Subcutaneous Transplantation Tumor Research Articles

PPAR-γ/NF-kB/AQP3 axis in M2 macrophage orchestrates lung adenocarcinoma progression by upregulating IL-6

Aquaporin 3 (AQP3), which is mostly expressed in pulmonary epithelial cells, was linked to lung adenocarcinoma (LUAD). However, the underlying functions and mechanisms of AQP3 in the tumor microenvironment (TME) of LUAD have not been elucidated. Single-cell RNA sequencing (scRNA-seq) was used to study the composition, lineage, and functional states of TME-infiltrating immune cells and discover AQP3-expressing subpopulations in five LUAD patients. Then the identifications of its function on TME were examined in vitro and in vivo. AQP3 was associated with TNM stages and lymph node metastasis of LUAD patients. We classified inter- and intra-tumor diversity of LUAD into twelve subpopulations using scRNA-seq analyses. The analysis showed AQP3 was mainly enriched in subpopulations of M2 macrophages. Importantly, mechanistic investigations indicated that AQP3 promoted M2 macrophage polarization by the PPAR-γ/NF-κB axis, which affected tumor growth and migration via modulating IL-6 production. Mixed subcutaneous transplanted tumor mice and Aqp3 knockout mice models were further utilized, and revealed that AQP3 played a critical role in mediating M2 macrophage polarization, modulating glucose metabolism in tumors, and regulating both upstream and downstream pathways. Overall, our study demonstrated that AQP3 could regulate the proliferation, migration, and glycometabolism of tumor cells by modulating M2 macrophages polarization through the PPAR-γ/NF-κB axis and IL-6/IL-6R signaling pathway, providing new insight into the early detection and potential therapeutic target of LUAD.

Urolithin C suppresses colorectal cancer progression via the AKT/mTOR pathway

Urolithin families are gut-microbial metabolites of ellagic acid (EA). Although urolithin A (UA) and urolithin B (UB) were reported to have antiproliferative activities in cancer cells, the role and related mechanisms of urolithin C (UC) in colorectal cancer (CRC) have not yet been clarified. In this study, we assess the antitumor activities of UC in vitro and in vivo and further explore the underlying mechanisms in CRC cell lines. We found that UC inhibited the proliferation and migration of CRC cells, induced apoptosis, and arrested the cell cycle at the G2/M phase in vitro, and UC inhibited tumor growth in a subcutaneous transplantation tumor model in vivo. Mechanically, UC blocked the activation of the AKT/mTOR signaling pathway by decreasing the expression of Y-box binding protein 1(YBX1). The AKT agonist SC79 could reverse the suppression of cell proliferation in UC-treated CRC cells. In conclusion, our research revealed that UC could prevent the progression of CRC by blocking AKT/mTOR signaling, suggesting that it may have potential therapeutic values.Graphical

FIGNL1 Promotes Hepatocellular Carcinoma Formation via Remodeling ECM-receptor Interaction Pathway Mediated by HMMR.

The development of novel biomarkers is crucial for the treatment of HCC. In this study, we investigated a new molecular therapeutic target for HCC. Fidgetin-like 1 (FIGNL1) has been reported to play a vital role in lung adenocarcinoma. However, the potential function of FIGNL1 in HCC is still unknown. This study aims to investigate the key regulatory mechanisms of FIGNL1 in the formation of HCC. The regulatory effect of FIGNL1 on HCC was studied by lentivirus infection. In vitro, the effects of FIGNL1 on the proliferation, migration and apoptosis of cells were investigated by CCK8, colony formation assay, transwell and flow cytometry. Meanwhile, the regulation of FIGNL1 on HCC formation in vivo was studied by subcutaneous transplanted tumors. In addition, using transcriptome sequencing technology, we further explored the specific molecular mechanism of FIGNL1 regulating the formation of HCC. Functionally, we demonstrated that FIGNL1 knockdown significantly inhibited HCC cell proliferation, migration and promoted cell apoptosis in vitro. Similarly, the knockdown of FIGNL1 meaningfully weakened hepatocarcinogenesis in nude mice. Transcriptome sequencing revealed that FIGNL1 affected the expression of genes involved in extracellular matrix-receptor (ECM-receptor) interaction pathway, such as hyaluronan mediated motility receptor (HMMR). Further validation found that overexpression of HMMR based on knockdown FIGNL1 can rescue the expression abundance of related genes involved in the ECM-receptor interaction pathway. Our study revealed that FIGNL1 could modulate the ECM-receptor interaction pathway through the regulation of HMMR, thus regulating the formation of HCC.

