Integrin-linked kinase-frizzled 7 interaction maintains cancer stem cells to drive platinum resistance in ovarian cancer.

BACKGROUND: Platinum-based chemotherapy regimens are a mainstay in the management of ovarian cancer (OC), but emergence of chemoresistance poses a significant clinical challenge. The persistence of ovarian cancer stem cells (OCSCs) at the end of primary treatment contributes to disease recurrence. Here, we hypothesized that the extracellular matrix protects CSCs during chemotherapy and supports their tumorigenic functions by activating integrin-linked kinase (ILK), a key enzyme in drug resistance. METHODS: TCGA datasets and OC models were investigated using an integrated proteomic and gene expression analysis and examined ILK for correlations with chemoresistance pathways and clinical outcomes. Canonical Wnt pathway components, pro-survival signaling, and stemness were examined using OC models. To investigate the role of ILK in the OCSC-phenotype, a novel pharmacological inhibitor of ILK in combination with carboplatin was utilized in vitro and in vivo OC models. RESULTS: In response to increased fibronectin secretion and integrin ß1 clustering, aberrant ILK activation supported the OCSC phenotype, contributing to OC spheroid proliferation and reduced response to platinum treatment. Complexes formed by ILK with the Wnt receptor frizzled 7 (Fzd7) were detected in tumors and correlated with metastatic progression. Moreover, TCGA datasets confirmed that combined expression of ILK and Fzd7 in high grade serous ovarian tumors is correlated with reduced response to chemotherapy and poor patient outcomes. Mechanistically, interaction of ILK with Fzd7 increased the response to Wnt ligands, thereby amplifying the stemness-associated Wnt/ß-catenin signaling. Notably, preclinical studies showed that the novel ILK inhibitor compound 22 (cpd-22) alone disrupted ILK interaction with Fzd7 and CSC proliferation as spheroids. Furthermore, when combined with carboplatin, this disruption led to sustained AKT inhibition, apoptotic damage in OCSCs and reduced tumorigenicity in mice. CONCLUSIONS: This "outside-in" signaling mechanism is potentially actionable, and combined targeting of ILK-Fzd7 may lead to new therapeutic approaches to eradicate OCSCs and improve patient outcomes.

Antitumor activity and phenolic profile of Melissa officinalis extract against human gastric adenocarcinoma cells.

Gastric cancer remains a global health concern, driving the exploration of natural products with anticancer potential. This study investigated the antiproliferative activity and chemical composition of a 70% ethanolic extract from Melissa officinalis L. against human gastric cancer cells. The extract was prepared and evaluated for total phenolic content, antioxidant capacity and flavonoid content. The MTT test checked how well it stopped the growth of human gastric adenocarcinoma (AGS) and normal dermal fibroblast (HDF) cells. Data analysis (SPSS Statistics) determined viable cell percentages and performed regression analysis (p<0.05). The extract exhibited significant antiproliferative activity against AGS cells compared to normal cells (p<0.05), with decreasing IC50 values (564.3, 258.0 and 122.5 µg/ml) over 24, 48 and 72 hours. It also displayed antioxidant activity (IC50=16.8±1.41µg/ml) and contained substantial phenolics (225.76±4.1 mg GAE/g) and flavonoids (22.36±2.6 mg RUT/g). This study suggests the 70% ethanolic extract of M. officinalis effectively suppresses AGS cell growth and possesses promising antioxidant properties, highlighting its potential as a natural source of anticancer and antioxidant agents, deserving further investigation.

