Proliferation Of T24 Cells Research Articles

Zn2+ enhanced the anti-breast cancer activity of fucoidan based nanoparticles by activating the cGAS-STING pathway

The incidence rate of breast cancer remains high, and lung metastasis often leads to treatment failure. Immunotherapy based on the cyclic guanosine monophosphate synthesis (cGAS) − interface gene stimulatory factor (STING) pathway is expected to bring benefits to the treatment of breast cancer. This study provided a delivery system based on fucoidan (Fu) and polyvinylpyrrolidone (PVP) for the potential molecule resveratrol (Res). The study confirmed that Fu and PVP were assembled into nanoparticles through hydrogen bonding and effectively loaded Res (Fu/PVP/Res). Interestingly, the adsorption of Zn2+ (Fu/PVP/Res Zn2+) was expected to provide impetus for the treatment of breast cancer. The results in vitro confirmed that Fu/PVP/Res Zn2+ significantly inhibited the proliferation of breast cancer 4T1 cells, induced the accumulation of mitochondrial reactive oxygen species (ROS) and the damage of mitochondrial membrane potential (MMP), and further induced the cytoplasmic release of mtDNA, which was the direct cause of the activation of cGAS-STING pathway. In vivo studies confirmed that Fu/PVP/Res Zn2+ treatment inhibited the growth of 4T1 transplanted tumors. Excitingly, Fu/PVP/Res Zn2+ treatment also prevented lung metastasis of in situ tumors. The in vivo mechanism confirmed that Fu/PVP/Res Zn2+ treatment activated the cGAS-STING pathway and induced abnormal expression of tumor apoptosis factors such as Bax/Bcl-2. In conclusion, our research provided potential for the treatment of breast cancer.

USP5 promotes tumor progression by stabilizing SLUG in bladder cancer.

Bladder cancer ranks as the second most prevalent urology malignancy globally. Invasive metastasis is a significant contributor to mortality among patients with bladder cancer, yet the underlying mechanisms remain elusive. Deubiquitinases are pivotal in carcinogenesis, with USP5 implicated in the malignant progression of hepatocellular carcinoma, colorectal cancer and non-small cell lung cancer. The present study assessed the role and mechanism of ubiquitin-specific proteinase 5 (USP5) in the malignant progression of bladder cancer. The association between USP5 expression and bladder cancer prognosis and stage was analyzed using The Cancer Genome Atlas database. Moreover, to elucidate the role of USP5 in bladder cancer, USP5 overexpression and knockdown cell lines were established using T24 cells. Cell viability, proliferation and migration were assessed using Cell Counting Kit-8, Transwell and scratch assays, respectively. Cyclohexanamide was used to evaluate the effect of USP5 expression on Snail family zinc finger 2 (SLUG) stability. Immunoprecipitation and immunofluorescence co-localization were utilized to probe the interaction between USP5 and SLUG. Changes in mRNA and protein levels were assessed using reverse transcription-quantitative PCR and western blotting, respectively. The results revealed that patients with bladder cancer with high USP5 expression had significantly shorter survival (P<0.05) and a higher clinicopathologic stage (P<0.05) than those with low USP5 expression. T24 cells overexpressing USP5 demonstrated significantly increased proliferation (P<0.05), invasion (P<0.05) and expression of epithelial-mesenchymal transition markers (P<0.05); whereas T24 cells with knocked-down USP5 expression revealed significantly reduced proliferation (P<0.05), invasion (P<0.05) and epithelial-mesenchymal transition markers (P<0.05). Immunoprecipitation experiments demonstrated the binding of USP5 to SLUG in bladder cancer cells, with further analysis revealing that USP5 upregulated protein levels of SLUG by inhibiting its ubiquitination. Furthermore, the treatment of bladder cancer cells with Degrasyn, a USP5 inhibitor, was associated with a significant inhibition of the proliferation (P<0.05) and invasion (P<0.05) of T24 cells. In conclusion, the findings of the present study underscore the role of USP5 in promoting the malignant progression of bladder cancer through the stabilization of SLUG. Targeting USP5 holds promise as a strategy for inhibiting bladder cancer progression.