Apoptosis-induced treatment for ovarian cancer from self-assembled nanocomposites of Fe3O4 nanoparticles and porous organic cages

HypothesisOvarian cancer is one of the most common tumors in gynecology and imposes a heavy burden on patients and their families. Fe3O4 nanoparticles are one of the few nanomaterials approved by the FDA for clinical use, which has been widely used in the diagnosis and treatment of tumor. ExperimentsThe aqueous colloids made from hydrophobic building blocks were fabricated through an oil-in-water emulsion confined assembly strategy. In vitro, the proliferation, migration, invasion of SKOV3 cell after the treatment of iron oxide based assemblies were detected using Test-EdU assay and wound-healing assay. The ovarian cancer of the Nude Mice model was used to investigate the anti-cancer effect of Fe3O4 assemblies and Fe3O4-CC3 co-assemblies in vivo. FindingsThe applied nanocomposites inhibited ovarian cancer cells by CytC/caspase-3 pathway. The results in vivo revealed that treatment of nanocomposites significantly restrained the growth of subcutaneous transplantation tumor in Ovarian Cancer of the Nude Mice model. Our study demonstrated that colloidal nanocomposites assembled from Fe3O4 nanoparticles and porous organic cages inhibits cancer progression and induces apoptosis via caspase pathway. The effect of Fe3O4-CC3 co-assemblies is more profound due to their better contact with the biological matter.

LEPR/FOS/JUNB signaling pathway contributes to chronic restraint stress-induced tumor proliferation

Background & aimsPsychosocial stress has become an unavoidable part of life, which was reported to promote tumor development. Chronic stress significantly promotes the norepinephrine (NE) secretion and the expression of leptin receptor (LEPR), leading to tumor invasion, metastasis, and proliferation. However, the mechanism of chronic stress-induced tumor proliferation remains unclear. MethodsTo reveal the effect of chronic stress on tumor proliferation, subcutaneous tumor models combined with chronic restraint stress (CRS) were established. Combined with the transcript omics database of liver cancer patients, the target pathways were screened and further verified by in vitro experiments. ResultsThe results showed that the CRS with subcutaneous tumor transplantation (CRS + tumor) group exhibited significantly larger tumor sizes than the subcutaneous tumor transplantation (tumor) group. Compared with the tumor group, CRS obviously increased the mRNA levels of LEPR, FOS, and JUNB of tumor tissues in the CRS + tumor group. Furthermore, the treatment with norepinephrine (NE) significantly elevated the survival rate of H22 cells and enhanced the expression of LEPR, FOS, and JUNB in vitro. Silencing LEPR significantly reduced the expression of FOS and JUNB, accompanied by a decrease in H22 cell viability. ConclusionsOur study demonstrated that CRS activates the LEPR–FOS–JUNB signaling pathway by NE, aggravating tumor development. These findings might provide a scientific foundation for investigating the underlying pathological mechanisms of tumors in response to chronic stress.

Abstract 2129: Efficacy and underlying mechanisms of surufatinib combined with PD-1 monoclonal antibody and chemotherapy in pancreatic cancer

Abstract Background: A phase 1b/2 study (NCT05218889) of surufatinib combined with camrelizumab, nab-paclitaxel, and S-1 (NASCA) as first-line therapy for metastatic pancreatic adenocarcinoma has shown promising results, potentially overcoming the limitations of current pancreatic immunotherapies. Animal studies are underway to further explore the synergistic effects and mechanisms of this combined therapeutic approach. Methods: Genetically engineered KPC mouse cells were used to establish a subcutaneous transplantation tumor model in female mice, which were then divided into control group, AG group (nab-paclitaxel + gemcitabine), and NASCA group. Tumor metrics were recorded until euthanasia after 21 days. Blood and tumor samples were analyzed using Flow Cytometry. Results: Compared with the control group, the NASCA and AG groups demonstrated tumor volume reductions of 14.7% (p=0.172) and -0.4% (p=0.972) on day 7, respectively. At day 14, the reductions were 48.8% and 29.9% (p<0.001*, p=0.002*). At day 21, the reductions were 37.4% and 21.5% (p=0.001*, p=0.144). The tumor growth inhibitions (TGIs) for NASCA and AG group were 56.7% (p<0.001*) and 21.2% (p=0.36), respectively. Compare to AG group, the tumor volume reduction in NASCA group were 15.1% (p=0.050*), 26.9% (p=0.001*) and 20.3% (p=0.116), respectively. And the TGIs for NASCA group compare AG group was 45.0% (p=0.020*). Distinguishingly, the NASCA group, when compared to the AG group, had a heightened proportion of M1 macrophages (28.5% vs. 15.5%, p=0.019*), mCD68+ Macrophages (99.7% vs. 99.1%, p=0,014*) and mPD-1+/mCD8+ T cells (24.8% vs. 9.8%, p=0.03*). Furthermore, T-regulatory cells (T-regs) in peripheral blood demonstrated uniformity across the groups, were no statistically. RNA-seq analysis revealed that the NASCA significantly suppressed multiple anti-angiogenic pathways while enhancing the cytokine pathway (p=0.005*), indicating potential immune system activation. Conclusion: The NASCA group outperformed the AG group in controlling tumor growth and early tumor shrinkage, potentially due to macrophage polarization regulation. Further research is vital to understand the benefits of integrating targeted therapy, immunotherapy, and chemotherapy for pancreatic cancer. Citation Format: Nan Zhang, Ru Jia, Guanghai Dai. Efficacy and underlying mechanisms of surufatinib combined with PD-1 monoclonal antibody and chemotherapy in pancreatic cancer [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 2129.

