Dose In Cells Research Articles

Quercetin and Mesenchymal Stem Cell Metabolism: A Comparative Analysis of Young and Senescent States.

Quercetin is a natural flavonoid renowned for its potent antioxidant, anti-inflammatory, anti-diabetic, and antibacterial properties, making it a highly promising candidate for the treatment of various medical conditions. Our current study investigates the influence of quercetin on energy metabolism, fatty acid composition, oxidative stress gene expression, and sirtuin expression in early- and late-stage passages of stem cells derived from human exfoliated deciduous teeth (SHEDs). Mitochondrial respiration was analyzed by measuring oxygen consumption following a 24 h quercetin treatment, while fatty acid profiles were examined using gas chromatography-mass spectrometry (GC-MS). Additionally, quantitative PCR (qPCR) was used to assess the expression of oxidative stress genes and sirtuins. In younger SHEDs, quercetin enhances metabolic activity and mitochondrial respiration, although higher doses may decrease mitochondrial activity. Conversely, in older, senescent SHEDs, quercetin supports mitochondrial function at lower concentrations but appears to inhibit respiration at higher doses. These results suggest that quercetin may hold therapeutic potential for maintaining SHED viability and function, especially at lower doses in older cells. Further research is essential to fully elucidate a dose-dependent effect of quercetin and optimize its applications in regenerative medicine.

Effects of Arborvitae (Thuja plicata) Essential Oil on Cervical Cancer Cells: Insights into Molecular Mechanisms.

This study aimed to assess the effects of AEO in an in vitro model of cell lines derived from cervical cancer-namely, HeLa and SiHa-by screening for AEO's cytotoxic properties and examining its influence on the modulation of gene expression. Cervical cancer stands as a prevalent global health concern, affecting millions of women worldwide. The current treatment modalities encompass surgery, radiation, and chemotherapy, but significant limitations and adverse effects constrain their effectiveness. Therefore, exploring novel treatments that offer enhanced efficacy and reduced side effects is imperative. Arborvitae essential oil, extracted from Thuja Plicata, has garnered attention for its antimicrobial, anti-inflammatory, immunomodulatory, and tissue-remodeling properties; however, its potential in treating cervical cancer remains uncharted. The objective of this study was to delve into the molecular mechanisms induced by arborvitae essential oil in order to learn about its anticancer effects on cervical cancer cell lines. The methods used in this study were assessments of cell viability using WST-1 and annexin V- propidium iodide, mRNA sequencing, and subsequent bioinformatics analysis. The findings unveiled a dose-dependent cytotoxic effect of arborvitae essential oil on both HeLa and SiHa cell lines. Minor effects were observed only at very low doses in the HaCaT non-tumorigenic human keratinocyte cells. RNA-Seq bioinformatics analysis revealed the regulatory impact of arborvitae essential oil on genes enriched in the following pathways: proteasome, adherens junctions, nucleocytoplasmic transport, cell cycle, proteoglycans in cancer, protein processing in the endoplasmic reticulum, ribosome, spliceosome, mitophagy, cellular senescence, and viral carcinogenesis, among others, in both cell lines. It is worth noting that the ribosome and spliceosome KEGG pathways are the most significantly enriched pathways in HeLa and SiHa cells. Arborvitae essential oil shows potential as a cytotoxic and antiproliferative agent against cervical cancer cells, exerting its cytotoxic properties by regulating many KEGG pathways.

Prediction of First-in-Human Dose of Chimeric Antigen Receptor-T (CAR-T) Cells from Mice.

In this study, six scaling methods were evaluated for the prediction of FIH dose for CAR-T cells. The methods were body weight-based fixed exponents such as 1.0 and 0.75, human equivalent dose (HED) using exponents 0.33, two modified HED methods such as using total animal dose (in place of per kg basis) and body surface area in place of body weight using total animal dose with exponent 0.33 and a physiological factor derived from physiological parameters. The FIH doses of six CAR-T cells were predicted in this study. The predicted human doses were compared with the recommended human dose by the US-FDA for four CAR-T cell products, and the literature data were used for the remaining two CAR-T cells. The results indicated that the two modified HED methods and physiological factor are the best and reliable methods for the prediction of FIH dose for CAR-T cells. The proposed methods are simple and accurate in their predictive power and can be used on a spreadsheet.

Cell death and DNA damage via ROS mechanisms after applied antibiotics and antioxidants doses in prostate hyperplasia primary cell cultures.

