Anti-apoptotic Genes Research Articles

METTL3 inhibits BMSC apoptosis and facilitates osteonecrosis repair via an m6A-IGF2BP2-dependent mechanism

Hypoxia-induced apoptosis of bone marrow mesenchymal stem cells (BMSCs) limits the efficacy of their transplantation for steroid-induced osteonecrosis of the femoral head (SONFH). As apoptosis and RNA methylation are closely related, exploring the role and mechanism of RNA methylation in hypoxic apoptosis of BMSCs is expected to identify new targets for transplantation of BMSCs for SONFH and enhance transplantation efficacy. We performed methylated RNA immunoprecipitation sequencing (MeRIP-seq) combined with RNA-seq on a hypoxia-induced apoptosis BMSC model and found that the RNA methyltransferase-like 3 (METTL3) is involved in hypoxia-induced BMSC apoptosis. The expression of METTL3 was downregulated in BMSCs after hypoxia and in BMSCs implanted in osteonecrosis areas. Knockdown of METLL3 under normoxic conditions promoted apoptosis of BMSCs. In contrast, overexpression of METTL3 promoted the survival of BMSCs under hypoxic conditions, and overexpression of METTL3 promoted the survival of BMSCs in the osteonecrosis area and the repair of the osteonecrosis area. Regarding the mechanism, the m6A levels of the mRNAs of anti-apoptotic genes Bcl-2, Mcl-1, and BIRC5 were significantly increased upon the overexpression of METTL3 under hypoxic conditions, which promoted the binding of Bcl-2, Mcl-1, and BIRC5 mRNAs to IGF2BP2, enhanced the mRNA stability, and increased the protein expression of the three anti-apoptotic genes. In conclusion, overexpression of METTL3 promoted m6A modification of mRNAs of Bcl-2, Mcl-1, and BIRC5, promoted the binding of IGF2BP2 to the above-mentioned mRNAs, enhanced mRNA stability, inhibited hypoxia-induced BMSC apoptosis, and promoted repair of SONFH, thereby providing novel targets for transplantation of BMSCs for SONFH.

Evaluation of the anticancer and antibacterial activities of moscatilin

Orchids (Dendrobium sp.) have been the subject of extensive research due to their ubiquitous pharmacological, antimicrobial, and anticancer properties. Moscatilin is a bibenzyl secondary metabolite enriched in orchids that exhibits anticancer and antimicrobial properties through mechanisms that have not yet been fully elucidated. The current study aimed to assess the in vitro anticancer and antibacterial potential of moscatilin. The in vitro anti-proliferative effects of moscatilin against breast cancer-MCF-7 and liver-HepG2 cells were assessed using the dimethylthiazol-diphenyltetrazolium bromide assay. Selected six pro-apoptotic (caspase-3, 8, 9, p53, p21 & Bax) and two anti-apoptotic (Bcl-xL & Bcl-2) gene markers were assessed via qPCR and tested antibacterial activity against various bacterial strains using disc diffusion and broth dilution methods. Moscatilin decreased the cellular viabilities of HepG2 and MCF-7 cancer cells, with anti-proliferation rates of 66 % (IC50 51 ± 5.18 μM) and 58 % (IC50 57 ± 4.18 μM), respectively. This effect was selectively observed in cancer cells, and the impact of moscatilin on non-cancerous MCF-12 cells was marginal. Moreover, moscatilin-treated cells exhibited higher mRNA levels of caspase-3,8, 9, Bax, p53, and p21, whereas lower levels of Bcl-2 and Bcl-xL, two anti-apoptotic markers, were observed. Furthermore, moscatilin exhibited varying degrees of antibacterial activity against the bacterial strains investigated. Notably, the highest antibacterial potentials were observed against Staphylococcus epidermidis and Klebsiella pneumonia, while the lowest inhibitory activity was observed in Escherichia coli and Pseudomonas aeruginosa. Overall, these findings demonstrated that moscatilin exerts potent anticancer effects via apoptosis and has antimicrobial properties against Gram-negative and Gram-positive bacteria that are clinically relevant. These findings highlight the potential of moscatilin as a natural therapeutic candidate for the treatment of cancer and clinically important bacterial pathogens.

