and widely found in transcripts. In cancer cells, m6A is upregulated, contributing to their malignant transformation. In this study, we analyzed gene expression and m6A modification in cancer tissues, ducts, and acinar cells derived from pancreatic cancer patients using MeRIP-seq. We found that dozens of RNAs highly modified by m6A were detected in cancer tissues compared with ducts and acinar cells. Among them, the m6A-activated mRNA TCEAL8 was observed, for the first time, as a potential marker gene in pancreatic cancer. Spatially resolved transcriptomic analysis showed that TCEAL8 was highly expressed in specific cells, and activation of cancer-related signaling pathways was observed relative to TCEAL8-negative cells. Furthermore, among TCEAL8-positive cells, the cells expressing the m6A-modifying enzyme gene METTL3 showed co-activation of Notch and mTOR signaling, also known to be involved in cancer metastasis. Overall, these results suggest that m6A-activated TCEAL8 is a novel marker gene involved in the malignant transformation of pancreatic cancer.

Antitumoral Activity of the Universal Methyl Donor S-Adenosylmethionine in Glioblastoma Cells.

Glioblastoma (GBM), the most frequent and lethal brain cancer in adults, is characterized by short survival times and high mortality rates. Due to the resistance of GBM cells to conventional therapeutic treatments, scientific interest is focusing on the search for alternative and efficient adjuvant treatments. S-Adenosylmethionine (AdoMet), the well-studied physiological methyl donor, has emerged as a promising anticancer compound and a modulator of multiple cancer-related signaling pathways. We report here for the first time that AdoMet selectively inhibited the viability and proliferation of U87MG, U343MG, and U251MG GBM cells. In these cell lines, AdoMet induced S and G2/M cell cycle arrest and apoptosis and downregulated the expression and activation of proteins involved in homologous recombination DNA repair, including RAD51, BRCA1, and Chk1. Furthermore, AdoMet was able to maintain DNA in a damaged state, as indicated by the increased γH2AX/H2AX ratio. AdoMet promoted mitotic catastrophe through inhibiting Aurora B kinase expression, phosphorylation, and localization causing GBM cells to undergo mitotic catastrophe-induced death. Finally, AdoMet inhibited DNA repair and induced cell cycle arrest, apoptosis, and mitotic catastrophe in patient-derived GBM cells. In light of these results, AdoMet could be considered a potential adjuvant in GBM therapy.

Exploring the Mechanism of Echinatin against Cervical Cancer Based on Network Pharmacology

Cervical cancer significantly impacts women’s health due to its high mortality rate and increasing prevalence among younger individuals, thereby posing a severe threat. Echinatin is the primary active component of licorice in traditional Chinese medicine. However, studies on its use in cervical cancer treatment are limited. In our study, 198 targets of Echinatin were identified by some databases. Among these, 40 core targets related to cervical cancer were selected. Enrichment analyses revealed that Echinatin operated through genes associated with cell cycle, apoptosis, senescence, and various cancer-related signaling pathways. Differential expression of intersecting targets was verified in the GEO database, and molecular docking also indicated a strong binding capacity between active compounds and identified targets. Moreover, the results of western blot provided further evidence at the protein level. Echinatin hindered the proliferation, migration, and invasion of HeLa and SiHa cells while increasing their apoptosis. This study predicted the potential targets and beneficial effects of Echinatin in cervical cancer treatment, which provides a new avenue for further research into the molecular mechanisms underlying the role of Echinatin in cervical cancer treatment.

Potential value of expression of receptor accessory protein 4 for evaluating the prognosis of lower-grade glioma patients.

REEP4 is involved in the regulation of the biological process of mitosis. Lower grade glioma (LGG), as a malignant tumor, is accompanied by abnormalities in mitosis, but there have been no reports of REEP4 so far. We collected transcriptome data, DNA methylation data and the clinical characteristics of thousands of patients with LGG. Various big data analysis methods and molecular biology experiments were employed to reveal the impact of REEP4 on the pathological process of LGG. It was found that the expression of REEP4 was significantly elevated and negatively regulated by its methylation site. Therefore, both the high expression of REEP4 and low methylation state of cg16311504 showed that the patients are correlated with lower patient survival rate. In addition, high REEP4 expression participates in the regulation of various cancer-related cellular signaling pathways, such as the cell cycle, MAPK signaling pathway, NOD-like receptor signaling pathway, etc. More importantly, the level of immune cell infiltration significantly increased in the high expression group of REEP4 in the LGG tumor microenvironment and REEP4 has a high positive correlation with PD-L1 and other immune checkpoints. In brief, this study is the first to introduce REEP4 in malignant tumors, which can be used as an independent risk factor that participates in the malignant process of LGG. More importantly, REEP4 has the potential to become a new star in the field of anti-tumor treatment.