A pH‐Sensitive Smart Monomer Prevents Oral Cancer Progression

Oral squamous cell carcinoma (OSCC) is a prevalent cancer worldwide, and the development of anti‐OSCC materials is urgent. The tumor microenvironment has been identified as a significant characteristic of cancer, with the pHe value in OSCC ranging from 6.56 to 6.97. Given the acidic nature of OSCC, the creation of pH‐sensitive antitumor materials has become a prominent area of research. A pH‐sensitive tertiary amine monomer, dodecylmethylaminoethyl methacrylate (DMAEM), has been previously synthesized. This study aims to evaluate the impact of DMAEM on OSCC. The results demonstrated that DMAEM inhibited the proliferation, migration, and invasion of OSCC cells. Furthermore, it promoted apoptosis and autophagy of OSCC cells, with its anti‐OSCC effect being strengthened in the acidic environment. In a subcutaneous transplantation tumor model, DMAEM inhibited the growth of OSCC and expression of Ki‐67. The analysis of 16S rDNA sequencing data revealed that DMAEM had no significant impact on the Alpha/Beta diversity of the gastrointestinal tract microbiota in mice and had minimal effect on its composition. Overall, this study suggests that DMAEM exhibits pH‐responsive behavior in the acidic tumor microenvironment, effectively inhibiting OSCC without disturbing the gastrointestinal microbiota. These findings highlight the potential of DMAEM for clinical applications in OSCC treatment.

Prognostic marker CXCL5 in glioblastoma polyformis and its mechanism of immune invasion

Glioblastoma multiforme (GBM) is the most aggressive brain cancer with a poor prognosis. Therefore, the correlative molecular markers and molecular mechanisms should be explored to assess the occurrence and treatment of glioma.WB and qPCR assays were used to detect the expression of CXCL5 in human GBM tissues. The relationship between CXCL5 expression and clinicopathological features was evaluated using logistic regression analysis, Wilcoxon symbolic rank test, and Kruskal–Wallis test. Univariate, multivariate Cox regression and Kaplan–Meier methods were used to assess CXCL5 and other prognostic factors of GBM. Gene set enrichment analysis (GSEA) was used to identify pathways associated with CXCL5. The correlation between CXCL5 and tumor immunoinfiltration was investigated using single sample gene set enrichment analysis (ssGSEA) of TCGA data. Cell experiments and mouse subcutaneous transplanted tumor models were used to evaluate the role of CXCL5 in GBM. WB, qPCR, immunofluorescence, and immunohistochemical assays showed that CXCL5 expression was increased in human GBM tissues. Furthermore, high CXCL5 expression was closely related to poor disease-specific survival and overall survival of GBM patients. The ssGSEA suggested that CXCL5 is closely related to the cell cycle and immune response through PPAR signaling pathway. GSEA also showed that CXCL5 expression was positively correlated with macrophage cell infiltration level and negatively correlated with cytotoxic cell infiltration level. CXCL5 may be associated with the prognosis and immunoinfiltration of GBM.

Carvedilol induces pyroptosis through NLRP3-caspase1-ASC inflammasome by nuclear migration of NF-κB in prostate cancer models.

Pyroptosis is an inflammatory type of programmed cell death, and could overcome the drug-resistance induced by anti-apoptotic effect of cancers. Carvedilol (CVL), a β-adrenergic receptors antagonist, has shown anti-inflammatory response and anti-cancer effect. The aim of this study is to investigate whether pyroptosis can be activated by CVL in prostate cancer (PCa). Datasets were used to analyze the expressions of pyroptosis-related proteins. Intracellular morphological change, cell viability, LDH and Il-1β release by cells,, and Hoechst/PI staining were used to detect the occurrence of pyroptosis. Realtime-PCR, western blot, immunofluorescence, and immunohistochemistry (IHC) were used to investigate the expressions of pyroptosis-related proteins. Datasets analyze showed the expressions of NLRP3, Caspase 1, ASC and GSDMD were all decreased in PCa comparing with normal tissues, but without prognostic significance. CVL treatment weakened the viabilities of PCa cells. Cell morphology changing, cytoplasmic vacuole formation, membrane integrity loss, LDH and IL-1β release and PI positive cells increasing were observed. NLRP3, Caspase 1, ASC, GSDMD and N-GSDMD expressions were elevated after CVL treatment, accompanied by a tendency of NF-κB transferring into nucleus. In vivo, CVL inhibited the growth of subcutaneous transplanted tumor. IHC showed CVL increased the expressions of NLRP3, ASC, and GSDMD, and decreased the expression of Ki-67 in transplanted tumor tissues. This study demonstrated that CVL could induce pyroptosis in PCa cells through NLRP3-caspase1-ASC inflammasome by promoting nuclear translocation of NF-κB, which would lay a foundation for the application of adrenergic receptor antagonist in PCa.