Igniting tumour microenvironment in triple-negative breast cancer using a mannose/hyaluronic acid dual-coated Ganoderma polysaccharide-superparamagnetic iron oxide nanocomplex for combinational therapies

Eliciting tumour microenvironment (TME) activation in triple-negative breast cancer (TNBC) is crucial for effective anti-tumour therapies. The aim of this study is to employ pharmaceutical approaches to precisely deliver Ganoderma polysaccharide (GPS) to tumour sites, thereby enhancing TME activation. We first established a direct link between the accumulation of GPS within tumours and its efficacy in the TME activation. Building upon this insight, we then engineered a mannose/hyaluronic acid dual-coated GPS-loaded superparamagnetic iron oxide nanocomplex (Man/HA/GPS-SPIONs) with a particle size of 33.8 ± 1.6 nm and a zeta potential of −22.4 ± 3.5 mV, capable of precise tumour accumulation through magnet-assisted targeting and internalisation by tumour-associated macrophages (TAMs) and tumour cells, facilitated by dual ligand modification. In vitro, Man/HA/GPS-SPIONs effectively induced M1 polarisation of macrophages (CD86+ cells: 38.6 ± 2.8%), curbed 4T1 cell proliferation (viability: 47.3 ± 2.9%) and heightened Th1 cytokine release. Significantly, in vivo, Man/HA/GPS-SPIONs notably suppressed tumour growth (tumour index: 0.048 ± 0.005), fostered M1 polarisation of TAMs (CD45+F4/80+CD86+ cells: 26.1 ± 7.2%), consequently bolstering intratumoural T cytotoxic cells. This enhancement was intricately tied to the efficient co-delivery of GPS and iron ions to the tumours, made possible by the Man/HA/GPS-SPIONs delivery system. The synergistic effects with paclitaxel (PTX, inhibition rate: 61.2 ± 4.3%) and PD-1 inhibitors (inhibition rate: 69.8 ± 7.6%) underscored the translational potential of this approach. By harnessing a well-conceived iron-based drug delivery strategy, this study amplifies the tumour immune modulatory potential of natural polysaccharides, offering insightful guidance for interventions in the TME and synergistic therapies.

Valproic acid (VPA) exerts different effects on proliferation and viability of 4T1 parental and 4T1 cells overexpressing adaptor protein Ruk/CIN85 by triggering metabolic reprograming in type specific manner

Valproic acid (VPA) exerts different effects on proliferation and viability of 4T1 parental and 4T1 cells overexpressing adaptor protein Ruk/CIN85 by triggering metabolic reprograming in type specific manner

Comprehensive Genomic Analysis of Puerarin in Inhibiting Bladder Urothelial Carcinoma Cell Proliferation and Migration.

Bladder urothelial carcinoma (BUC) ranks second in the incidence of urogenital system tumors, and the treatment of BUC needs to be improved. Puerarin, a traditional Chinese medicine (TCM), has been shown to have various effects such as anti-cancer effects, the promotion of angiogenesis, and anti-inflammation. This study investigates the effects of puerarin on BUC and its molecular mechanisms. Through GeneChip experiments, we obtained differentially expressed genes (DEGs) and analyzed these DEGs using the Ingenuity® Pathway Analysis (IPA®), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) pathway enrichment analyses. The Cell Counting Kit 8 (CCK8) assay was used to verify the inhibitory effect of puerarin on the proliferation of BUC T24 cells. String combined with Cytoscape® was used to create the Protein-Protein Interaction (PPI) network, and the MCC algorithm in cytoHubba plugin was used to screen key genes. Gene Set Enrichment Analysis (GSEA®) was used to verify the correlation between key genes and cell proliferation. A total of 1617 DEGs were obtained by GeneChip. Based on the DEGs, the IPA® and pathway enrichment analysis showed they were mainly enriched in cancer cell proliferation and migration. CCK8 experiments proved that puerarin inhibited the proliferation of BUC T24 cells, and its IC50 at 48 hours was 218μmol/L. Through PPI and related algorithms, 7 key genes were obtained: ITGA1, LAMA3, LAMB3, LAMA4, PAK2, DMD, and UTRN. GSEA showed that these key genes were highly correlated with BUC cell proliferation. Survival curves showed that ITGA1 upregulation was associated with poor prognosis of BUC patients. Our findings support the potential antitumor activity of puerarin in BUC. To the best of our knowledge, bioinformatics investigation suggests that puerarin demonstrates anticancer mechanisms via the upregulation of ITGA1, LAMA3 and 4, LAMB3, PAK2, DMD, and UTRN, all of which are involved in the proliferation and migration of bladder urothelial cancer cells.