Abstract 2887: Impact of ionizing radiation on the energy metabolism of normal and tumor cells in the brain

Abstract This study aims to analyze irradiation-induced metabolic alterations in glioblastoma cells and astrocytes to enhance understanding of cellular responses and facilitate the development of more effective radiotherapeutic strategies. Metabolic reprogramming of tumor cells is considered one of the hallmarks of cancer. Studies have shown that abnormal activation of oncogenes and cancer-related signaling pathways, as well as the inactivation of tumor suppressor genes can induce metabolic reprogramming. These metabolic aberrations facilitate rapid proliferation, continuous growth, invasion, metastasis, and immune evasion. Following the application of radiotherapy, the activity of several metabolic pathways significantly changes, potentially leading to the development of radioresistance. However, a differential effect of ionizing radiation on tumor and normal cells is not well understood. Hence, quantifying irradiation-induced metabolic changes in glioblastoma cells and astrocytes is of high importance. We have developed a novel approach to measure dynamic metabolite changes upon irradiation, in vivo, in situ, and in real-time. This tool combines imaging of recently developed fluorescent biosensors via 2-photon microscopy and millimeter-scaled irradiation. Murine glioblastoma cells and astrocytes were generated with stable expression of biosensors for the quantification of intracellular concentrations of various metabolites, including lactate. Cytosolic concentrations of lactate depend on the balance between glycolytic production, mitochondrial consumption of pyruvate and lactate, and the exchange with the extracellular lactate pool via monocarboxylate transporters. Many cancer cells are characterized by an excessive conversion of glucose to lactate even under normoxic conditions. We quantified this so-called Warburg Effect in glioblastoma cells and astrocytes as the ratio between basal lactate production and the lactate increase after OXPHOS inhibition and determined irradiation-induced metabolic changes on the single-cell level. Complementarily, we used metabolomics and Seahorse analyses before and after irradiation. Our data indicate that irradiation increases oxygen consumption in glioblastoma cells but not in astrocytes. Also, we established an orthotopic glioblastoma mouse model via injection of sensor-expressing tumor cells for in vivo measurements of metabolite changes during and after irradiation. These data are expected to significantly advance the understanding of the cellular response to irradiation on a molecular level in situ and in real-time. This knowledge will help to develop combination treatments and increase the efficiency of radiotherapy. Citation Format: Marvin Kreuzer, Pascal Imseng, Bruno Weber, Martin Pruschy. Impact of ionizing radiation on the energy metabolism of normal and tumor cells in the brain [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2887.

Lycium barbarum polysaccharide reverses drug resistance in oxaliplatin-resistant colon cancer cells by inhibiting PI3K/AKT-dependent phosphomannose isomerase.