Carbon ion irradiation exerts antitumor activity by inducing cGAS-STING activation and immune response in prostate cancer-bearing mice.

As an advanced radiotherapy technique, carbon ion radiotherapy has demonstrated good efficacy and low toxicity for prostate cancer patients, but the radiobiological mechanism of killing tumor cells has not been fully elucidated. This study aims to explore the antitumor effects of carbon ion irradiation (CIR) through investigating the immune response induced by CIR in prostate cancer-bearing mice and the underlying molecular mechanism. We established subcutaneous transplantation tumor models of prostate cancer to evaluate the tumor inhibition effect of CIR. Investigation of immunophenotype alterations were assessed by flow cytometry. Immunofluorescence, western blot, and real-time quantitative PCR was employed to analyze the activation of cGAS-STING pathway. CIR showed more powerful tumor growth control than photon irradiation in immunocompetent syngeneic C57BL/6 mice. CIR exerts antitumor effect by triggering immune response, characterized by increased CD4+ T cells and macrophages in tumor, enhanced CD8+ T cells and T effector memory cells in spleen, improved IFN-γ production of CD8+ tumor-infiltrating lymphocytes, and reduced exhausted T cells in tumor and spleen. Additionally, production of cytoplasmic double-stranded DNA, protein levels of p-TBK1 and p-IRF3 in the cGAS-STING pathway, and gene expression levels of downstream interferon-stimulated genes were significantly increased after CIR in a dose-dependent manner. Treatment of RM1 tumor-bearing mice with the STING inhibitor C-176 impaired the antitumor effect of CIR. The excellent antitumor activity of CIR in immunocompetent prostate cancer-bearing C57BL/6 mice may be attributed to stronger induction of antitumor immune response and higher activation of cGAS-STING pathway.

Transformation of Ginsenosides by Lactiplantibacillus plantarum MB11 Fermentation: Minor Ginsenosides Conversion and Enhancement of Anti-Colorectal Cancer Activity.

The present study aimed to increase the content of minor ginsenosides and enhance the anti-colorectal cancer activity of ginsenosides via biotransformation by Lactiplantibacillus plantarum MB11 screened from fermented foods. A subcutaneous transplantation tumor model of murine colorectal cancer CT26 cells was established in mice to study the anticarcinogenic activities and mechanism of fermented total ginsenosides (FTGs). The results showed that L. plantarum MB11 fermentation increased the content of minor ginsenosides and decreased that of major ginsenosides. FTGs reduced the tumor weight and size compared with the model group. Immunofluorescence and TdT-mediated dUTP nick end labeling (TUNEL) analysis showed that FTGs significantly increase the number of caspase-3 cells in tumor tissue and induce cell apoptosis. Mechanically, FTGs activate AMPK/mTOR autophagy pathway and regulate JAK2/STAT3 and Bax/Bcl-2/caspase-3 apoptosis pathway. Overall, fermentation with L. plantarum MB11 enhanced minor ginsenosides in total ginsenosides, and FTGs induced subcutaneous transplantation tumor autophagy and apoptosis in mice.

CCL5 promotes the proliferation and metastasis of bladder cancer via the JAK2/STAT3 signaling pathway.

Non-muscle invasive bladder cancer (NMIBC) is one of the most common malignant tumors of the urinary system. There is an urgent need for further studies to elucidate the underlying mechanisms of bladder cancer (BC) progression. It has been observed that C-C chemokine ligand 5 (CCL5) and its receptor C-C chemokine receptor type 5 (CCR5) are expressed abnormally and activated in solid tumors and hematological malignancies, which is gaining increasing attention. However, the underlying mechanism of CCL5 in BC remains unclear. The expression levels of CCL5 were analyzed by real-time polymerase chain reaction (RT-PCR) and western blot. Proliferation analysis of cells was carried out using Cell Counting Kit-8 (CCK-8). The assessment of the migration was conducted using a wound-healing assay. A Matrigel-coated transwell chamber was used to test cell invasiveness. A subcutaneous transplantation tumor model and tail vein injection pulmonary metastasis tumor model were used to evaluate the proliferation and metastasis of BC cell in vivo. This study showed that CCL5 promotes proliferative, migratory, and tumor-growing BC cells in vitro and tumor metastasizing BC cells in vivo. Moreover, we found that the tumor-promotive role of CCL5 is dependent on activation of the JAK2/STAT3 signaling pathway. CCL5 may play an oncogenic role in BC and may also serve as a potential diagnostic and prognostic biomarker.