Integrated ceRNAs regulating relationship and bioinformatics analysis to study the molecular mechanisms of the inhibition of puerarin on bladder cancer cell

Based on previous experiments, we demonstrated puerarin inhibited the proliferation of BC T24 cells. To further explore the molecular mechanisms, whole transcriptome sequencing combined with bioinformatics analysis was performed. The results showed puerarin significantly inhibited T24 proliferation and pathway enrichment analysis of differentially expressed RNAs were mainly enriched in Cell cycle, PI3K/AKT, Ras family chromatin remodeling. lncRNAs and circRNAs may regulate miRNAs, thereby regulating the expression of ITGA1, PAK2 and UTRN. The predicted upstream transcription factor ERG and puerarin were well docked, which may be one of the underlying mechanisms by which puerarin inhibiting BC cells.

Hippophae Rhamnoides-derived Phytomedicine Nano-System Modulates Bax/Fas Pathways to Reduce Proliferation in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is the most common primary tumor of the breast with limited effectual drug availability. Therefore, the aim of the study is to develop an innovative phyto-nanomedicine (PNM) to cure TNBC with the least genotoxicity. Hereinafter, the sea buckthorn' extracted polyphenols (SBP), combine with metformin (MET), are synthesized as a novel PNM to evaluate its anti-tumor properties, effectiveness, and mechanism of action in TNBC in vitro and in vivo models. The SBP exhibits 16 new kinds of polyphenols that are been reported earlier which regulated cell development, proliferation, and programmed cell death (PCD) effectively. SBP-MET PNM inhibits MDA-MB-231 (47%), MDA-MB-436 (46%), and 4T1 (46%) cell proliferation but does not affect L929 normal murine cell development and successfully induce PCD (73.19%) in MDA-MB-231 cells. Mechanistically, in vivo SBP-MET proteome expression profiling reveals upregulation of proapoptotic Bax protein and activation of Fas signaling pathways convince downstream Daxx and FADD proteins, which further triggers Caspase-3 that prompts apoptosis in human TNBC cells by cleaving PARP-1 protein. Current findings establish innovative highly biocompatible phyto-nanomedicine that has significant potential to inhibit TNBC cell growth and induce regulated cell death (RCD) in vivo model, thereby opening a new arena for TNBC therapy.

Redox-responsive hyaluronic acid-celastrol prodrug micelles with glycyrrhetinic acid co-delivery for tumor combination therapy

Combining cytotoxic drugs with tumor microenvironment (TME) modulator agents is an effective strategy to enhance anti-tumor effects. In this study, two natural anti-tumor active ingredients celastrol (CEL) and glycyrrhetinic acid (GA) were combined for tumor treatment. In order to ensure the precise co-delivery and controllable synchronous release of combined drugs to tumors, it is necessary to construct a suitable nano-drug delivery platform. Based on this, we coupled hyaluronic acid (HA) with CEL by amide reaction to obtain an amphiphilic polymer prodrug HA-SS-CEL, and GA was spontaneously loaded into polymer micelles by self-assembly to obtain G/HSSC-M. G/HSSC-M has ideal size distribution, redox-responsive synchronous drug release, enhanced tumor cell internalization and in vivo tumor targeting. Compared with free drugs, the construction of multifunctional polymer micelles makes G/HSSC-M show better anticancer effect at the same concentration, and can significantly inhibit the proliferation and migration of HepG2 and 4T1 cells. In the in vivo experiments, G/HSSC-M inhibited tumor as high as 75.12% in H22 tumor-bearing mice. The mechanism included regulation of M1/M2 macrophage polarization, inhibition of Janus kinase 1/signal transducer and activator of transcription 3 (JAK1/STAT3) signaling pathway, and remodeling of tumor blood vessels. Therefore, the development of prodrug micelles co-loaded with CEL and GA provides a promising drug co-delivery strategy for combined cancer therapy.

A recombinant adeno-associated virus expressing secretory TGF-β type Ⅱ receptor inhibits triple-negative murine breast cancer 4T1 cell proliferation and lung metastasis in mice