Objective: Here, we aimed to explore the effect of LBP in combination with Oxaliplatin (OXA) on reversing drug resistance in colon cancer cells through in vitro and in vivo experiments. We also aimed to explore the possible mechanism underlying this effect. Finally, we aimed to determine potential targets of Lycium barbarum polysaccharide (LBP) in colon cancer (CC) through network pharmacology and molecular docking. Methods: The invasion ability of colon cancer cells was assessed using the invasion assay. The migration ability of these cells was assessed using the migration assay and wound healing assay. Cell cycle analysis was carried out using flow cytometry. The expression levels of phosphomannose isomerase (PMI) and ATP-binding cassette transport protein of G2 (ABCG2) proteins were determined using immunofluorescence and western blotting. The expression levels of phosphatidylinositol3-kinase (PI3K), protein kinase B (AKT), B-cell lymphoma 2 (Bcl-2), and BCL2-Associated X (Bax) were determined using western blotting. Forty BALB/c nude mice purchased from Weitong Lihua, Beijing, for the in vivo analyses. The mice were randomly divided into eight groups. They were administered HCT116 and HCT116-OXR cells to prepare colon cancer xenograft models and then treated with PBS, LBP (50mg/kg), OXA (10mg/kg), or LBP + OXA (50mg/kg + 10mg/kg). The tumor weight and volume of treated model mice were measured, and organ toxicity was evaluated using hematoxylin and eosin staining. The expression levels of PMI, ABCG2, PI3K, and AKT proteins were then assessed using immunohistochemistry. Moreover, PMI and ABCG2 expression levels were analyzed using immunofluorescence and western blotting. The active components and possible targets of LBP in colon cancer were explored using in silico analysis. GeneCards was used to identify CC targets, and an online Venn analysis tool was used to determine intersection targets between these and LBP active components. The PPI network for intersection target protein interactions and the PPI network for interactions between the intersection target proteins and PMI was built using STRING and Cytoscape. To obtain putative targets of LBP in CC, we performed GO function enrichment and KEGG pathway enrichment analyses. Results: Compared with the HCT116-OXR blank treatment group, both invasion and migration abilities of HCT116-OXR cells were inhibited in the LBP + OXA (2.5mg/mL LBP, 10μΜ OXA) group (p < 0.05). Cells in the LBP + OXA (2.5mg/mL LBP, 10μΜ OXA) group were found to arrest in the G1 phase of the cell cycle. Knockdown of PMI was found to downregulate PI3K, AKT, and Bcl-2 (p < 0.05), while it was found to upregulate Bax (p < 0.05). After treatment with L. barbarum polysaccharide, 40 colon cancer subcutaneous tumor models showed a decrease in tumor size. There was no difference in the liver index after LBP treatment (p > 0.05). However, the spleen index decreased in the OXA and LBP + OXA groups (p < 0.05), possibly as a side effect of oxaliplatin. Immunohistochemistry, immunofluorescence, and western blotting showed that LBP + OXA treatment decreased PMI and ABCG2 expression levels (p < 0.05). Moreover, immunohistochemistry showed that LBP + OXA treatment decreased the expression levels of PI3K and AKT (p < 0.05). Network pharmacology analysis revealed 45 active LBP components, including carotenoids, phenylpropanoids, quercetin, xanthophylls, and other polyphenols. It also revealed 146 therapeutic targets of LBP, including AKT, SRC, EGFR, HRAS, STAT3, and MAPK3. KEGG pathway enrichment analysis showed that the LBP target proteins were enriched in pathways, including cancer-related signaling pathways, PI3K/AKT signaling pathway, and IL-17 signaling pathways. Finally, molecular docking experiments revealed that the active LBP components bind well with ABCG2 and PMI. conclusion: Our in vitro experiments showed that PMI knockdown downregulated PI3K, AKT, and Bcl-2 and upregulated Bax. This finding confirms that PMI plays a role in drug resistance by regulating the PI3K/AKT pathway and lays a foundation to study the mechanism underlying the reversal of colon cancer cell drug resistance by the combination of LBP and OXA. Our in vivo experiments showed that LBP combined with oxaliplatin could inhibit tumor growth. LBP showed no hepatic or splenic toxicity. LBP combined with oxaliplatin could downregulate the expression levels of PMI, ABCG2, PI3K, and AKT; it may thus have positive significance for the treatment of advanced metastatic colon cancer. Our network pharmacology analysis revealed the core targets of LBP in the treatment of CC as well as the pathways they are enriched in. It further verified the results of our in vitro and in vivo experiments, showing the involvement of multi-component, multi-target, and multi-pathway synergism in the drug-reversing effect of LBP in CC. Overall, the findings of the present study provide new avenues for the future clinical treatment of CC.

Redox proteomics of PANC-1 cells reveals the significance of HIF-1 signaling protein oxidation in pancreatic ductal adenocarcinoma pathogenesis

BackgroundProtein cysteine oxidation is substantially involved in various biological and pathogenic processes, but its implications in pancreatic cancer development remains poorly understood.Methods and resultsIn this study, we performed a global characterization of protein oxidation targets in PDAC cells through iodoTMT-based quantitative proteomics, which identified over 4300 oxidized cysteine sites in more than 2100 proteins in HPDE6c7 and PANC-1 cells. Among them, 1715 cysteine residues were shown to be differentially oxidized between HPDE6c7 and PANC-1 cells. Also, charged amino acids including aspartate, glutamate and lysine were significantly overrepresented in flanking sequences of oxidized cysteines. Differentially oxidized proteins in PANC-1 cells were enriched in multiple cancer-related biological processes and signaling pathways. Specifically, the HIF-1 signaling proteins exhibited significant oxidation alterations in PANC-1 cells, and the reduced PHD2 oxidation in human PDAC tissues was correlated with lower survival time in pancreatic cancer patients.ConclusionThese investigations provided new insights into protein oxidation-regulated signaling and biological processes during PDAC pathogenesis, which might be further explored for pancreatic cancer diagnosis and treatment.