Cinobufagin Induces Apoptosis of DLBCL Cells By Targeting the Inhibition of G6PD Activity

Background:Globally, there are 150,000 new cases of diffuse large B-cell lymphoma (DLBCL) each year, accounting for approximately 30% of all non-Hodgkin lymphoma cases. Although most patients can be cured with R-CHOP immunochemotherapy, 30-40% of patients progress or relapse after treatment. Therefore, there is an urgent need to find new treatments to improve the survival rate of this group of patients. Natural small molecule drugs have unique advantages as anticancer agents due to their low toxicity and multiple targets. This project aims to explore potentially effective natural compounds as new therapeutic strategies for DLBCL. Methods:Search for suitable natural compounds through GEO database, CMAP server and bioinformatics analysis. The effects of natural compounds on the biological function of DLBCL cells were studied by IC50 assay, apoptosis assay, cell cycle assay and subcutaneous transplanted tumor model. RNA sequencing, metabolomics detection, SEA server and 3D molecular simulation docking analysis were used to explore the drug target. The drug mechanism was verified by G6PD activity assay, EdU incorporation assay, NADPH assay and ROS assay. Results:We used GEO and CMAP databases to identify 20 natural compounds that target DLBCL prognostic genes. Cinobufagin, a natural bufodienol isolated from toad venom, became the main object of this study because of its low IC50 value. Flow cytometry analysis showed that cinobufagin can effectively induce G2 arrest and apoptosis of DLBCL cells. In addition, cinobufagin significantly inhibited the growth of tumor tissue in a mouse model of heterotopic subcutaneous tumor transplantation. The results of transcriptome sequencing suggested that cinobufagin treatment inhibited multiple metabolism-related pathways. Metabolomic analysis showed a decrease in nucleoside metabolite content and inhibition of niacinamide synthesis. Through SEA server and 3D molecular docking simulation analysis, it was found that glucose-6-phosphate dehydrogenase (G6PD) may be a potential target of cinobufagin. G6PD activity assay showed that cinobufagin could inhibit the catalytic ability of G6PD. EdU incorporation experiments showed that cinobufagin inhibited DNA synthesis in DLBCL cells. In addition, cinobufagin also promoted the up-regulation of NADP+/NADPH ratio and the increase of ROS. The addition of ROS scavengers and nucleotides partially reversed the apoptosis-inducing ability of cinobufagin. Conclusion:G6PD is a target of cinobufagin. By inhibiting the enzyme activity of G6PD, cinobufagin can inhibit DNA synthesis and the production of reducing NADPH, thereby inducing apoptosis of DLBCL cells. This suggests that Cinobufagin has potential application as a new drug for the treatment of DLBCL.

Transcription factor LHX9 (LIM Homeobox 9) enhances pyruvate kinase PKM2 activity to induce glycolytic metabolic reprogramming in cancer stem cells, promoting gastric cancer progression

BackgroundGlycolytic metabolic reprogramming is a phenomenon in which cells undergo altered metabolic patterns during malignant transformation, mainly involving various aspects of glycolysis, electron transport chain, oxidative phosphorylation, and pentose phosphate pathway. This reprogramming phenomenon can be used as one of the markers of tumorigenesis and development. Pyruvate kinase is the third rate-limiting enzyme in the sugar metabolism process by specifically catalyzing the irreversible conversion of PEP to pyruvate.PurposeThis study aimed to reveal the critical mediator(s) that regulate glycolytic metabolism reprogramming in gastric cancer and their underlying molecular mechanism and then explore the molecular mechanisms by which LHX9 may be involved in regulating gastric cancer (GC) progression.MethodsFirstly, we downloaded the GC and glycolysis-related microarray datasets from TCGA and MSigDB databases and took the intersection to screen out the transcription factor LHX9 that regulates GC glycolytic metabolic reprogramming. Software packages were used for differential analysis, single gene predictive analysis, and Venn diagram. In addition, an enrichment analysis of the glycolytic pathway was performed. Immunohistochemical staining was performed for LHX9 and PKM2 protein expression in 90 GC patients, and the association between their expressions was evaluated by Spearman's correlation coefficient method. Three human GC cell lines (AGS, NCI-N87, HGC-27) were selected for in vitro experimental validation. Flow cytometry was utilized to determine the stem cell marker CD44 expression status in GCSCs. A sphere formation assay was performed to evaluate the sphere-forming capabilities of GCSCs. In addition, RT-qPCR and Western blot experiments were employed to investigate the tumor stem cell markers OCT4 and SOX2 expression levels in GCSCs. Furthermore, a lentiviral expression vector was constructed to assess the impact of downregulating LHX9 or PKM2 on the glycolytic metabolic reprogramming of GCSCs. The proliferation, migration, and invasion of GCSCs were then detected by CCK-8, EdU, and Transwell assays. Subsequently, the mutual binding of LHX9 and PKM2 was verified using chromatin immunoprecipitation and dual luciferase reporter genes. In vivo experiments were verified by establishing a subcutaneous transplantation tumor model in nude mice, observing the size and volume of tumors in vivo in nude mice, and obtaining fresh tissues for subsequent experiments.ResultsBioinformatics analysis revealed that LHX9 might be involved in the occurrence and development of GC through regulating glycolytic metabolism. High LHX9 expression could be used as a reference marker for prognosis prediction of GC patients. Clinical tissue assays revealed that LHX9 and PKM2 were highly expressed in GC tissues. Meanwhile, GC tissues also highly expressed glycolysis-associated protein GLUT1 and tumor cell stemness marker CD44. In vitro cellular assays showed that LHX9 could enhance its activity and induce glycolytic metabolic reprogramming in GCSCs through direct binding to PKM2. In addition, the knockdown of LHX9 inhibited PKM2 activity and glycolytic metabolic reprogramming and suppressed the proliferation, migration, and invasive ability of GCSCs. In vivo animal experiments further confirmed that the knockdown of LHX9 could reduce the tumorigenic ability of GCSCs in nude mice by inhibiting PKM2 activity and glycolytic metabolic reprogramming.ConclusionThe findings suggest that both LHX9 and PKM2 are highly expressed in GCs, and LHX9 may induce the reprogramming of glycolytic metabolism through transcriptional activation of PKM2, enhancing the malignant biological properties of GCSCs and ultimately promoting GC progression.