Different recovery patterns of the surviving bivalve Mytilus galloprovincialis based on transcriptome profiling exposed to spherical or fibrous polyethylene microplastics

Microplastics (MPs) are pervasive pollutants exuded from anthropogenic activities and ingested by animals in different ecosystems. This transcriptomic profiling study aimed to explore the impact of polyethylene MPs on Mytilus galloprovincialis, an ecologically significant bivalve species. The toxicity of two MPs types was found to result in increased cellular stress when exposed up to 14 days. Moreover, recovery mechanisms were also observed in progress. Mussels exhibited different gene expression patterns and molecular regulation in response to cellular reactive oxygen species (ROS) stress. The transcriptome analysis demonstrated a notable hindrance in cilia movement as MPs ingested through gills. Subsequent entry resulted in a significant disruption in the cytoskeletal organization, cellular projection, and cilia beat frequency. On day 4 (D4), signal transduction and activation of apoptosis evidenced the signs of toxic consequences. Mussels exposed to spherical MPs shown significant recovery on day 14 (D14), characterized by the upregulation of anti-apoptotic genes and antioxidant genes. The expression of P53 and BCL2 genes was pivotal in controlling the apoptotic process and promoting cell survival. Mussels exposed to fibrous MPs displayed a delayed cell survival effect. However, the elevated physiological stress due to fibrous MPs resulted in energy transfer by compensatory regulation of metabolic processes to expedite cellular recovery. These observations highlighted the intricate and varied reaction of cell survival mechanisms in mussels to recover toxicity. This study provides critical evidence of the ecotoxicological impacts of two different MPs and emphasizes the environmental risks they pose to aquatic ecosystems. Our conclusion highlights the detrimental effects of MPs on M. galloprovincialis and the need for more stringent regulations to protect marine ecosystems.

Comparison of 177Lu-octreotate and 177Lu-octreotide for treatment in human neuroblastoma-bearing mice

BackgroundPatients with high-risk neuroblastoma (NB) have a 5-year event-free survival of less than 50 %, and novel and improved treatment options are needed. Radiolabeled somatostatin analogs (SSTAs) could be a treatment option. The aims of this work were to compare the biodistribution and the therapeutic effects of 177Lu-octreotate and 177Lu-octreotide in mice bearing the human CLB-BAR NB cell line, and to evaluate their regulatory effects on apoptosis-related genes. MethodsThe biodistribution of 177Lu-octreotide in mice bearing CLB-BAR tumors was studied at 1, 24, and 168 h after administration, and the absorbed dose was estimated to tumor and normal tissues. Further, animals were administered different amounts of 177Lu-octreotate or 177Lu-octreotide. Tumor volume was measured over time and compared to a control group given saline. RNA was extracted from tumors, and the expression of 84 selected genes involved in apoptosis was quantified with qPCR. ResultsThe activity concentration was generally lower in most tissues for 177Lu-octreotide compared to 177Lu-octreotate. Mean absorbed dose per administered activity to tumor after injection of 1.5 MBq and 15 MBq was 0.74 and 0.03 Gy/MBq for 177Lu-octreotide and 2.9 and 0.45 Gy/MBq for 177Lu-octreotate, respectively. 177Lu-octreotide treatment resulted in statistically significant differences compared to controls. Fractionated administration led to a higher survival fraction than after a single administration. The pro-apoptotic genes TNSFS8, TNSFS10, and TRADD were regulated after administration with 177Lu-octreotate. Treatment with 177Lu-octreotide yielded regulation of the pro-apoptotic genes CASP5 and TRADD, and of the anti-apoptotic gene IL10 as well as the apoptosis-related gene TNF. Conclusion177Lu-octreotide gave somewhat better anti-tumor effects than 177Lu-octreotate. The similar effect observed in the treated groups with 177Lu-octreotate suggests saturation of the somatostatin receptors. Pronounced anti-tumor effects following fractionated administration merited receptor saturation as an explanation. The gene expression analyses suggest apoptosis activation through the extrinsic pathway for both radiopharmaceuticals.

Trichinella spiralis Larval Extract as a Biological Anti-Tumor Therapy in a Murine Model of Ehrlich SolidCarcinoma.