Hsa_circ_0096157 silencing suppresses autophagy and reduces cisplatin resistance in non-small cell lung cancer by weakening the Nrf2/ARE signaling pathway.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the leading cause of cancer morbidity and mortality worldwide, and new diagnostic markers are urgently needed. We aimed to investigate the mechanism by which hsa_circ_0096157 regulates autophagy and cisplatin (DDP) resistance in NSCLC. METHODS: A549 cells were treated with DDP (0 µg/mL or 3 µg/mL). Then, the autophagy activator rapamycin (200 nm) was applied to the A549/DDP cells. Moreover, hsa_circ_0096157 and Nrf2 were knocked down, and Nrf2 was overexpressed in A549/DDP cells. The expression of Hsa_circ_0096157, the Nrf2/ARE pathway-related factors Nrf2, HO-1, and NQO1, and the autophagy-related factors LC3, Beclin-1, and p62 was evaluated by qRT‒PCR or western blotting. Autophagosomes were detected through TEM. An MTS assay was utilized to measure cell proliferation. The associated miRNA levels were also tested by qRT‒PCR. RESULTS: DDP (3 µg/mL) promoted hsa_circ_0096157, LC3 II/I, and Beclin-1 expression and decreased p62 expression. Knocking down hsa_circ_0096157 resulted in the downregulation of LC3 II/I and Beclin-1 expression, upregulation of p62 expression, and decreased proliferation. Rapamycin reversed the effect of interfering with hsa_circ_0096157. Keap1 expression was lower, and Nrf2, HO-1, and NQO1 expression was greater in the A549/DDP group than in the A549 group. HO-1 expression was repressed after Nrf2 interference. In addition, activation of the Nrf2/ARE pathway promoted autophagy in A549/DDP cells. Moreover, hsa_circ_0096157 activated the Nrf2/ARE pathway. The silencing of hsa_circ_0096157 reduced Nrf2 expression by releasing miR-142-5p or miR-548n. Finally, we found that hsa_circ_0096157 promoted A549/DDP cell autophagy by activating the Nrf2/ARE pathway. CONCLUSION: Knockdown of hsa_circ_0096157 inhibits autophagy and DDP resistance in NSCLC cells by downregulating the Nrf2/ARE signaling pathway.

Secretion of functional interferon by the type 3 secretion system of enteropathogenic Escherichia coli.

BACKGROUND: Type I interferons (IFN-I)-a group of cytokines with immunomodulatory, antiproliferative, and antiviral properties-are widely used as therapeutics for various cancers and viral diseases. Since IFNs are proteins, they are highly susceptible to degradation by proteases and by hydrolysis in the strong acid environment of the stomach, and they are therefore administered parenterally. In this study, we examined whether the intestinal bacterium, enteropathogenic Escherichia coli (EPEC), can be exploited for oral delivery of IFN-Is. EPEC survives the harsh conditions of the stomach and, upon reaching the small intestine, expresses a type III secretion system (T3SS) that is used to translocate effector proteins across the bacterial envelope into the eukaryotic host cells. RESULTS: In this study, we developed an attenuated EPEC strain that cannot colonize the host but can secrete functional human IFNα2 variant through the T3SS. We found that this bacteria-secreted IFN exhibited antiproliferative and antiviral activities similar to commercially available IFN. CONCLUSION: These findings present a potential novel approach for the oral delivery of IFN via secreting bacteria.

Fluoride impairs vascular smooth muscle A7R5 cell lines via disrupting amino acids metabolism.

Given the insidious and high-fatality nature of cardiovascular diseases (CVDs), the emergence of fluoride as a newly identified risk factor demands serious consideration alongside traditional risk factors. While vascular smooth muscle cells (VSMCs) play a pivotal role in the progression of CVDs, the toxicological impact of fluoride on VSMCs remains largely uncharted. In this study, we constructed fluorosis model in SD rats and A7R5 aortic smooth muscle cell lines to confirm fluoride impaired VSMCs. Fluoride aggravated the pathological damage of rat aorta in vivo. Then A7R5 were exposed to fluoride with concentration ranging from 0 to 1200 µmol/L over a 24-h period, revealing a dose-dependent inhibition of cell proliferation and migration. The further metabolomic analysis showed alterations in metabolite profiles induced by fluoride exposure, notably decreasing organic acids and lipid molecules level. Additionally, gene network analysis underscored the frequency of fluoride's interference with amino acids metabolism, potentially impacting the tricarboxylic acid (TCA) cycle. Our results also highlighted the ATP-binding cassette (ABC) transporters pathway as a central element in VSMC impairment. Moreover, we observed a dose-dependent increase in osteopontin (OPN) and α-smooth muscle actin (α-SMA) mRNA level and a dose-dependent decrease in ABC subfamily C member 1 (ABCC1) and bestrophin 1 (BEST1) mRNA level. These findings advance our understanding of fluoride as a CVD risk factor and its influence on VSMCs and metabolic pathways, warranting further investigation into this emerging risk factor.

Mechanisms of cordycepin in the treatment of pulmonary arterial hypertension in rats based on metabonomics and transcriptomics.