Simvastatin induces pyroptosis via ROS/caspase-1/GSDMD pathway in colon cancer

BackgroundThe outcome of patients with colon cancer is still unsatisfied nowadays. Simvastatin is a type of statins with anti-cancer activity, but its effect on colon cancer cells remains unclear. The present study is intended to determine the underlying mechanism of simvastatin in treatment of colon cancer.MethodsThe viability and pyroptosis rate of cells treated and untreated with simvastatin were analysed by CCK-8 and flow cytometry assays, respectively. We used DCFH-DA and flow cytometry to detect reactive oxygen species (ROS) production. Levels of pyroptosis markers were detected by western blotting analysis or immunofluorescence staining. Besides, the anticancer properties of simvastatin on colon cancer were further demonstrated using a cell line based xenograft tumor model.ResultsSimvastatin treatment in HCT116 and SW620 induced pyroptosis and suppressed cell proliferation, with changes in the expression level of NLPR3, ASC, cleaved-caspase-1, mature IL-1β, IL-18 and GSDMD-N. Moreover, inhibition of caspase-1 and ROS attenuated the effects of simvastatin on cancer cell viability. In addition, it was identified that simvastatin has an anti-tumor effect by down-regulating ROS production and inducing downstream caspase-1 dependent pyroptosis in the subcutaneous transplantation tumors of HCT116 cells in BALB/c nude mice.ConclusionsOur in vitro and in vivo results indicated that simvastatin induced pyroptosis through ROS/caspase-1/GSDMD pathway, thereby serving as a potential agent for colon cancer treatment.7bxpkuYS9tFbFRGefzbb2gVideo

The upregulation of immune checkpoints after photodynamic therapy reducing immune effect for treating breast cancer.

The immune effect induced by photodynamic therapy (PDT) has a limited effect on breast tumor. This study hypothesized that suppressive immune checkpoints on T cells might upregulate after PDT, which may reduce the antitumor effect of PDT for treating breast tumor. This study explored the alteration of immune checkpoint for the first time. A bilateral subcutaneous transplanted breast tumor mice model was established, and right tumors imitated primary tumors, and left tumors imitated distant tumors. Primary tumors were treated with PDT mediated by hematoporphyrin derivatives (HpD-PDT). Costimulatory molecules (ICOS, OX40, and 4-1BB) and immune checkpoints (PD1, LAG-3, CTLA-4, TIM-3, TIGIT) on tumor infiltrating T cells after HpD-PDT were analyzed by flow cytometry. Antitumor and immune effects were also assessed after HpD-PDT combined with anti-PD1 and LAG-3 antibodies. Primary tumors were suppressed, but distant tumors could not be inhibited after HpD-PDT. The number of T cells was increased, but function did not enhance after HpD-PDT. Additionally, costimulatory molecules (ICOS, OX40, and 4-1BB) were not elevated, but the suppressive immune checkpoints on tumor infiltrating T cells were upregulated after HpD-PDT. Notably, PD1+ LAG-3+ CD4+ T and PD1+ LAG-3+ CD8+ T cells were significantly increased. When PD1 and LAG-3 blockade combined with HpD-PDT, both primary and distant tumors were significantly suppressed, and antitumor immune effects were significantly enhanced. HpD-PDT could upregulate the PD1+ LAG-3+ CD4+ T and PD1+ LAG-3+ CD8+ T cells. Dual blockade of PD1 and LAG-3 immune checkpoints can enhance the antitumor effect of HpD-PDT.