To investigate the effects of an adeno-associated virus (AAV2) vector expressing secretory transforming growth factor-β (TGF-β) type Ⅱ receptor (sTβRⅡ) extracellular domain-IgG2a Fc fusion protein (sTβRⅡ-Fc) on proliferation and migration of triple-negative murine breast cancer 4T1 cells in mice. The pAAV-sTβRⅡ-Fc vector expressing sTβRⅡ-Fc fusion protein constructed by molecular cloning, the capsid protein-expressing vector pAAV2 and the helper vector were co-transfected into HEK 293T cells to prepare the recombinant AAV2-sTβRⅡ virus, which was purified by density gradient centrifugation with iodixanol. Western blotting was used to examine the effects of AAV-sTβRⅡ virus on Smad2/3 phosphorylation in 4T1 cells and on expression levels of E-cadherin, vimentin and p-Smad2/3 in 4T1 cell xenografts in mice. BALB/c mice bearing subcutaneous xenografts of luciferase-expressing 4T1 cells received intravenous injections of AAV-sTβRⅡ virus, AAV-GFP virus or PBS (n=6) through the tail vein, and the proliferation and migration of 4T1 cells were analyzed with in vivo imaging. Ki67 expression in the tumor tissues and sTβRⅡ protein expressions in mouse livers were detected with immunohistochemistry and immunofluorescence staining, and tumor metastases in the vital organs were examined with HE staining. The recombinant pAAV-sTβRⅡ-Fc vector successfully expressed sTβRⅡ in HEK 293T cells. Infection with AAV2-sTβRⅡ virus significantly reduced TGF-β1-induced Smad2/3 phosphorylation in 4T1 cells and effectively inhibited proliferation and lung metastasis of 4T1 xenografts in mice (P<0.05). In the tumor-bearing mice, intravenous injection of AAV-sTβRⅡ virus significantly increased E-cadherin expression, reduced vimentin and Ki67 protein expressions and Smad2/3 phosphorylation level in the tumor tissues (P<0.05 or 0.01), and induced liver-specific sTβRⅡ expression without causing body weight loss or heart, liver, spleen or kidney pathologies. The recombinant AVV2 vector encoding sTβRⅡ extracellular domain is capable of blocking the TGF-β signaling pathway to inhibit the proliferation and lung metastasis of 4T1 cells in mice.

Quercetin exerts anti-tumor immune mechanism by regulating IL-6/JAK2/STAT3 signaling pathway to deplete Treg cells

Breast cancer is still the leading cause of death among women worldwide. Due to the lack of effective drug targets, triple-negative breast cancer has a worse prognosis and higher mortality compared with other types of breast cancer, and chemotherapy is still the main treatment for triple-negative breast cancer at present. Quercetin (QUE) is a flavonoid compound found in a variety of fruits and vegetables. The mechanism of QUE has been extensively studied, such as prostate cancer, colon cancer, ovarian cancer, etc. However, the anti-tumor immune mechanism of QUE in triple-negative breast cancer remains unclear. Therefore, we assessed the anti-tumor immune effects of QUE on triple-negative breast cancer using both 4T1 cells and a xenograft mouse model of 4T1 cells. In vitro, we examined the inhibitory effects of QUE on 4T1 cells and its molecular mechanisms through MTT, Transwell, ELISA, and Western blotting. In vivo, by establishing a xenograft mouse model, we utilized flow cytometry, immunohistochemistry, ELISA, and Western blotting to evaluate the anti-tumor immune effects of QUE on triple-negative breast cancer. The results indicate that QUE inhibits the proliferation, migration, and invasion of 4T1 cells, concurrently significantly suppressing the IL-6/JAK2/STAT3 signaling pathway. Furthermore, it depletes Treg cell content in 4T1 xenograft mice, thereby improving the tumor immune microenvironment and promoting the cytotoxicity of relevant tumor immune cells. These findings suggest that QUE may serve as a potential adjuvant for immune therapy in triple-negative breast cancer.

Brazilin Actuates Ferroptosis in Breast Cancer Cells via p53/SLC7A11/GPX4 Signaling Pathway.

To investigate the mechanism of induction of ferroptosis by brazilin in breast cancer cells. Breast cancer 4T1 cells were divided into 6 groups: control, brazilin 1/2 half maximal inhibitory concentration (IC50), IC50, 2×IC50, erastin (10 µg/mL) and capecitabine (10 µg/mL) groups. The effect of brazilin on the proliferation of 4T1 cells was detected by cell counting kit-8 assay, and the treatment dose of brazilin was screened. The effect of brazilin on the mitochondrial morphology of 4T1 cells, and the mitochondrial damage was evaluated under electron microscopy. The levels of Fe2+, reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH) and glutathione peroxidase 4 (GPX4) were estimated using various detection kits. The invasion and migration abilities of 4T1 cells were detected by scratch assay and transwell assay. The expressions levels of tumor protein p53, solute carrier family 7 member 11 (SLC7A11), GPX4 and acyl-CoA synthetase long-chain family member 4 (ACSL4) proteins were quantified by Western blot assay. Compared to the control group, the 10 (1/2 IC50), 20 (IC50) and 40 (2×IC50) µg/mL brazilin, erastin, and capecitabine groups showed a significant decrease in the cell survival rate, invasion and migration abilities, GSH, SLC7A11 and GPX4 protein expression levels, and mitochondrial volume and ridge (P<0.05), and a significant increase in the mitochondria membrane density, Fe2+, ROS and MDA levels, and p53 and ACSL4 protein expression levels (P<0.05). Brazilin actuated ferroptosis in breast cancer cells, and the underlying mechanism is mainly associated with the p53/SLC7A11/GPX4 signaling pathway.