A novel CD8+ T cell-related gene signature as a prognostic biomarker in hepatocellular carcinoma.

CD8+ T cells have great roles in tumor suppression and elimination of various tumors including hepatocellular carcinoma (HCC). Nonetheless, potential prognostic roles of CD8+ T cell-related genes (CD8Gs) in HCC remains unknown. In our study, 416 CD8Gs were identified in HCC, which were enriched in inflammatory and immune signaling pathways. Using The Cancer Genome Atlas dataset, a 5-CD8Gs risk model (KLRB1, FYN, IL2RG, FCER1G, and DGKZ) was constructed, which was verified in International Cancer Genome Consortium and gene expression omnibus datasets. Furthermore, we found that overall survival was independently correlated with the CD8Gs signature, and it was associated with immune- and cancer-related signaling pathways and immune cells infiltration. Finally, drug sensitivity data indicated that 10 chemotherapeutic drugs held promise as therapeutics for HCC patients with high-risk. In conclusion, multi-databases analysis showed that 5-CD8Gs and their signature could be an indicator to predict candidate drugs for HCC therapy.

Potential diagnostic value of serum microRNAs for 19 cancer types: a meta-analysis of bioinformatics data

Cancer is the second most common cause of mortality worldwide, accounting for almost 10 million deaths in 2020. These deaths were partly due to delayed diagnosis that led to deferred treatment. Therefore, new diagnostic methods are necessary to enhance the accuracy of noninvasive cancer detection. The present study developed a microRNA (miRNA)-based serum diagnostic marker for detecting a wide range of cancers. The study involved 61,019 serum samples from 19 different cancer types. A miRNA prediction model was established through bioinformatics analysis of serum samples from various cancer pathologies and qRT-PCR results from studies in PubMed aligned to the analysis criteria. R software v.4.1.1 with the limma data analysis package was used for single gene expression series data series, and batchNormalize and robustRankAggreg were used to predict the changes in miRNA expression in multiple datasets. GO and KEGG analyses showed that these miRNAs play a role in cancer-related biological signaling pathways. Finally, the diagnostic capability of these miRNA biomarkers was assessed using area under the curve analysis. The study predicted that 7 miRNAs were upregulated and 10 miRNAs were downregulated in 19 different types of cancer. Some miRNAs showed significant differential expression in a specific cancer type. Additionally, downstream genes regulated by miRNAs focused on many cancer-related molecular signaling pathways. In this review, we summarize the current understanding of miRNAs in various cancers, with a particular focus on their potential as future noninvasive diagnostic biomarkers. The emphasis is on their capacity for achieving high accuracy and cost savings compared to conventional biomarkers. Communicated by Ramaswamy H. Sarma

Region-specific DNA hydroxymethylation along the malignant progression of IDH-mutant gliomas.

The majority of low-grade isocitrate dehydrogenase-mutant (IDHmt) gliomas undergo malignant progression (MP), but their underlying mechanism remains unclear. IDHmt gliomas exhibit global DNA methylation, and our previous report suggested that MP could be partly attributed to passive demethylation caused by accelerated cell cycles. However, during MP, there is also active demethylation mediated by ten-eleven translocation, such as DNA hydroxymethylation. Hydroxymethylation is reported to potentially contribute to gene expression regulation, but its role in MP remains under investigation. Therefore, we conducted a comprehensive analysis of hydroxymethylation during MP of IDHmt astrocytoma. Five primary/malignantly progressed IDHmt astrocytoma pairs were analyzed with oxidative bisulfite and the Infinium EPIC methylation array, detecting 5-hydroxymethyl cytosine at over 850,000 locations for region-specific hydroxymethylation assessment. Notably, we observed significant sharing of hydroxymethylated genomic regions during MP across the samples. Hydroxymethylated CpGs were enriched in open sea and intergenic regions (p < 0.001), and genes undergoing hydroxymethylation were significantly associated with cancer-related signaling pathways. RNA sequencing data integration identified 91 genes with significant positive/negative hydroxymethylation-expression correlations. Functional analysis suggested that positively correlated genes are involved in cell-cycle promotion, while negatively correlated ones are associated with antineoplastic functions. Analyses of The Cancer Genome Atlas clinical data on glioma were in line with these findings. Motif-enrichment analysis suggested the potential involvement of the transcription factor KLF4 in hydroxymethylation-based gene regulation. Our findings shed light on the significance of region-specific DNA hydroxymethylation in glioma MP and suggest its potential role in cancer-related gene expression and IDHmt glioma malignancy.