Trichinella spiralis (T. spiralis) is an immunomodulating parasite that can adversely affect tumor growth and extend host lifespan. The aim of this study was to elucidate the mechanisms by which T. spiralis larval antigens achieve this effect using Ehrlich solid carcinoma (ESC) murine model. Assessment was done by histopathological and immunohistochemical analysis of caspase-3, TNF-α, Ki-67 and CD31. Additionally, Bcl2 and Bcl2-associated protein X (Bax) relative gene expression was assessed by molecular analysis for studying the effect of T. spiralis crude larval extract (CLE) antigen on tumor necrosis, apoptosis, cell proliferation and angiogenesis. We found that both T. spiralis infection and CLE caused a decrease in the areas of necrosis in ESC. Moreover, they led to increased apoptosis through activation of caspase-3, up-regulation of pro-apoptotic gene, Bax and down-regulation of anti-apoptotic gene, Bcl2. Also, T. spiralis infection and CLE diminished ESC proliferation, as evidenced by decreasing Ki-67. T. spiralis infection and CLE were able to suppress the development of ESC by inhibiting tumor proliferation, inducing apoptosis and decreasing tumor necrosis, with subsequent decrease in tumor metastasis. T. spiralis CLE antigen may be considered as a promising complementary immunotherapeutic agent in the treatment of cancer.

Evaluating the Anti-proliferative and Apoptotic Role of Atrial Natriuretic Peptide in Colon Cancer Cell Lines

The use of small peptides and conventional anticancer drugs is gaining importance in oncology as small peptides may help increase chemo or radiation sensitivity. The present study aimed to study the impact of atrial natriuretic peptide (ANP) in reducing colon cancer cell proliferation in primary and metastatic colon cancer cell lines. The proliferation of colon cancer cell lines (SW480 and SW620) was analysed by CCK-8 assay and cell damage was analyzed by lactate dehydrogenase activity. Catalase activity assay was performed to measure the oxidative stress and antioxidant defense mechanisms in SW480 and SW620 cell lines. Subsequently, up or downregulation of cancer-specific gene expression such as BAX, Caspase-3, BCL-2, CDK-6, and PCNA genes were assessed after the treatment of small peptide ANP in SW480 and SW620 colon cancer cell lines. The ANP treatment decreased the colon cancer cell proliferation, upregulated the apoptosis-related gene expression (Caspase-3, BAX), downregulated the anti-apoptotic gene (BCL-2) expression, and proliferation-related genes (CDK-6, PCNA) in SW480 and SW620 colon cancer cell lines, and the differences were found to be statistically significantly. Further, increased levels of catalase in the colon cancer cell lines after ANP-treatment suggested the therapeutic role of ANP. Subsequently, the LDH analysis showed the potential of ANP in inducing colon cancer cell damage. Collectively, the current study clearly shows that ANP is a potential molecule in reducing uncontrolled cancer cell growth. However, additional research using animal models and additional colon cancer cell lines is needed to validate its potential usage in clinical studies.

Synthesis, bioactivity assessment, molecular docking and ADMET studies of new chromone congeners exhibiting potent anticancer activity

In consideration of the chromones' therapeutic potential and anticancer activity, a new series of chromanone derivatives have been synthesized through a straightforward reaction between 6-formyl-7-hydroxy-5-methoxy-2-methylchromone (2) and various organic active compounds. The cytotoxic activity of the newly synthesized congeners was investigated against MCF-7 (human breast cancer), HCT-116 (colon cancer), HepG2 (liver cancer), and normal skin fibroblast cells (BJ1). The obtained data indicated that compounds 14b, 17, and 19 induce cytotoxic activity in the breast MCF7, while compounds 6a, 6b, 11 and 14c showed highly potent activity in the colon cancer cell lines. Overall, the results demonstrate that the potential cytotoxic effects of the studied compounds may be based on their ability to induce DNA fragmentation in cancer cell lines, down-regulate the expression level of CDK4 as well as the anti-apoptotic gene Bcl-2 and up-regulate the expression of the pro-apoptotic genes P53 and Bax. Furthermore, compounds 14b and 14c showed a dual mechanism of action by inducing apoptosis and cell cycle arrest. The docking studies showed that the binding affinity of the most active cytotoxic compounds within the active pocket of the CDK4 enzyme is stronger due to hydrophobic and H-bonding interactions. These results were found to be consistent with the experimental results.

Therapeutic Effects of Essential Oils and Their Bioactive Compounds on Prostate Cancer Treatment.