Pulmonary arterial hypertension (PAH) is a fatal disease featured by high morbidity and mortality. Although Cordycepin is known for its anti-inflammatory, antioxidant and immune-enhancing effects, its role in PAH treatment and the underlying mechanisms remain unclear. The therapeutic effects of Cordycepin on rats with PAH were investigated using a monocrotaline (MCT)-induced rat model. The metabolic effects of Cordycepin were assessed based on the plasma metabolome. The potential mechanisms of Cordycepin in PAH treatment were investigated through transcriptome sequencing and validated in pulmonary artery smooth muscle cells (PASMC). Evaluations included hematoxylin and eosin staining for pulmonary vascular remodeling, CCK-8 assay, EDU, and TUNEL kits for cell viability, proliferation, and apoptosis, respectively, and western blot for protein expression. Cordycepin significantly reduced right ventricular systolic pressure (RVSP) and right ventricular hypertrophy index (RVHI) in PAH rats, and mitigated pulmonary vascular remodeling. Plasma metabolomics showed that Cordycepin could reverse the metabolic disorders in the lungs of MCT-induced PAH rats, particularly impacting linoleic acid and alpha-linolenic acid metabolism pathways. Transcriptomics revealed that the P53 pathway might be the primary pathway involved, and western blot results showed that Cordycepin significantly increased P53 and P21 protein levels in lung tissues. Integrated analysis of transcriptomics and metabolomics suggested that these pathways were mainly enriched in linoleic acid metabolism and alpha-linolenic acid metabolism pathway. In vitro experiments demonstrated that Cordycepin significantly inhibited the PDGFBB (PD)-induced abnormal proliferation and migration of PASMC and promoted PD-induced apoptosis. Meanwhile, Cordycepin enhanced the expression levels of P53 and P21 proteins in PD-insulted PASMC. However, inhibitors of P53 and P21 eliminated these effects of Cordycepin. Cordycepin may activate the P53-P21 pathway to inhibit abnormal proliferation and migration of PASMC and promote apoptosis, offering a potential approach for PAH treatment.

Identification of small molecular inhibitors of SIRT3 by computational and biochemical approaches a potential target of breast cancer.

Sirtuin 3 (SIRT3) belongs to the Sirtuin protein family, which consists of NAD+-dependent lysine deacylase, involved in the regulation of various cellular activities. Dysregulation of SIRT3 activity has been linked to several types of cancer, including breast cancer. Because of its ability to stimulate adaptive metabolic pathways, it can aid in the survival and proliferation of breast cancer cells. Finding new chemical compounds targeted towards SIRT3 was the primary goal of the current investigation. Virtual screening of ~ 800 compounds using molecular docking techniques yielded 8 active hits with favorable binding affinities and poses. Docking studies verified that the final eight compounds formed stable contacts with the catalytic domain of SIRT3. Those compounds have good pharmacokinetic/dynamic properties and gastrointestinal absorption. Based on excellent pharmacokinetic and pharmacodynamic properties, two compounds (MI-44 and MI-217) were subjected to MD simulation. Upon drug interaction, molecular dynamics simulations demonstrate mild alterations in the structure of proteins and stability. Binding free energy calculations revealed that compounds MI-44 (- 45.61 ± 0.064 kcal/mol) and MI-217 (- 41.65 ± 0.089 kcal/mol) showed the maximum energy, suggesting an intense preference for the SIRT3 catalytic site for attachment. The in-vitro MTT assay on breast cancer cell line (MDA-MB-231) and an apoptotic assay for these potential compounds (MI-44/MI-217) was also performed, with flow cytometry to determine the compound's ability to cause apoptosis in breast cancer cells. The percentage of apoptotic cells (including early and late apoptotic cells) increased from 1.94% in control to 79.37% for MI-44 and 85.37% for MI-217 at 15 µM. Apoptotic cell death was effectively induced by these two compounds in a flow cytometry assay indicating them as a good inhibitor of human SIRT3. Based on our findings, MI-44 and MI-217 merit additional investigation as possible breast cancer therapeutics.

SPAG5 deficiency activates autophagy to reduce atherosclerotic plaque formation in ApoE-/- mice.