Tumor heterogeneity results in aggressive cancer phenotypes with acquired resistance. However, combining chemical treatment with adjuvant therapies that cause cellular structure and function perturbations may diminish the ability of cancer cells to resist at chemical treatment and lead to a less aggressive cancer phenotype. Applied treatments on prostate hyperplasia primary cell cultures exerted their antitumor activities through mechanisms including cell cycle blockage, oxidative stress, and cell death induction by flow cytometry methods. A 5.37 mM Chloramphenicol dose acts on prostate hyperplasia cells by increasing the pro-oxidant status, inducing apoptosis, autophagy, and DNA damage, but without ROS changes. Adding 6.30 mM vitamin C or 622 µM vitamin E as a supplement to 859.33 µM Chloramphenicol dose in prostate hyperplasia cells determines a significant increase of ROS level for a part of cells. However, other cells remain refractory to initial ROS, with significant changes in apoptosis, autophagy, and cell cycle arrest in G0/G1 or G2/M. When the dose of Chloramphenicol was increased to 5.37 mM for 6.30 mM of vitamin C, prostate hyperplasia cells reacted by ROS level drastically decreased, cell cycle arrest in G2/M, active apoptosis, and autophagy. The pro-oxidant action of 1.51 mM Erythromycin dose in prostate hyperplasia cell cultures induces changes in the apoptosis mechanisms and cell cycle arrest in G0/G1. Addition of 6.30 mM vitamin C to 1.51 mM Erythromycin dose in hyperplasia cell cultures, the pro-oxidant status determines diminished caspase 3/7 mechanism activation, but ROS level presents similar changes as Chloramphenicol dose and cell cycle arrest in G2/M. Flow cytometric analysis of cell death, oxidative stress, and cell cycle are recommended as laboratory techniques in therapeutic and diagnostic fields.

Exploring antiviral activity of Betanin and Glycine Betaine against dengue virus type-2 in transfected Hela cells

Dengue virus (DENV) infection is a worldwide public health concern infecting approximately 400 million individuals and about 40,000 mortalities yearly. Despite this, no licensed or readily available antiviral medication is currently available specifically for DENV infection, and therapy is typically symptomatic. Therefore, the objective of the study was to investigate the antiviral activity of Beta vulgaris L. phytoconstituents against DENV-2 targeting NS3 protein. The antiviral activity of phytochemicals was examined through virtual ligand-based screening, antiviral inhibition and dosage response assays, western blotting analysis and MD simulations. We conducted toxicological, and pharmacokinetic analysis to assess plant-based natural compound's efficacy, safety, and non-toxic doses. Molecular docking and MD simulation results revealed that the nonstructural protein-3 (NS3) might prove as a funamental target for Betanin and Glycine Betaine against Dengue virus. Betanin and Glycine betaine were initially studied for their non-toxic doses in HeLa, CHO, and Vero cells via MTT assay. HeLa cells were transiently transfected with cloned vector pcDNA3.1/Zeo(+)/DENV-2 NS3 along with non-toxic doses (80 μM–10 μM) of selected phytochemicals. The dose-response assay illustrated downregulated expression of DENV-2 NS3 gene after administration of Betanin (IC50 = 4.35 μM) and Glycine Betaine (IC50 = 4.49 μM). Dose response analysis of Betanin (80 μM–10 μM) depicted the significant inhibition of NS3 protein expression as well. These results suggested downregulated expression of DENV-2 NS3 at mRNA and protein level portraying the DENV replication inhibition. Based on our study findings, NS3 protease is depicted as distinctive DENV-2 inhibitor target. We will channel our study further into in vitro characterization employing the mechanistic study to understand the role of host factors in anti-flavi therapeutic.

Toxicity of CuFe and Co-doped CuFe layered double hydroxides on algae: Insights into synthesis, growth inhibition, morphological changes, and antioxidative enzyme responses