Since prostate cancer (PCa) relies on limited therapies, more effective alternatives are required. Essential oils (EOs) and their bioactive compounds are natural products that have many properties including anticancer activity. This review covers studies published between 2000 and 2023 and discusses the anti-prostate cancer mechanisms of the EOs from several plant species and their main bioactive compounds. It also provides a critical perspective regarding the challenges to be overcome until they reach the market. EOs from chamomile, cinnamon, Citrus species, turmeric, Cymbopogon species, ginger, lavender, Mentha species, rosemary, Salvia species, thyme and other species have been tested in different PCa cell lines and have shown excellent results, including the inhibition of cell growth and migration, the induction of apoptosis, modulation in the expression of apoptotic and anti-apoptotic genes and the suppression of angiogenesis. The most challenging aspects of EOs, which limit their clinical uses, are their highly lipophilic nature, physicochemical instability, photosensitivity, high volatility and composition variability. The processing of EO-based products in the pharmaceutical field may be an interesting alternative to circumvent EOs' limitations, resulting in several benefits in their further clinical use. Identifying their bioactive compounds, therapeutic effects and chemical structures could open new perspectives for innovative developments in the field. Moreover, this could be helpful in obtaining versatile chemical synthesis routes and/or biotechnological drug production strategies, providing an accurate, safe and sustainable source of these bioactive compounds, while looking at their use as gold-standard therapy in the close future.

Cytotoxic activity and mechanism of action of Smp43 scorpion peptide against colorectal cancer cell line HCT-116

This study was conducted to extend our previous investigations to examine the cytotoxic efficacy of Smp43 scorpion peptide against colorectal cancer cell line (HCT-116) and revealing its molecular mechanisms of action. Using MTT assay, Smp43 significantly induced cytotoxic effects on HCT-116 cancer cells compared to normal colon epithelial cell line (FHC) of IC50 4.11 and 62.17 µg/ml, respectively. Flow cytometric apoptosis assay showed that IC50 (4.11 µg/ml) of Smp43 induced a remarkable increase in the proportion of HCT-116 cancer cells undergoing late apoptotic cell death. Also, Smp43 caused cell cycle arrest at the S phase. RT-PCR analysis showed highly significant downregulation of Bcl-2 anti-apoptotic gene and ki-67 proliferative markers in HCT-116 treated with IC50 of Smp43. In contrast, caspase-3, −8, −9, Bax, and p53 apoptotic genes were upregulated. Protein expression levels of caspase-3, −8, −9, and p53 were similarly elevated in treated cells using a western blot analysis. The apoptotic characteristics were also confirmed through scanning and transmission electron microscopy. The Smp43 treated HCT-116 cancer cells showed cell membrane blebbing, pore formation, heterochromatin condensation toward the nuclear envelope, leakage of cell organelles, and formation of apoptotic bodies. Consequently, Smp43 peptide provides a great starting point for creating effective cytotoxic agents against human colorectal cancer.

Anticancer potential of thiocolchicoside and lauric acid loaded chitosan nanogel against oral cancer cell lines: a comprehensive study

The present study explored the anticancer activity of a Chitosan-based nanogel incorporating thiocolchicoside and lauric acid (CTL) against oral cancer cell lines (KB-1). Cell viability, AO/EtBr dual staining and Cell cycle analysis were done to evaluate the impact of CTL nanogel on oral cancer cells. Real-time PCR was performed to analyze proapoptotic and antiapoptotic gene expression in CTL-treated KB-1 cells. Further, molecular docking analysis was conducted to explore the interaction of our key ingredient, thiocolchicoside and its binding affinities. The CTL nanogel demonstrated potent anticancer activity by inhibiting oral cancer cell proliferation and inducing cell cycle arrest in cancer cells. Gene expression analysis indicated alterations in Bax and Bcl-2 genes; CTL nanogel treatment increased Bax mRNA expression and inhibited the Bcl-2 mRNA expression, which showed potential mechanisms of the CTL nanogel's anticancer action. It was found that thiocolchicoside can stabilize the protein's function or restore it as a tumour suppressor. The CTL nanogel exhibited excellent cytotoxicity and potent anticancer effects, making it a potential candidate for non-toxic chemotherapy in cancer nanomedicine. Furthermore, the nanogel's ability to modulate proapoptotic gene expression highlights its potential for targeted cancer therapy. This research contributes to the growing interest in Chitosan-based nanogels and their potential applications in cancer treatment.