Fabrication of Quercetin-Functionalized Morpholine and Pyridine Motifs-Laden Silk Fibroin Nanofibers for Effective Wound Healing in Preclinical Study.

Choosing suitable wound dressings is crucial for effective wound healing. Spun scaffolds with bioactive molecule functionalization are gaining attention as a promising approach to expedite tissue repair and regeneration. Here, we present the synthesis of novel multifunctional quercetin with morpholine and pyridine functional motifs (QFM) embedded in silk fibroin (SF)-spun fibers (SF-QFM) for preclinical skin repair therapies. The verification of the novel QFM structural arrangement was characterized using ATR-FTIR, NMR, and ESI-MS spectroscopy analysis. Extensive characterization of the spun SF-QFM fibrous mats revealed their excellent antibacterial and antioxidant properties, biocompatibility, biodegradability, and remarkable mechanical and controlled drug release capabilities. SF-QFM mats were studied for drug release in pH 7.4 PBS over 72 h. The QFM-controlled release is mainly driven by diffusion and follows Fickian's law. Significant QFM release (40%) occurred within the first 6 h, with a total release of 79% at the end of 72 h, which is considered beneficial in effectively reducing bacterial load and helping expedite the healing process. Interestingly, the SF-QFM-spun mat demonstrated significantly improved NIH 3T3 cell proliferation and migration compared to the pure SF mat, as evidenced by the complete migration of NIH 3T3 cells within 24 h in the scratch assay. Furthermore, the in vivo outcome of SF-QFM was demonstrated by the regeneration of fresh fibroblasts and the realignment of collagen fibers deposition at 9 days post-operation in a preclinical rat full-thickness skin defect model. Our findings collectively indicate that the SF-QFM electrospun nanofiber scaffolds hold significant capability as a cost-effective and efficient bioactive spun architecture for use in wound healing applications.

Abstract 4278: Role of WNK1 in extracellular matrix modification in breast cancer

Abstract Introduction: Breast cancer (BCa) is the most common cancer and the second leading cause of cancer related deaths in women in the United States. Metastatic dissemination of cancer cells remains the leading cause of mortality in BCa. Therefore, determination of the mechanisms that promote metastasis is important for development of new therapeutic strategies. The extracellular matrix (ECM), which is the main non-cellular component of the tumor microenvironment (TME), plays an important role in BCa progression and metastasis. The cancer-associated fibroblasts (CAFs) are the principal cell types that together with BCa cells drive the modification of ECM during cancer development. Lysine deficient receptor protein kinase 1 (WNK1), a widely expressed serine threonine kinase that regulates blood pressure and electrolyte balance in the body, and is highly overexpressed in BCa microenvironment. The aim of the present study was to investigate the role of WNK1 in modification of ECM in BCa. Methods: The clinical relevance of WNK1 overexpression in BCa was determined by immunohistochemically staining human BCa sections, analyzing data from publicly available databases and creating orthotopic mouse models of BCa using 4T1 mouse BCa cells followed by treatment of the tumor bearing mice with WNK1 inhibitors, WNK-IN-11 and WNK463 (10 mg/kg , oral for 10 days). In vitro assays were conducted using 4T1 cells and mouse embryonic fibroblasts (MEFs) to determine the regulatory role of WNK1 on cancer cells and fibroblasts. Results and Conclusion: Our results indicated that WNK1 is strongly expressed in BCa cells and fibroblasts, the two most abundant cell types in the BCa microenvironment. Furthermore, WNK1 expression strongly correlated with expressions of matrix regulatory and cross-linking genes. Our results demonstrated that WNK1 inhibitors significantly reduced orthotopic 4T1 growth and metastasis. WNK1 inhibition also significantly inhibited collagen deposition in tumor tissues, which is a negative prognostic indicator in BCa. In vitro proliferation and migration assays using 4T1 cells and MEFs identified that WNK1 inhibitors (1-100nM) can significantly inhibit the proliferation and migration of 4T1 cells and MEFs. Results from co-culture studies with 4T1 cells and MEFs further indicated that WNK1 inhibition in MEFs could significantly reduce the invasive properties of 4T1 cells. Taken together, our results suggest that blocking the actions of WNK1 in BCa may be a promising therapeutic approach. Citation Format: Prabhat Suman, Sooraj Kakkat, Suhas S. Patil, Veronica Ramirez, Elba A. Turbat-Herrera, Seema Singh, Chandrani Sarkar, Debanjan Chakroborty. Role of WNK1 in extracellular matrix modification in breast 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 4278.