The Mechanism of Fuzheng Xiaoji Decoction in the Treatment of Non-Small Cell Lung Cancer Based on Network Pharmacology and Molecular Docking Technology

With approximately 2 million lung cancer patients and 1.8 million deaths per year, the world ranks among the highest incidence rates in the world. Non-small cell lung cancer (NSCLC) accounts for up to 85% of lung cancer patients and has a poor prognosis. To investigate the mechanism of Fuzheng Xiaoji Decoction in the treatment of NSCLC. Fuzheng Xiaoji Decoction, consisting of ten traditional Chinese medicines, was analyzed using the TCMSP database to identify its active ingredients and their corresponding targets. Simultaneously, NSCLC-related targets were obtained from GeneCards. The study aimed to predict Fuzheng Xiaoji Decoction’s targets in treating NSCLC and establish a network diagram illustrating the relationship between traditional Chinese medicine, active ingredients, targets, and the disease. The STRING platform was used to construct a PPI network, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and Gene Ontology (GO) function enrichment analysis were performed on the intersecting targets. Based on the interaction network analysis, the core target proteins, represented by the TOP5 nodes with the highest degrees, were selected. These core targets were then subjected to molecular docking with the top five key active components of Fuzheng Xiaoji Decoction to construct the docking model. We identified 132 active components in Fuzheng Xiaoji Decoction and picked the TOP 5 proteins from the PPI network as the primary active elements in Fuzheng Xiaoji Decoction for NSCLC treatment, including kaempferol, quercetin, ingenin, luteolin, and osterol, involving a total of 209 targets. The top five core targets obtained by PPI network were AKT1, TP53, STAT3, TNF and IL6. A total of 7879 targets related to NSCLC were retrieved, and 164 drug-disease intersection targets were identified. GO functional analysis results showed that 164 gene sets were involved in 377 biological processes, mainly involved in 79 processes related to cell composition and 46 molecular function expression processes. KEGG enrichment results showed 132 significantly enriched signaling pathways. Molecular docking findings indicated favorable binding interactions between the receptor and ligand, suggesting a strong affinity between the key NSCLC treatment protein and the primary active ingredient in Fuzheng Xiaoji Decoction. The potential mechanism of Fuzheng Xiaoji Decoction in NSCLC treatment may involve flavonoids, triterpenoids and other compounds regulate lung cancer-related signaling pathways by acting on AKT1, TP53, STAT3 and other proteins to affect the expression of P53, IL6 and other cytokines. This may help regulate immune function and inhibit tumor cell proliferation.

Diagnostic value of long noncoding RNA SNHG15 in gastric cancer: in vitro and in silico studies

LncRNA SNHG15 has been recognized as the main factor in the progression of various cancer types. However, the underlying mechanisms are not well clarified. This research aimed to explore the diagnostic potential of SNHG15 in gastric cancer (GC) patients and also the effects of SNHG15-miRNA-mRNA network in GC pathobiology. The expression level of SNHG15 in GC tissues and adjacent normal tissues (ANTs) was evaluated by qRT-PCR and also considered in relation to clinicopathologic factors. The ROC curve was explored to consider the specificity and sensitivity of SNHG15. Gene ontology functional annotation and KEGG pathway analysis were performed in order to predict the effects of SNHG15-miRNA-mRNA network in GC pathobiology. SNHG15 was overexpressed in GC tissues compared to ANTs (fold change= 3.87 and P-value = 0.0022). The SNHG15 expression level was not significantly associated with clinicopathologic factors. ROC curve indicated the specificity of 63.51 and sensitivity of 79.73 and the AUC of 0.744 (P-value < 0.0001). Further gene network analysis revealed that SNHG15 interacts with has-miR-613, has-miR-542-3p, and has-miR-1236-3p, and may be involved in the GC pathobiology by affecting the EGFR tyrosine kinase inhibitor resistance, HIF-1 signaling pathway, and VEGF signaling pathway. It can be concluded that SNHG15 may be a diagnostic factor in GC and may contribute in a variety of cancer-related signaling pathways.