Understanding the mechanism of ulcerative colitis-induced colorectal cancer and revealing the potential of Dillenia Indica in its management

Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD) that significantly increases the risk of developing colorectal cancer (CRC). The overall prevalence of UC-induced CRC is on the rise due to the rapidly increasing frequency of UC. It is widely acknowledged that chronic inflammation that persists for a long period is a significant contributing factor to the development of CRC in individuals with UC. The activation of various proinflammatory pathways, such as nuclear factor kappa B (NF-κB), interleukin-6/signal transducer and activator of transcription 3 (IL-6/STAT3), cyclooxygenase-2/prostaglandin E2 (COX-2/PGE2), and interleukin-23/T-helper 17 (IL-23/Th17), and gut microbiota contributes to the initiation and progression of tumorigenesis by a different mechanism, such as triggering the production of inflammatory mediators, increasing the expression of antiapoptotic genes, and stimulating cell proliferation as well as angiogenesis. Managing this risk and finding potential nutraceutical interventions, such as Dillenia indica (DI), is of significant interest. This review delves into the literature concerning the mechanism correlated to the development of CRC in UC individuals and investigates the potential of DI, as a nutraceutical in managing this rapid risk of UC-CRC. By examining the different mechanisms involved in UC-induced CRC, we aim to provide insight into the therapeutic promise of DI in mitigating these mechanisms.

NCI 159456 PERK Inhibitor as a Targeted Therapy for Lung Cancer: An In Vitro Study.

Non-small cell lung cancer (NSCLC) represents the most common histological type of lung cancer, characterized by a five-year survival rate of 15% and poor prognosis. Accumulating evidence indicates a prominent role of endoplasmic reticulum (ER) stress and the protein kinase RNA-like ER kinase (PERK)-dependent pathway of the unfolded protein response (UPR) in the pathogenesis of NSCLC. Increased expression of downstream targets of PERK was observed in various subtypes of NSCLC, and it was associated with a more aggressive phenotype, high risk of recurrence, and poor prognosis. Therefore, the present study aimed to investigate the biological effect of the selective PERK inhibitor NCI 159456 on A549 NSCLC cells and Human Pulmonary Fibroblasts (HPF) in vitro. Treatment of both normal and ER-stressed A549 cells with NCI 159456 resulted in a significant increase in the mRNA expression level of pro-apoptotic genes like activating transcription factor 4 (ATF4), DNA damage inducible transcript 3 (DDIT3), and BCL2 Associated X, Apoptosis Regulator (BAX) as well as a decreased level of the anti-apoptotic gene B-cell lymphoma 2 (Bcl-2). Cytotoxicity and genotoxicity analyses revealed that NCI 159456 significantly decreased viability and increased DNA damage in A549 cells under normal and ER stress conditions. Caspase-3 and reactive oxygen species (ROS) detection assays demonstrated that NCI 159456 significantly induced apoptosis and increased the ROS level in normal and ER-stressed A549 cells. Importantly, treatment with the inhibitor did not affect substantially normal HPF cells at any used concentration. The results indicate that PERK inhibitors could potentially be applied as a targeted therapy for NSCLC.

Multifaceted physiological and therapeutical impact of curcumin on hormone-related endocrine dysfunctions: A comprehensive review.

Over the past five decades, Curcumin (Cur), derived from turmeric (Curcuma longa), has gained considerable attention for its potential therapeutic applications. Synthesizing insights from clinical trials conducted over the last 25 years, this review delves into diseases where Cur has demonstrated promise, offering a nuanced understanding of its pharmacokinetics, safety, and effectiveness. Focusing on specific examples, the impact of Cur on various human diseases is explored. Endocrine glands and associated signaling pathways are highlighted, elucidating how Cur influences cellular signaling. The article underscores molecular mechanisms such as hormone level alteration, receptor interaction, cytokine and adipokine expression inhibition, antioxidant enzyme activity, and modulation of transcription factors. Cur showcases diverse protective mechanisms against inflammation and oxidative damage by suppressing antiapoptotic genes and impeding tumor promotion. This comprehensive overview emphasizes the potential of Cur as a natural agent for countering aging and degenerative diseases, calling for further dedicated research in this realm.

Allosteric Modulation of the YAP/TAZ-TEAD Interaction by Palmitoylation and Small-Molecule Inhibitors.