BACKGROUND: Autophagy, as a regulator of cell survival, plays an important role in atherosclerosis (AS). Sperm associated antigen 5 (SPAG5) is closely associated with the classical autophagy pathway, PI3K/Akt/mTOR signaling pathway. This work attempted to investigate whether SPAG5 can affect AS development by regulating autophagy. METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with oxidized-low density lipoprotein (ox-LDL) to induce cell damage. ApoE-/- mice were fed a Western diet to establish an AS mouse model. Haematoxylin and eosin (H&E) staining and Oil Red O staining evaluated the pathological changes and in lipid deposition in aortic tissues. CCK-8 and flow cytometry detected cell proliferation and apoptosis. Immunohistochemistry, Enzyme linked immunosorbent assay, qRT-PCR and western blotting assessed the levels of mRNA and proteins. RESULTS: Ox-LDL treatment elevated SPAG5 expression and the expression of autophagy-related proteins, LC3-I, LC3-II, Beclin-1, and p62, in HUVECs. GFP-LC3 dots were increased in ox-LDL-treated HUVECs and LPS-treated HUVECs. SPAG5 knockdown reversed both ox-LDL and LPS treatment-mediated inhibition of cell proliferation and promotion of apoptosis in HUVECs. SPAG5 silencing further elevated autophagy and repressed the expression of PI3K, p-Akt/Akt, and p-mTOR/mTOR in ox-LDL-treated HUVECs. 3-MA (autophagy inhibitor) treatment reversed SPAG5 silencing-mediated increase of cell proliferation and decrease of apoptosis in ox-LDL-treated HUVECs. In vivo, SPAG5 knockdown reduced atherosclerotic plaques in AS mice through activating autophagy and inhibiting PI3K/Akt/mTOR signaling pathway. CONCLUSION: This work demonstrated that SPAG5 knockdown alleviated AS development through activating autophagy. Thus, SPAG5 may be a potential target for AS therapy.

Hispidin Increases Cell Apoptosis and Ferroptosis in Prostate Cancer Cells Through Phosphatidylinositol-3-Kinase and Mitogen-activated Protein Kinase Signaling Pathway.

BACKGROUND/AIM: Chemotherapy is mainly used in the clinical treatment of prostate cancer. Different anticancer mechanisms can induce cell death in various cancers. Reactive oxygen species (ROS) play crucial roles in cell proliferation, differentiation, apoptosis, and signal transduction. It is widely accepted that ROS accumulation is closely related to chemical drug-induced cancer cell death. MATERIALS AND METHODS: We utilized the MTT assay to detect changes in cell proliferation. Additionally, colony formation and wound healing assay were conducted to investigate the effect of hispidin on cell colony formation and migration ability. Fluorescence microscopy was used to detect intracellular and mitochondrial ROS levels, while western blot was used for detection of cell apoptosis. RESULTS: Hispidin treatment significantly decreased viability of PC3 and DU145 cancer cells but exhibited no cytotoxicity in WPMY-1 cells. Furthermore, hispidin treatment inhibited cell migration and colony formation and triggered cellular and mitochondrial ROS accumulation, leading to mitochondrial dysfunction and mitochondrion-dependent apoptosis. Moreover, hispidin treatment induced ferroptosis in PC3 cells. Scavenging of ROS with N-acetyl cysteine significantly inhibited hispidin-induced apoptosis by altering the expression of apoptosis-related proteins, such as cleaved caspase-3, 9, Bax, and Bcl2. Furthermore, hispidin treatment dramatically up-regulated MAPK (involving p38, ERK, and JNK proteins) and NF-kB signaling pathways while down-regulating AKT phosphorylation. Hispidin treatment also inhibited ferroptosis signaling pathways (involving P53, Nrf-2, and HO-1 proteins) in PC3 cells. In addition, inhibiting these signaling pathways via treatment with specific inhibitors significantly reversed hispidin-induced apoptosis, cellular ROS levels, mitochondrial dysfunction, and ferroptosis. CONCLUSION: Hispidin may represent a potential candidate for treating prostate cancer.

The Combination of Auranofin and Celecoxib Suppresses Local Synovial Sarcoma Progression In Vivo.

BACKGROUND/AIM: Synovial sarcoma (SS) is a rare malignant tumor with a poor survival rate. We previously reported that a combination of auranofin (AUR), a thioredoxin reductase inhibitor, and celecoxib (CE), an anti-inflammatory drug, significantly impedes the local progression of osteosarcoma (OS). However, the role of redox regulation in SS remains to be elucidated. This study aimed to investigate the efficacy of combined treatment of AUR and CE on the local progression of SS in vivo. MATERIALS AND METHODS: Nu/nu mice were implanted with the human SS cell line, Aska-SS, and treated with vehicle control, AUR, or a combination of AUR and CE (AUR-CE). Primary tumor size and weight were evaluated for the study duration and upon resection, respectively. Hematoxylin and eosin (H&E) and Ki-67 staining were performed to assess the local progression of SS. RESULTS: A statistically significant reduction in tumor size and weight was observed in the AUR- and AUR-CE-treated groups upon excision compared to that in the vehicle-treated group. The AUR-CE-treated group showed synergistic inhibition of local tumor growth. H&E staining of local SS tumors revealed decreased cell density and nuclear deformation in the AUR- and AUR-CE-treated groups compared to those in the vehicle-treated group. Immunohistochemical staining revealed a statistically significant decrease in Ki-67-positive cells in the AUR-CE-treated group compared to the vehicle-treated group. CONCLUSION: The combination of AUR and CE showed significant potential for delaying the local progression of SS. These findings support the repurposing of AUR and CE as early treatment options for SS.