IGF2BP1 enhances the stability of SIK2 mRNA through m6A modification to promote non-small cell lung cancer progression

BackgroundNon-small cell lung cancer (NSCLC) is a significant public health concern globally. Evidence suggests that Salt-inducible kinase 2 (SIK2) is differentially expressed across various cancers and is also implicated in cancer progression. Despite this, the precise function of SIK2 in NSCLC is yet to be elucidated and requires further investigation. MethodsSIK2 expression was evaluated in both HBEC and NSCLC cells, utilizing quantitative real-time PCR (qRT-PCR) and Western blot (WB) analyses. Furthermore, to identify the influence of SIK2 on cell proliferation, migration, invasion, and apoptosis, a range of techniques were employed. To evaluate N6-methyladenosine (m6A) modification levels of total RNA and SIK2 within cells, RNA m6A colorimetry and methylated RNA immunoprecipitation (MeRIP) techniques were employed. Additionally, to confirm the interaction between SIK2 and insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1), bioinformatics analysis was executed, and the results were validated through RIP. The stability of SIK2 mRNA was determined using actinomycin D experiment. Furthermore, to validate the in vivo functionality of SIK2, a subcutaneous transplantation tumor model was established in nude mice. ResultsIn this study, upregulation of SIK2 in NSCLC cells was observed. Overexpression of SIK2 was found to lead to promotion of cell proliferation, migration, invasion, and suppression of the Hippo/yes-associated protein (YAP) pathway, while inhibiting apoptosis. RIP analysis showed that IGF2BP1 protein interacted with SIK2 mRNA. Knockdown of IGF2BP1 decreased mRNA stability and m6A modification levels of SIK2. Additionally, knockdown of IGF2BP1 resulted in inhibition of cell proliferation, migration, invasion, suppression of the Hippo/YAP pathway, and promoting apoptosis. Overexpression of SIK2 overturned the impact of IGF2BP1 on NSCLC cells, which was then confirmed through in vivo experiments. ConclusionIGF2BP1 stabilized SIK2 mRNA through m6A modification to promote NSCLC progression, potentially offering new diagnostic and therapeutic insights for NSCLC.

Umbilical Cord Blood-Derived M1 Macrophage Exosomes Loaded with Cisplatin Target Ovarian Cancer In Vivo and Reverse Cisplatin Resistance.

We investigated the therapeutic efficacy of umbilical cord blood (UCB)-derived M1 macrophage exosomes loaded with cisplatin (CIS) in ovarian cancer and platinum resistance. M1 macrophages were purified by using CD14 magnetic beads and characterized by flow cytometry. Our analyses included morphology, particle size, particle concentration, potential, drug loading capacity, counts of entry into cells, antitumor effect in vivo, and the ability to reverse drug resistance. A2780, SKOV3, and A2780/DDP, SKOV3/DDP ovarian cancer cells (CIS-sensitive and CIS-resistant cell lines, respectively) were treated with CIS or CIS-loaded M1 macrophage exosomes (M1exoCISs). The encapsulation efficiency of CIS loading into M1 macrophage exosomes was approximately 30%. In vitro, M1exoCIS treatment reduced the CIS IC50 values of both A2780, SKOV3, and A2780/DDP, SKOV3/DDP cells. We evaluated the effect of M1exoCIS on tumor growth using a mouse ovarian cancer subcutaneous transplantation tumor model inoculated with A2780/DDP cells. M1exoCIS was observed in the liver, spleen, and tumor sites 24 h posttreatment; the fluorescence intensity of M1exoCIS is higher than that of CIS. After 7 days, M1exoCIS significantly inhibited the growth of subcutaneously transplanted tumors compared with CIS alone and had a longer survival time. Moreover, the toxicity test shows that M1exoCIS has less hepatorenal toxicity than CIS. To investigate the mechanism of M1exoCIS targeting, homing, and reversing drug resistance, we performed RT-PCR, Western blotting, and Proteome Profiler Human Receptor Array analyses. We found that A2780 and A2780/DDP cells expressed the integrin β1/CD29 receptor, while M1 exosomes expressed integrin β1/CD29. In addition, M1exos carries long noncoding RNA H19, implicated in PTEN protein upregulation and miR-130a and Pgp gene downregulation, leading to the reversal of CIS drug resistance. Therefore, UCB-derived M1exoCIS target tumor sites of ovarian cancer in vivo and can be used to increase the CIS sensitivity and cytotoxicity.