The Hippo signaling pathway is a highly conserved signaling network that plays a central role in regulating cellular growth, proliferation, and organ size. This pathway consists of a kinase cascade that integrates various upstream signals to control the activation or inactivation of YAP/TAZ proteins. Phosphorylated YAP/TAZ is sequestered in the cytoplasm; however, when the Hippo pathway is deactivated, it translocates into the nucleus, where it associates with TEAD transcription factors. This partnership is instrumental in regulating the transcription of progrowth and antiapoptotic genes. Thus, in many cancers, aberrantly hyperactivated YAP/TAZ promotes oncogenesis by contributing to cancer cell proliferation, metastasis, and therapy resistance. Because YAP and TAZ exert their oncogenic effects by binding with TEAD, it is critical to understand this key interaction to develop cancer therapeutics. Previous research has indicated that TEAD undergoes autopalmitoylation at a conserved cysteine, and small molecules that inhibit TEAD palmitoylation disrupt effective YAP/TAZ binding. However, how exactly palmitoylation contributes to YAP/TAZ-TEAD interactions and how the TEAD palmitoylation inhibitors disrupt this interaction remains unknown. Utilizing molecular dynamics simulations, our investigation not only provides detailed atomistic insight into the YAP/TAZ-TEAD dynamics but also unveils that the inhibitor studied influences the binding of YAP and TAZ to TEAD in distinct manners. This discovery has significant implications for the design and deployment of future molecular interventions targeting this interaction.

Functionalized siRNA-chitosan nanoformulations promote triple-negative breast cancer cell death via blocking the miRNA-21/AKT/ERK signaling axis: in-silico and in vitro studies

Oncogenic microRNA (miRNA), especially miRNA-21 upregulation in triple-negative breast cancer (TNBC), suggests a new class of therapeutic targets. In this study, we aimed to create GE11 peptide-conjugated small interfering RNA-loaded chitosan nanoparticles (GE11-siRNA-CSNPs) for the targeting of EGFR overexpressed TNBC and selectively inhibit miRNA-21 expression. A variety of in-silico and in vitro cellular and molecular studies were conducted to investigate the binding affinities of specific targets used as well as the anticancer efficacies and mechanisms of GE11-siRNA-CSNPs in TNBC cells. An in-silico assessment reveals a distinct binding affinity of miRNA-21 with siRNA as well as between the extracellular domain of EGFR and synthesized peptides. Notably, the in vitro results showed that GE11-siRNA-CSNPs were revealed to have better cytotoxicity against TNBC cells. It significantly inhibits miRNA-21 expression, cell migration, and colony formation. The results also indicated that GE11-siRNA-CSNPs impeded cell cycle progression. It induces cell death by reducing the expression of the antiapoptotic gene Bcl-2 and increasing the expression of the proapoptotic genes Bax, Caspase 3, and Caspase 9. Additionally, the docking analysis and immunoblot investigations verified that GE1-siRNA-CSNPs, which specifically target TNBC cells and suppress miRNA-21, can prevent the effects of miRNA-21 on the proliferation of TNBC cells via controlling EGFR and subsequently inhibiting the PI3K/AKT and ERK1/2 signaling axis. The GE11-siRNA-CSNPs design, which specifically targets TNBC cells, offers a novel approach for the treatment of breast cancer with improved effectiveness. This study suggests that GE11-siRNA-CSNPs could be a promising candidate for further assessment as an additional strategy in the treatment of TNBC.Graphical

A comparative study on targeted gene expression in zebrafish and its gill cell line exposed to chlorpyrifos.

Chlorpyrifos (CPF) is an organophosphorus-based insecticide, which is known to pose a serious risk to aquatic animals. However, the mechanisms of CPF toxicity in animals still remain unclear. The present investigation aimed to compare the potential effects of CPF in zebrafish (Danio rerio) and its gill cell line (DrG cells). Based on the in vivo study, the LC50 was calculated as 18.03µg/L and the chronic toxic effect of CPF was studied by exposing the fish to 1/10th (1.8µg/L) and 1/5th (3.6µg/L) of the LC50 value. Morphological changes were observed in fish and DrG cells which were exposed to sublethal concentrations of CPF. The results of MTT and NR assays showed significant decline in the survival of cells exposed to CPF at 96h. The production of reactive oxygen species in DrG cells and expression levels of antioxidant markers, inflammatory response genes (cox2a and cox2b), cyp1a, proapoptotic genes (bax), antiapoptotic gene (bcl2), apoptotic genes (cas3 and p53), and neuroprotective gene (ache) were determined in vivo using zebrafish and in vitro using DrG cells after exposure to CPF. Significant changes were found in the ROS production (DrG cells) and in the expression of inflammatory, proapoptotic, and apoptotic genes. This study showed that DrG cells are potential alternative tools to replace the use of whole fish for toxicological studies.