A Potential Combination of Targeting HSP90 and mTOR in Breast Cancer Cell Growth, Migration, and Invasion Through Inhibiting AKT Phosphorylation and F-actin Organization.

BACKGROUND/AIM: Breast cancer is the most prevalent form of cancer among women worldwide, with a high mortality rate. While the most common cause of breast cancer death is metastasis, there is currently no potential treatment for patients at the metastatic stage. The present study investigated the potential of using a combination of HSP90 and mTOR inhibitor in the treatment of breast cancer cell growth, migration, and invasion. MATERIALS AND METHODS: Gene Expression Profiling Interactive Analysis (GEPIA) was used to investigate the gene expression profiles. Western blot analysis and fluorescence staining were used for protein expression and localization, respectively. MTT, wound healing, and transwell invasion assays were used for cell proliferation, migration, and invasion, respectively. RESULTS: GEPIA demonstrated that HSP90 expression was significantly higher in breast invasive carcinoma compared to other tumor types, and this expression correlated with mTOR levels. Treatment with 17-AAG, an HSP90 inhibitor, and Torkinib, an mTORC1/2 inhibitor, significantly inhibited cell proliferation. Moreover, combination treatment led to down-regulation of AKT. Morphological changes revealed a reduction in F-actin intensity, a marked reduction of YAP, with interference in nuclear localization. CONCLUSION: Targeting HSP90 and mTOR has the potential to suppress breast cancer cell growth and progression by disrupting AKT signaling and inhibiting F-actin polymerization. This combination treatment may hold promise as a therapeutic strategy for breast cancer treatment that ameliorates adverse effects of a single treatment.

Combination Treatment of Biochanin A and Atorvastatin Alters Mitochondrial Bioenergetics, Modulating Cell Metabolism and Inducing Cell Cycle Arrest in Pancreatic Cancer Cells.

BACKGROUND/AIM: Pancreatic cancer is an aggressive type of cancer, with a dismally low survival rate of <5%. FDA-approved drugs like gemcitabine have shown little therapeutic success, prolonging survival by a mere six months. Isoflavones, such as biochanin A and daidzein, are known to exhibit anti-cancer activity, whereas statins reportedly have anti-proliferative effects. This study investigated the effects of combination treatment of biochanin A and atorvastatin on pancreatic cancer cells. MATERIALS AND METHODS: Pancreatic cancer cells AsPC-1, PANC-1, and MIA PaCa-2 were procured from ATCC. The cell viability studies were carried out using MTT & cell count assays. Flow cytometry was used to study cell apoptosis whereas cell metabolism studies were carried out using the Seahorse Mito stress test and XF-PMP assay. The effects of treatment on cell signaling pathways & cell cycle associated proteins were investigated using western blot whereas invasiveness of cancer cells was evaluated using gelatin zymography. RESULTS: The combination treatment decreased the survival and enhanced pro-apoptotic responses compared to single treatments in the pancreatic cancer cells. In PANC-1 cells, the combination treatment decreased invasiveness, reduced expression of activated STAT3 and expression of critical mediators of cell cycle progression. Furthermore, the combination treatment induced a differential inhibition of respiratory complexes in the pancreatic cancer cells. CONCLUSION: The combination treatment of biochanin A and atorvastatin exerts enhanced anti-cancer effects, inducing apoptosis, down-regulating cell cycle associated proteins and invasiveness in pancreatic cancer cells and merits further investigation for new, improved treatments for pancreatic cancer.

BET inhibition reforms the immune microenvironment and alleviates T cell dysfunction in chronic lymphocytic leukemia.