Simultaneous Visualization and Depletion of Peroxynitrite by a Simple Aggregation-Induced Emission Nanoprobe for Preventing Breast Cancer Metastasis after Surgery.

Inflammation has been confirmed to be closely related to the development of tumors, while peroxynitrite (ONOO-) is one of the most powerful oxidative pro-inflammatory factors. Although ONOO- can kill bacteria through oxidation, it will activate matrix metalloproteinases (MMPs), accelerate the degradation of the extracellular matrix (ECM), and subsequently lead to the activation and release of other tumor promotion factors existing in the ECM, promoting tumor metastasis and invasion. Herein, we report a simple aggregation-induced emission (AIE) nanoprobe (NP), TPE-4NMB, that can simultaneously visualize and deplete ONOO-. The probe can light up the endogenous and exogenous ONOO- in cells and selectively inhibit the proliferation and migration of 4T1 cells by inducing an intracellular redox homeostasis imbalance through ONOO- depletion. After being modified with DSPE-PEG2000, the TPE-4NMB NPs can be used to image ONOO- induced by various models in vivo; especially, it can monitor the dynamic changes of ONOO- level in the residual tumor after surgery, which can provide evidence for clarifying the association between surgery, ONOO-, and cancer metastasis. Excitingly, inhibited tumor volume growth and decreased counts of lung metastases were observed in the TPE-4NMB NPs group, which can be attributed to the downregulated expression of MMP-9 and transforming growth factor-β (TGF-β), increased cell apoptosis, and inhibited epithelial-mesenchymal transition (EMT) mediated by ONOO-. The results will provide new evidence for clarifying the relationship between surgery, ONOO-, and tumor metastasis and serve as a new intervention strategy for preventing tumor metastasis after tumor resection.

TFRC, associated with hypoxia and immune, is a prognostic factor and potential therapeutic target for bladder cancer

BackgroundBladder cancer is a common malignancy of the urinary system, and the survival rate and recurrence rate of patients with muscular aggressive (MIBC) bladder cancer are not ideal. Hypoxia is a pathological process in which cells acquire special characteristics to adapt to anoxic environment, which can directly affect the proliferation, invasion and immune response of bladder cancer cells. Understanding the exact effects of hypoxia and immune-related genes in BLCA is helpful for early assessment of the prognosis of BLCA. However, the prognostic model of BLCA based on hypoxia and immune-related genes has not been reported.PurposeHypoxia and immune cell have important role in the prognosis of bladder cancer (BLCA). The aim of this study was to investigate whether hypoxia and immune related genes could be a novel tools to predict the overall survival and immunotherapy of BLCA patients.MethodsFirst, we downloaded transcriptomic data and clinical information of BLCA patients from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. A combined hypoxia and immune signature was then constructed on the basis of the training cohort via least absolute shrinkage and selection operator (LASSO) analysis and validated in test cohort. Afterwards, Kaplan–Meier curves, univariate and multivariate Cox and subgroup analysis were employed to assess the accuracy of our signature. Immune cell infiltration, checkpoint and the Tumor Immune Dysfunction and Exclusion (TIDE) algorithm were used to investigate the immune environment and immunotherapy of BLCA patients. Furthermore, we confirmed the role of TFRC in bladder cancer cell lines T24 and UMUC-3 through cell experiments.ResultsA combined hypoxia and immune signature containing 8 genes were successfully established. High-risk group in both training and test cohorts had significantly poorer OS than low-risk group. Univariate and multivariate Cox analysis indicated our signature could be regarded as an independent prognostic factor. Different checkpoint was differently expressed between two groups, including CTLA4, HAVCR2, LAG3, PD-L1 and PDCD1. TIDE analysis indicated high-risk patients had poor response to immunotherapy and easier to have immune escape. The drug sensitivity analysis showed that high-risk group patients were more potentially sensitive to many drugs. Meanwhile, TFRC could inhibit the proliferation and invasion ability of T24 and UMUC-3 cells.ConclusionA combined hypoxia and immune-related gene could be a novel predictive model for OS and immunotherapy estimation of BLCA patients and TFRC could be used as a potential therapeutic target in the future.