The potential anticancer effect of bee venom in combination with sorafenib against HepG2 cell lines via induction of apoptosis and autophagy candidate genes

Abstract Background Hepatocellular carcinoma (HCC) is a severe threat and a main reason for cancer-related deaths around the world. Drug resistance to sorafenib (Sorf), the effective HCC first-line therapy, is very common. A number of natural compounds, notably bee venom (BV), have been claimed to show a great impact against cancer when administered on its own or in conjunction with chemotherapy. Thus, this study aimed to investigate the anticancer effect of BV alone and/or combined with Sorf on HepG2 liver cancer cell lines. Methods Both mRNA and protein expressions of Bax, Bcl-2 and Beclin-1 were investigated by quantitative real-time PCR (qPCR) and western blot respectively, to examine the apoptotic and autophagic regulatory effects of BV and Sorf single treatments plus BV/Sorf combination on HepG2 cell lines. Results Our findings showed that BV and Sorf had considerable dose-dependent anti-proliferative effects on HepG2 cells whether administered alone or in combination, with the greatest impact for the combined therapies. Single BV and Sorf treatments showed IC50 of 93.21 and 7.28 μg/ml respectively, while combined treatment showed IC50 of 6.73 μg/ml BV + 6.73 μg/ml Sorf. Moreover, both the pro-apoptotic gene Bax and the autophagy-related gene Beclin-1 showed significant up-regulation in their mRNA expression, while the anti-apoptotic Bcl-2 mRNA gene expression showed significant down-regulation after BV/Sorf treatment as compared to either BV or Sorf single treatment. These qPCR results were further confirmed by western blot. Conclusions These findings indicate that BV synergistically potentiates the anticancer effect of Sorf on HepG2 cells through induction of apoptotic and autophagic machineries.

Abstract LB092: Novel tumor gene expression signatures predictive of outcome in large B cell lymphoma treated with CAR T cell therapy (axicabtagene ciloleucel)

Abstract We leveraged ZUMA-7, the first and largest, phase III randomized CAR-T cell clinical study of relapsed/refractory large B cell lymphoma (LBCL) in the 2nd line setting, to discover tumor gene expression (GE) signatures associated with outcome to anti-CD19 CAR-T cell therapy, Axicabtagene Ciloleucel (axi-cel). We performed GE profiling on pre-treatment LBCL tumor samples from 134 axi-cel-treated (safety dataset) patients using IO-360 Nanostring panel (769 genes). Multivariate analysis with penalized Cox models was conducted to identify genes whose expression (transcripts) associated with event-free survival (EFS), progression-free survival (PFS), and duration of response (DOR), leading to identification of two predictive GE signatures. For technical validation, the predictive value of these signatures was reproduced in a subset of ZUMA-7 samples profiled by RNA-seq (n=124). High levels (> median) of a favorable 6-transcript GE signature (6-GES) composed of CD19, CD45RA, CCL22, KLRK1, SOX11, and SIGLEC5 positively correlated with EFS (HR: 0.27, 95% CI: 0.16–0.44; P=1.82 × 10−8) and PFS (HR: 0.27, 95% CI: 0.16–0.46; P=1.35 × 10−7) in patients treated with axi-cel. The 6-GES may capture lymphomas with abundant adhesion molecules, a relatively low inflammation, and abundant expression of the targeted antigen (CD19). Conversely, high levels of an unfavorable 17-transcript GE signature (17-GES; CD45RO, BCL2, IL-18R1, TNFSF4 [OX40L], KLRB1 [CD161], KIR3DL2, ITGB8, DUSP5, GPC4, PSMB5, RPS6KB1, SERPINA9, NBN, GLUD1, ESR1, ARID1A, and SLC16A1) negatively correlated with Axi-cel EFS (HR: 6.19, 95% CI: 3.60–10.65; P=1.51 × 10−13) and PFS (HR: 7.58, 95% CI: 4.16–13.81; P=2.70 × 10−14). The 17-GES signature is consistent with a high level of immune infiltration and inflammation paralleled by the activation of immune-escape mechanisms, such as the upregulation of anti-apoptotic genes. Notably, the 17-GES was elevated at progression after axi-cel treatment (n=18). These signatures did not associate with outcome to 2nd-line SOC from ZUMA-7 (n = 122 for nanostring and n = 125 for ZUMA-7 RNAseq datasets, respectively), nor with outcome to 1st -line R-CHOP from two online datasets, indicating their predictive rather than prognostic value. In conclusion, our transcriptomic analysis of ZUMA-7 dataset identified novel GE signatures predictive of outcome with axi-cel. These GE signatures could support risk-stratification of LBCL patients. Citation Format: Yuan Tian, Justin Budka, Frederick L. Locke, Jason R. Westin, Christina To, Shilpa Shahani, Davide Bedognetti, Rhine R. Shen, Jorge Andrade, Simone Filosto. Novel tumor gene expression signatures predictive of outcome in large B cell lymphoma treated with CAR T cell therapy (axicabtagene ciloleucel) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB092.