Redundant tumor microenvironment (TME) immunosuppressive mechanisms and epigenetic maintenance of terminal T cell exhaustion greatly hinder functional antitumor immune responses in chronic lymphocytic leukemia (CLL). Bromodomain and extraterminal (BET) proteins regulate key pathways contributing to CLL pathogenesis and TME interactions, including T cell function and differentiation. Herein, we report that blocking BET protein function alleviates immunosuppressive networks in the CLL TME and repairs inherent CLL T cell defects. The pan-BET inhibitor OPN-51107 reduced exhaustion-associated cell signatures resulting in improved T cell proliferation and effector function in the Eµ-TCL1 splenic TME. Following BET inhibition (BET-i), TME T cells coexpressed significantly fewer inhibitory receptors (IRs) (e.g., PD-1, CD160, CD244, LAG3, VISTA). Complementary results were witnessed in primary CLL cultures, wherein OPN-51107 exerted proinflammatory effects on T cells, regardless of leukemic cell burden. BET-i additionally promotes a progenitor T cell phenotype through reduced expression of transcription factors that maintain terminal differentiation and increased expression of TCF-1, at least in part through altered chromatin accessibility. Moreover, direct T cell effects of BET-i were unmatched by common targeted therapies in CLL. This study demonstrates the immunomodulatory action of BET-i on CLL T cells and supports the inclusion of BET inhibitors in the management of CLL to alleviate terminal T cell dysfunction and potentially enhance tumoricidal T cell activity.

Novel angiogenesis inhibitors with superoxide anion radical amplification effect: Surmounting the Achilles' heels of angiogenesis inhibitors and photosensitizers.

Angiogenesis inhibitors and photosensitizers are pivotal in tumor clinical treatment, yet their utilization is constrained. Herein, eleven novel angiogenesis inhibitors were developed through hybridization strategy to overcome their clinical limitations. These title compounds boast excitation wavelengths within the "therapeutic window", enabling deep tissue penetration. Notably, they could generate superoxide anion radicals via the Type I mechanism, with compound 36 showed the strongest superoxide anion radical generating capacity. Biological evaluation demonstrated remarkable cellular activity of all the title compounds, even under hypoxic conditions. Among them, compound 36 stood out for its superior anti-proliferative activity in both normoxic and hypoxic environments, surpassing individual angiogenesis inhibitors and photosensitizers. Compound 36 induced cell apoptosis via superoxide anion radical generation, devoid of dark toxicity. Molecular docking revealed that the target-recognizing portion of compound 36 was able to insert into the ATP binding pocket of the target protein similar to sorafenib. Collectively, our results suggested that hybridization of angiogenesis inhibitors and photosensitizers was a potential strategy to address the limitations of their clinical use.

Discovery of an efficacious KDM5B PROTAC degrader GT-653 up-regulating IFN response genes in prostate cancer.

Epigenetic alterations promote cancer development by regulating the expression of various oncogenes and anti-oncogenes. Histone methylation modification represents a pivotal area in epigenetic research and numerous publications have demonstrated that aberrant histone methylation is highly correlated with tumorigenesis and development. As a key histone demethylase, lysine-specific demethylase 5B (KDM5B) demethylates lysine 4 of histone 3 (H3K4) and serves as a transcriptional repressor of certain tumor suppressor genes. Meanwhile, KDM5B inhibits STING-induced intrinsic immune response of tumor cells or recruits SETDB1 through non-enzymatic function to silence reverse transcription elements to promote immune escape. The conventional small molecule inhibitors can only inhibit the enzymatic function of KDM5B with no effect on the non-enzymatic function. In the article, we present the development of the first series of KDM5B degraders based on CPI-455 to inhibit the non-enzymatic function. Among them, GT-653 showed optimal KDM5B degradation efficiency in a ubiquitin proteasome-dependent manner. GT-653 efficiently reduced KDM5B protein levels without affecting KDM5B transcription. Interestingly, GT-653 increased H3K4me3 levels and activated the type-I interferon signaling pathway in 22RV1 cells without significant phenotypic response on cell proliferation.

Integrating network pharmacology, bioinformatics, and experimental validation to unveil the molecular targets and mechanisms of galangin for treating hepatocellular carcinoma.