IGF-PLGA microspheres promote angiogenesis and accelerate skin flap repair and healing by inhibiting oxidative stress and regulating the Ang 1/Tie 2 signaling pathway

Random flaps are widely used in the treatment of injuries, tumors, congenital malformations, and other diseases. However, postoperative skin flaps are prone to ischemic necrosis, leading to surgical failure. Insulin-like growth factor- 1(IGF-1) belongs to the IGF family and exerts its growth-promoting effects in various tissues through autocrine or paracrine mechanisms. Its application in skin flaps and other traumatic diseases is relatively limited. Poly (lactic-co-glycolic acid) (PLGA) is a degradable high-molecular-weight organic compound commonly used in biomaterials. This study prepared IGF-PLGA sustained-release microspheres to explore their impact on the survival rate of flaps both in vitro and in vivo, as well as the mechanisms involved. The research results demonstrate that IGF-PLGA has a good sustained-release effect. At the cellular level, it can promote 3T3 cell proliferation by inhibiting oxidative stress, inhibit apoptosis, and enhance the tube formation ability of human umbilical vein endothelial cells (HUVEC) . At the animal level, it accelerates flap healing by promoting vascularization through the inhibition of oxidative stress. Furthermore, this study reveals the role of IGF-PLGA in activating the Angiopoietin-1(Ang1)/Tie2 signaling pathway in promoting flap vascularization, providing a strong theoretical basis and therapeutic target for the application of IGF-1 in flaps and other traumatic diseases.

ARV-825 Showed Antitumor Activity against BRD4-NUT Fusion Protein by Targeting the BRD4.

The bromodomain-containing 4 (BRD4) is a member of the bromodomain and extra terminal domain (BET) family, which is an important epigenetic reader. It is currently a promising oncology target. In some tumors, BET bromodomain inhibitors have demonstrated promising results. Proteolysis-targeting methods (PROTAC), which rapidly and effectively degrade BRD4, have displayed considerable potential in the treatment of tumors in recent years. The purpose of this study is to examine the potential impact of BRD4 PROTAC compounds ARV-825 on oncogene BRD4-NUT fused protein in NUT carcinoma. The effectiveness of ARV-825 was evaluated at the cellular level using the cell counting kit 8 test, wound healing, cell transfection, western blotting analysis, and RNA sequencing. The effectiveness of ARV-825 was also examined in vivo using a xenograft model. The BRD4-NUT fusion gene was overexpressed in 3T3 cells, and the pathogenic fusion gene was simulated. The results showed that the overexpression of BRD4-NUT could promote the proliferation and migration of 3T3 cells, but the expression of BRD4 protein was degraded after the addition of the novel cereblon-based PROTAC compound ARV-825 against BRD4, resulting in inhibition of BRD4-NUT 3T3 cell proliferation and migration. Further RNA-seq analysis showed that overexpression of BRD4-NUT was accompanied by increased expression of gene (e.g., Myc, E2F, TRAFs, Wnt, Gadd45g, and Sox6) with significantly enriched pathway (e.g., small cell lung cancer, NF-kappa B signaling pathway, and breast cancer), promoted cell cycle from G 1 phase to S phase, and increased cell proliferation and migration, activated the antiapoptosisi signal, led to abnormal cell growth, and ultimately led to tumorigenesis. The addition of ARV-825 effectively rescued this process and effectively inhibited cell vitality, proliferation, and migration. In vivo studies demonstrated that treatment with ARV-825 greatly suppressed tumor growth without causing harmful side effects and downregulated the BRD4-NUT expression level. Through the induction of BRD4 protein degradation, ARV-825 can successfully limit BRD4-NUT 3T3 cell proliferation in vitro and in vivo. These findings suggested that the BRD4 inhibitor ARV-825 would be an effective therapeutic strategy for treating NUT carcinoma that with the genetic feature of BRD4-NUT fusion event.