A26 BIOLOGICAL CHARACTERIZATION OF THE INTERACTION BETWEEN SIRT1 AND HNF4Α2 IN INTESTINAL EPITHELIAL CELLS

Abstract Background Colorectal cancer (CRC) is the third most common cancer in Canada. HNF4A locus is amplified in CRC, while additional reports suggest that hepatocyte nuclear factor 4 alpha (HNF4α) is associated with increased proliferation and disease development. This dual role as a tumor suppressor or oncoprotein might be due to the production of 12 spliced isoforms with structural differences and variable tissue expression. It suggests distinct functions for each isoform according to their specific interaction complexes. However, little is known about the nature of these protein complexes and their biological functions during intestinal physiopathology. Recently, using a BioID2 and mass spectrometry approach, our laboratory showed a possible interaction between HNF4α2 and sirtuin 1 (SIRT1) in intestinal epithelial cells from a colon carcinoma (HCT116). HNF4α2 is one of the most potent isoforms in the control of transcriptional activity of multiple intestinal epithelial genes related to regulating cell death, angiogenesis, apoptosis, response to injury, and response to drugs, among others. Aims To characterize the possible interaction between HNF4α2 and SIRT1 in intestinal epithelial cells at the biological level. Methods We performed SIRT1 knockdown using shRNAs in HCT116 cells expressing the HNF4α2 isoform in an inducible manner by doxycycline (DOX). Subsequently, we conducted a proximity ligation assay to validate the interaction between HNF4α2 and SIRT1 in HCT116 cells. Additionally, protein purification was performed to validate their physical interaction through EMSA. A transcriptome analysis was carried out using RNA-seq to define the biological processes involved in the interaction between the two proteins. Results We observed an interaction signal between HNF4α2 and SIRT1 in HCT116 cells, similar to that observed for IRFBB2, whose interaction with HNF4α2 has been previously validated. This signal decreased when cells were treated with shRNA for SIRT1 and was not observed in cells not induced by DOX. EMSA analyses revealed a direct interaction signal between HNF4α2 and SIRT1. RNA-seq analyses showed a loss in the regulation of over 95% of genes targeted by HNF4α2 when cells were treated with shRNA for SIRT1. An enrichment analysis for GO annotations using ShyniGO v. 0.77 software showed an enrichment of genes for biological processes such as apoptosis, inflammation, cell migration, cell invasion, cell death, and various cancer-related signaling pathways. Conclusions SIRT1 displays physical interaction with HNF4α2 and significantly affects the transcriptional regulation of this isoform in the context of epithelial cells. Their interaction is involved in processes related to intestinal inflammation and cancer progression. Pharmacological targeting of SIRT1 could represent a viable strategy in treating CRC and other intestinal diseases Funding Agencies CIHRNSERC

Therapeutic Targeting of Hypoxia-Inducible Factors in Cancer.

In the realm of cancer therapeutics, targeting the hypoxia-inducible factor (HIF) pathway has emerged as a promising strategy. This study delves into the intricate web of HIF-associated mechanisms, exploring avenues for future anticancer therapies. Framing the investigation within the broader context of cancer progression and hypoxia response, this article aims to decipher the pivotal role played by HIF in regulating genes influencing angiogenesis, cell proliferation, and glucose metabolism. Employing diverse approaches such as HIF inhibitors, anti-angiogenic therapies, and hypoxia-activated prodrugs, the research methodologically intervenes at different nodes of the HIF pathway. Findings showcase the efficacy of agents like EZN-2968, Minnelide, and Acriflavine in modulating HIF-1α protein synthesis and destabilizing HIF-1, providing preliminary proof of HIF-1α mRNA modulation and antitumor activity. However, challenges, including toxicity, necessitate continued exploration and development, as exemplified by ongoing clinical trials. This article concludes by emphasizing the potential of targeted HIF therapies in disrupting cancer-related signaling pathways.