BACKGROUND: Galangin, a flavonoid compound, is derived from Alpinia officinarum Hance. Previous studies have shown that galangin can inhibit the proliferation of hepatocellular carcinoma (HCC), but its mechanism is still unclear. This study aims to investigate the potential targets and molecular mechanisms of galangin on HCC through network pharmacology, bioinformatics, molecular docking, and experimental in vitro validation. METHODS: In this study, network pharmacology was used to investigate the targets and mechanisms of galangin in the treatment of HCC. AutoDockTools software was used to simulate and calculate the binding of galangin to its core targets. GO and KEGG enrichment analyses were conducted in the DAVID database to explore the main biological functions and signaling pathways impacted by galangin intervention. In addition, bioinformatics was applied to examine the correlation between the differential expressions of the anti-HCC core targets of galangin and the survival of patients with HCC. Finally, the findings obtained from network pharmacology and bioinformatics were verified in cell experiments. RESULTS: A total of 67 overlapping target genes of galangin and HCC were identified. Through the analysis of the protein-protein interaction (PPI) network, 10 hub genes with the highest degree of freedom were identified, including SRC, ESR1, MMP9, CDK4, CCNB1, MMP2, CDK2, CDK1, CHK1, and PLK1. These genes were found to be closely related to the degradation of the extracellular matrix, signal transduction, and the cell cycle. GO and KEGG enrichment analyses revealed that galangin exerts an anti-HCC role by affecting various signaling pathways, including the cell cycle, pathways in cancer, and the PI3K-Akt signaling pathway. The results of molecular docking indicated a significant interaction between galangin and CCNB1, CDK4, CDK1, and PLK1. Bioinformatics analysis revealed that CCNB1, CDK4, CDK1, and PLK1 were upregulated in the liver of patients with HCC at both the mRNA and protein levels. Flow cytometry analysis showed that galangin induced G0/G1 phase arrest and cell apoptosis in HepG2 and Huh7 cells. Additionally, galangin suppressed the expression of key proteins and mRNAs involved in the cell cycle pathway. CONCLUSIONS: These results suggest that galangin inhibits the growth of HCC cells by arresting the cell cycle at the G0/G1 phase.

Paracrine- and cell-contact-mediated immunomodulatory effects of human periodontal ligament-derived mesenchymal stromal cells on CD4+ T lymphocytes.

BACKGROUND: Mesenchymal stromal cells (MSCs) isolated from the periodontal ligament (hPDL-MSCs) have a high therapeutic potential, presumably due to their immunomodulatory properties. The interaction between hPDL-MSCs and immune cells is reciprocal and executed by diverse cytokine-triggered paracrine and direct cell-to-cell contact mechanisms. For the first time, this study aimed to directly compare the contribution of various mechanisms on this reciprocal interaction using different in vitro co-culture models at different inflammatory milieus. METHODS: Three co-culture models were used: indirect with 0.4 µm-pored insert, and direct with or without insert. After five days of co-culturing mitogen-activated CD4+ T lymphocytes with untreated, interleukin (IL)-1ß, or tumor necrosis factor (TNF)-α- treated hPDL-MSCs, the CD4+ T lymphocyte proliferation, viability, and cytokine secretion were investigated. The gene expression of soluble and membrane-bound immunomediators was investigated in the co-cultured hPDL-MSCs. RESULTS: Untreated hPDL-MSCs decreased the CD4+ T lymphocyte proliferation and viability more effectively in the direct co-culture models. The direct co-culture model without inserts showed a strikingly higher CD4+ T lymphocyte cell death rate. Adding IL-1ß to the co-culture models resulted in substantial CD4+ T lymphocyte response alterations, whereas adding TNF resulted in only moderate effects. The most changes in CD4+ T lymphocyte parameters upon the addition of IL-1ß or TNF-α in a direct co-culture model without insert were qualitatively different from those observed in two other models. Additionally, the co-culture models caused variability in the immunomediator gene expression in untreated and cytokine-triggered hPDL-MSCs. CONCLUSION: These results suggest that both paracrine and cell-to-cell contact mechanisms contribute to the reciprocal interaction between hPDL-MSCs and CD4+ T lymphocytes. The inflammatory environment affects each of these mechanisms, which depends on the type of cytokines used for the activation of MSCs' immunomodulatory activities. This fact should be considered by comparing the outcomes of the different models.

Caffeic Acid Enhances Anticancer Drug-induced Apoptosis in Acid-adapted HCT116 Colon Cancer Cells.