Kiwi-derived extracellular vesicles for oral delivery of sorafenib

Sorafenib is an oral treatment for hepatocellular carcinoma (HCC). However, poor water solubility, harsh gastrointestinal environment and off-target effects contribute to the low bioavailability of oral sorafenib. Plant-derived extracellular vesicles (PDEVs) are biological nanovesicles with various bioactive functions that offer significant advantages in the field of oral drug delivery: protection from degradation by gastrointestinal fluids; crossing the intestinal epithelial barrier; specific targeting; safety; and abundant yield. However, there are fewer studies applying PDEVs for anti-tumor drug delivery to extra-digestive tissues. In this study, kiwifruit-derived extracellular vesicles (KEVs) were isolated and purified from kiwifruit, and their natural hepatic accumulation properties were exploited for targeted delivery of sorafenib (KEVs-SFB). Evidence showed that encapsulation of KEVs reduced the leakage of sorafenib in the gastrointestinal environment and enhanced the ability to cross the intestinal epithelium; KEVs-SFB was able to achieve liver accumulation and was predominantly taken up by HepG2 cells; KEVs-SFB was effective in inhibiting 4T1 cell proliferation; in the orthotopic liver cancer model, oral administration of KEVs-SFB inhibited tumor growth and improved the side effects of SFB. This PDEVs-based oral drug delivery platform is important for improving oral bioavailability and reducing drug side effects.

CDK2 and CDK4 targeted liensinine inhibits the growth of bladder cancer T24 cells

Bladder cancer (BCa) is a urinary tumor with limited treatment options and high mortality. Liensinine (LIEN), a natural bisbenzylisoquinoline alkaloid, has shown excellent anti-tumor effects in numerous preclinical studies. However, the anti-BCa effect of LIEN remains unclear. To the best of our knowledge, this is the first study to investigate the molecular mechanism of LIEN in the management of BCa. First, we identified the treatment-related targets of BCa; those that repeatedly occur in more than two databases, including GeneCards, Online Mendelian Inheritance in Man, DisGeNET, Therapeutic Target Database, and Drugbank. The SwissTarget database was used to screen LIEN-related targets, and those with a probability >0 were possible LIEN targets. The prospective targets of LIEN in the treatment of BCa were then determined using a Venn diagram. Second, we discovered that the PI3K/AKT pathway and senescence mediated the anti-BCa action of LIEN by using GO and KEGG enrichment analysis to explore the function of LIEN therapeutic targets. A protein-protein interaction network was created using the String website, and six algorithms of the CytoHubba plug-in were then used in Cytoscape to assess the core targets of LIEN for the therapy of BCa. The outcomes of molecular docking and dynamics simulation demonstrated that CDK2 and CDK4 proteins were the direct targets of LIEN in the management of BCa, among which CDK2 was more stable in binding to LIEN than CDK4. Finally, in vitro experiments showed that LIEN inhibited the activity and proliferation of T24 cells. The expression of p-/AKT, CDK2, and CDK4 proteins progressively decreased, while the expression and fluorescence intensity of the senescence-related protein, γH2AX, gradually increased with increasing LIEN concentration in T24 cells. Therefore, our data suggest that LIEN may promote senescence and inhibit proliferation by inhibiting the CDK2/4 and PI3K/AKT pathways in BCa.

ROS Responsive Nanoparticles Encapsulated with Natural Medicine Remodel Autophagy Homeostasis in Breast Cancer.

In photodynamic therapy (PDT), elevated reactive oxygen species (ROS) activate tumor cell protective autophagy, therefore attenuating the antitumor function of therapy. Hence, inhibition of protective autophagy in tumors can improve the antitumor effect of PDT. Herein, an innovative nanotraditional Chinese medicine system ((TP+A)@TkPEG NPs), which remodeled autophagy homeostasis, was fabricated. A photosensitizer aggregation inducing emission (AIE) and autophagy modulator triptolide (TP, an active ingredient of Tripterygium wilfordii Hook F) were encapsulated into ROS-responsive nanoparticles to improve antitumor effect of PDT in treatment of triple negative breast cancer. We proved that (TP+A)@TkPEG NPs effectively elevated intracellular ROS levels, activated ROS-responsive release of TP and inhibited the proliferation of 4T1 cells in vitro. More importantly, it sharply reduced autophagy related genes transcription and proteins expression in 4T1 cells, then promote cell apoptosis. In addition, this nanoherb therapeutic system effectively orientated to tumor sites, achieved efficient inhibition of tumor, and extended the survival time of 4T1-bearing mice in vivo. Further results confirmed that (TP+A)@TkPEG NPs remarkably inhibit the expression level of autophagy related initiation gene (becline-1) and elongation protein (light chain 3B) in tumor microenvironment and then block PDT induced protective autophagy. In brief, this system can remodel autophagy homeostasis and serve as an innovative approach for treatment of triple negative breast cancer.