The antitumoral effect of the esculetin in HeLa cells through endoplasmic reticulum stress.

Despite the multiple available treatment modalities, cervical cancer is one of the leading causes of mortality and morbidity among female gynecological cancers. Endoplasmic Reticulum (ER) is an effective organelle in ensuring cell homeostasis and is closely related to the development of cancer. Esculetin is a coumarin derivative that has anticancer, anti-inflammatory, antioxidant, and neuroprotective effects. Esculetin may have an anticancer effect by inducting apoptosis and ER stress. In this study, we evaluate that esculetin has an anti-tumor effect on human cervical cancer-derived (HeLa) cells via ER stress. Esculetin was applied to the HeLa cells, and a viability test was performed using the methyl thiazolyl tetrazolium proliferation (MTT) assay. Expression levels of apoptotic genes and anti-apoptotic genes were determined by real-time polymerase chain reaction. The results were statistically evaluated. Analysis of the MTT assay detected that esculetin inhibited HeLa cell viability development. Based on Western blot and quantitative real-time polymerase chain reaction (qPCR) analyses, esculetin destroyed cervical cancer cells via the ER stress pathway. The results showed that esculetin may have a potent antitumoral effect. It can potentially be utilized in the pharmacological therapy of cervical cancer.

Resolvin D1 alleviates apoptosis triggered by endoplasmic reticulum stress in IPEC-J2 cells

BackgroundResolvin D1 (RvD1), a specialized pro-resolving lipid mediator (SPM), is derived from docosahexaenoic acid (DHA). It plays a key role in actively resolving inflammatory responses, which further reduces small intestinal damage. However, its regulation of the apoptosis triggered by endoplasmic reticulum (ER) stress in intestinal epithelial cells is still poorly understood. The intestinal porcine epithelial cells (IPEC-J2) were stimulated with tunicamycin to screen an optimal stimulation time and concentration to establish an ER stress model. Meanwhile, RvD1 (0, 1, 10, 20, and 50 nM) cytotoxicity and its impact on cell viability and the effective concentration for reducing ER stress and apoptosis were determined. Finally, the effects of RvD1 on ER stress and associated apoptosis were furtherly explored by flow cytometry analysis, AO/EB staining, RT-qPCR, and western blotting.ResultsThe ER stress model of IPEC-J2 cells was successfully built by stimulating the cells with 1 µg/mL tunicamycin for 9 h. Certainly, the increased apoptosis and cell viability inhibition also appeared under the ER stress condition. RvD1 had no cytotoxicity, and its concentration of 1 nM significantly decreased cell viability inhibition (p= 0.0154) and the total apoptosis rate of the cells from 14.13 to 10.00% (p= 0.0000). RvD1 at the concentration of 1 nM also significantly reduced the expression of glucose-regulated protein 78 (GRP-78, an ER stress marker gene) (p= 0.0000) and pro-apoptotic gene Caspase-3 (p= 0.0368) and promoted the expression of B cell lymphoma 2 (Bcl-2, an anti-apoptotic gene)(p= 0.0008).ConclusionsCollectively, the results shed light on the potential of RvD1 for alleviating apoptosis triggered by ER stress, which may indicate an essential role of RvD1 in maintaining intestinal health and homeostasis.