BACKGROUND/AIM: Apoptosis resistance in cancer cells adapted to acidic microenvironments poses a challenge for effective treatment. This study investigated the potential use of caffeic acid as an adjunct therapy to overcome drug resistance in colorectal cancer cells under acidic conditions. MATERIALS AND METHODS: Long-term exposure to low-pH conditions induced resistance in HCT116 colorectal cancer cells. The effects of caffeic acid on proliferation, clonogenicity, and apoptosis induction were assessed alone and in combination with oxaliplatin and 5-Fluorouracil. The signaling pathways involved in drug resistance were examined by assessing the activities of PI3K/Akt and ERK1/2. RESULTS: Caffeic acid inhibited the proliferation and clonogenicity of acid-adapted cancer cells, and enhanced apoptosis when combined with anticancer drugs. Mechanistically, caffeic acid attenuated the hyperactivation of the PI3K/Akt and ERK1/2 signaling pathways associated with drug resistance. CONCLUSION: Caffeic acid is a promising therapeutic agent for targeting resistant cancer cells in acidic microenvironments. Its ability to inhibit proliferation, sensitize cells to apoptosis, and modulate signaling pathways highlights its potential for overcoming drug resistance in cancer therapy.

Ex Vivo Evaluation of Antiangiogenic Drugs in Oral Cancer: Potential Implications for Targeting Vasculogenic Mimicry.

BACKGROUND/AIM: Oral squamous cell carcinoma (OSCC) is characterized by early metastasis, clinical resistance and poor prognosis. Recently, we showed that aggressive OSCC cells co-express endothelial cell markers and can form tube-like structures, known as vasculogenic mimicry (VM), a process associated with poor prognosis in head and neck cancers. Given the limited success of current antiangiogenic therapy in treating OSCC, this study sought to explore the efficiency of these drugs in targeting an ex vivo model of VM. MATERIALS AND METHODS: OSCC cell lines from the tongue and floor of the mouth in addition to human endothelial cells were used. The treatments comprised a set of clinically relevant antiangiogenic drugs: sorafenib, sunitinib, and axitinib, which were administered in different doses. Multiple ex vivo approaches including cell tubulogenesis, proliferation, apoptosis, and migration assays were used. RESULTS: Although these drugs inhibited the formation of endothelial cell capillaries, they showed clear differential effects on OSCC cell-derived VM and cell morphology. Sorafenib inhibited the tubulogenesis of aggressive OSCC cells compared with the limited effect of sunitinib and axitinib. Furthermore, our data consistently demonstrated a preferential efficacy of certain drugs over others. Sorafenib and sunitinib exhibited anti-cancer effects on tumor cell proliferation, apoptosis, and cell migration, compared with the limited effect of axitinib. CONCLUSION: The antiangiogenic drugs, except sorafenib, had limited effect on VM formation in vitro and exhibited varying anti-cancer effects on OSCC cells. These data support the notion that VM formation may in part explain the development of drug resistance in OSCC cells.

Antitumor Activity of Tasurgratinib as an Orally Available FGFR1-3 Inhibitor in Cholangiocarcinoma Models With FGFR2-fusion.

BACKGROUND/AIM: Cholangiocarcinoma (CCA) is an aggressive tumor with limited treatment options especially in 2nd line or later treatments. Targeting fibroblast growth factor receptor (FGFR) 2 has recently emerged as a promising treatment option for patients with CCA harboring FGFR2-fusion. This study investigated the antitumor activities of tasurgratinib as an orally available FGFR1-3 inhibitor, in preclinical FGFR2-driven CCA models. MATERIALS AND METHODS: Antitumor activities of tasurgratinib were examined in vitro and in vivo using NIH/3T3 cells expressing FGFR2-fusion as FGFR2-driven CCA models, and in vivo using a CCA patient-derived xenograft model. The molecular mechanism of action of tasurgratinib was elucidated through co-crystal structure analysis with FGFR1, manual complex model analysis with FGFR2, and binding kinetics analysis with FGFR2. Furthermore, the cell-based inhibitory activities against acquired resistant FGFR2 mutations in patients with CCA treated with FGFR inhibitors were evaluated. RESULTS: Tasurgratinib showed antitumor activity in preclinical FGFR2-driven CCA models by inhibiting the FGFR signaling pathway in vitro and in vivo. Furthermore, cell-based target engagement assays indicated that tasurgratinib had potent inhibitory activities against FGFR2 mutations, such as N549H/K, which are the major acquired mutations in CCA. We also confirmed that tasurgratinib exhibited fast association and slow dissociation kinetics with FGFR2, binding to the ATP-binding site and the neighboring region, and adopting an Asp-Phe-Gly (DFG)-"in" conformation. CONCLUSION: These data demonstrate the therapeutic potential of tasurgratinib in FGFR2-driven CCA and provide molecular mechanistic insights into its unique inhibitory profile against secondary FGFR2 resistance mutations in patients with CCA treated with FGFR inhibitors.