The Effect of MDSC-Derived Exosomes Played in Esophageal Squamous Carcinoma Cells after Ionizing Radiation

Radiotherapy is the main treatment for esophageal cancer. Previous studies have shown that radiotherapy not only kills tumor cells directly, but also reshapes the immune microenvironment of the tumor. It has been reported an increase in the recruitment of myeloid-derived suppressor cells (MDSC) can occur in tumor tissue after ionizing radiation. Exosomes are mediators of intercellular information exchange and are also involved in the regulation of the tumor microenvironment. In this study, we wanted to understand whether MDSC in esophageal cancer tissue are involved in the regulation of tumor cell response to ionizing radiation via exosomes. KYSE-150 was used to construct a subcutaneous transplantation tumor model in nude mice. And then mice irradiated with 5 Gy×5fx and 0 Gy×5fx respectively. After irradiation, the spleens of the mice were used to isolate MDSC, and collect the cell supernatants to extract the exosomes. Based on the exosomes, we divided the experiment into three groups (control, exosomes, exosomes+radiation). Exosomes were injected into a nude mouse model of esophageal cancer via the tail vein or co-cultured with KYSE-150 cells. Mice were irradiated with a 5 Gy×5fx after completion of injection, and KYSE-150 cells were irradiated with a single dose 4 Gy. After radiation, KYSE-150 cells were used to detect cell cloning, apoptosis and cell cycle by flow cytometry, cell proliferation by CCK 8. XRCC4,XRCC5,XRCC6,γH2AX,ATM expression in cells and tumor tissue were measured by Western blot and RT-PCR. The tumor volume was significantly reduced after 5 Gy x 5fx radiation. When exosomes co-cultured with KYSE-150 cells, decrease in apoptosis and increase in cell cloning and cell proliferation were found in the exosomes+radiation group and exosomes group after radiation when compared with the control group, with this change being more pronounced in the exosome+radiation group. The results of the cell cycle assay showed that after ionizing radiation, the proportion of cells in the G0/G1 phase was significantly lower, and the proportion of cells in the S and G2/M phases were significantly higher in the exosomes+radiation group and exosomes group when compared to the Control group. The protein and mRNA expression of XRCC4,XRCC5,XRCC6,γH2AX,ATM in cells were increased in exosomes+radiation group and exosomes group after radiation when compared with the control group, with this change being more obvious in the exosome+radiation group. After irradiation, tumor volumes were measured in nude mice and the results showed that exosomes+radiation group tumors were the largest in volume, while the control group regressed most significantly after irradiation. MDSC-derived exosomes have a tumor growth-promoting effect in esophageal squamous carcinoma, which is enhanced by ionizing radiation, and this may be related to the accelerated repair of damage in tumor tissue after radiation.

Impact of PRaG Therapy on Immune Microenvironment of Bilateral Subcutaneous Tumor Model of Colon Cancer

Immune microenvironment is closely related to the efficacy of PD-1 inhibitors. The immune microenvironment contains a variety of immune cells, including effector T cells (cytotoxic CD8+T cells and effector CD4+T cells), dendritic cells (DC), and Myeloid-derived suppressor cells (MDSCs). The antitumor effects of PRaG therapy have been confirmed in bilateral subcutaneous transplantation tumor model of colon cancer. But the impact of PRaG therapy on immune microenvironment of such model is unclear. Therefore, the study continued to reveal the changes of immune microenvironment in mice. 80 male Balb/c mice aged 6-8 weeks were divided into five groups: control group, PD-1 inhibitor group, radiation group, radiation + PD-1 inhibitor group, and radiation + PD-1 inhibitor +GM-CSF (PRaG therapy) group. Bilateral subcutaneous tumor model of colon cancer in mice was constructed. 3×105 CT26.WT cells were inoculated subcutaneously in the right thigh root, and then the left thigh root 3 days later. Right subcutaneous tumor was selected for radiotherapy of 8 Gy×3. GM-CSF (100ng, i.p.) was given on the 1st day and PD-1 inhibitor (0.25mg/kg, i.p.) was given on the 2nd day after radiotherapy with one cycle every 3 days. On day 15, the spleen, left inguinal lymph node and left subcutaneous tumor of mice were collected. The proportion of immune cells was detected by flow cytometry. Compared with other groups, PRaG therapy decreased the proportion of cDC1 in left inguinal lymph node, increased the proportion of cDC2 in left subcutaneous tumor and left inguinal lymph node. Moreover, PRaG therapy increased the proportion of CD8+ effector memory T cells and CD226+CD8+T cells in left inguinal lymph nodes. Finally, PRaG therapy increased the proportion of CD4+, CD8+ central memory T cells and CD69+CD8+T cells and reduced the proportion of M-MDSCs in spleen. PRaG therapy can improve the immune microenvironment of spleen, unirradiated tumors and inguinal draining lymph nodes of bilateral subcutaneous tumor model of colon cancer in mice.