Layered double hydroxides (LDH) are widely used in a variety of industries due to their unique structural characteristics. It is essential to comprehend the environmental behavior and toxicological impacts of these substances to address potential risks caused by LDH release into the environment. In this study, CuFe and Cobalt (Co)-doped CuFe LDHs were synthesized and their toxicities to Chlorella vulgaris were investigated. In the scanning electron microscope images, the Co-doped and undoped catalysts appeared as uniformly dispersed flakes. The X-ray diffraction pattern of the Co-doped CuFe LDH confirmed the successful incorporation of Co into the crystalline lattice of the LDH. The growth of C. vulgaris was negatively affected by the presence of 1 mg/L LDHs, with membrane damage and cell wrinkling observed with 20 mg/L. The CuFe LDH-exposed algae exhibited a significantly greater decline in chlorophyll content compared to that of the Co-doped LDH-exposed algae. However, superoxide dismutase activity was elevated in algal cells exposed to the Co-doped CuFe LDH. Catalase activity increased up to 20 mg/L, followed by a decline at higher doses in CuFe LDH-exposed cells. From an ecological perspective, the lack of increased toxicity after Co doping is favorable for aquatic life. The extensive characterization, together with a rigorous toxicity assessment, provides new information about the environmental safety of cobalt doping to aid in the development of safer and more sustainable LDH-based products.

Propofol orchestrates long non-coding RNAs in MCF7 cells, unraveling new avenues for breast cancer intervention

Long non-coding RNAs (lncRNAs) play a dynamic role in gene expression regulation and serve as potential therapeutic targets in breast cancer. The anticancer effect of propofol, an anesthetic agent, has been proven, but its interaction with lncRNAs has not been adequately investigated. This study aims to reveal the interactions between propofol and lncRNAs and contribute to the understanding of its therapeutic potential in the treatment of breast cancer. We evaluated the effects of propofol on cell viability, apoptosis, and mitochondrial membrane potential in MCF7 cells. The study used qRT-PCR to analyze cancer-related lncRNA expressions following propofol treatment; this was supported by RNA-RNA interaction predictions and in silico functional analysis using selected datasets and the R cluster Profiler GSEABase package. Propofol showed a cytotoxic effect at higher doses in MCF7 breast cancer cells, inducing necrosis. Propofol regulated (IGF2-AS, MRPL23-AS1, PANDAR, HULC) and down-regulated (IWT1-AS, HOXA-AS2, H19, GACAT1, MIAT) the expression levels of various lncRNAs in MCF7 cells. Our research revealed complex interactions of MALAT1 lncRNA with both upregulated and downregulated genes. Additionally, three rRNA genes (LSU-rRNA, RNA45SN3, and SSU-rRNA) were identified to interact with both groups of lncRNAs. Propofol potentially targets chemotherapy resistance by regulating UCA1, LINC-RoR1, and MEG3. Wikipathways' pathway enrichment analysis identified two downregulated lncRNAs, UCA1 and LINC-RoR1, and an upregulated MEG3, implicated in lncRNA-mediated chemotherapeutic resistance mechanisms. Our study illuminates the intricate interplay of lncRNAs and their potential contribution to propofol's anti-cancer effects in breast cancer, offering new avenues for therapeutic exploration and advancement.

The protective effect of venlafaxine on hydrogen peroxide-induced cytotoxicity in C6 glioma cells

Aim: Neurodegeneration is the progressive loss and structural deterioration of neuronal cells. Hydrogen peroxide (H2O2) is formed by dismutation and causes oxidative stress in neuronal cells. Venlafaxine is a drug that increases both serotonin and noradrenaline in the synaptic gap.In this study, the effect of venlafaxine on H2O2-induced cytotoxicity in C6 cells was investigated. Methods: First of all, different doses of venlafaxine (25, 50, and 100 µM) were tried to find the appropriate dose in C6 glioma cells. Then, the effect of venlafaxine on H2O2-induced cytotoxicity in the cells was investigated. For this purpose, cell viability rate, proinflammatory markers IL-1β and TNF-α, and NO and iNOS levels were examined by ELISA kits. Results: H2O2-treated caused cytotoxicity in the C6 glioma cells; when venlafaxine 25, 50, and 100 μM doses were evaluated in terms of cell viability, it was observed that the 100 μM venlafaxine applied group significantly increased cell viability compared to the other groups. When we look at the levels of IL-1β and TNF-α, it is observed that there is an increase in the H2O2 applied group and a significant decrease in the venlafaxine (100 μM) applied group. It was observed that NO and iNOS levels increased in the H2O2 applied group compared to the other groups. It was observed that Venlafaxine treatment reduced the increased NO and iNOS levels caused by H2O2. Conclusion: The study results showed that venlafaxine may have a protective effect on H2O2-induced cytotoxicity in C6 glioma cells.

Bryonia aspera Root Extracts Induce Programmed Cell Death in Selected Cell Lines of Glioblastoma, Ovarian, and Breast Cancer

Background. Medicinal plants have been remarkable sources of current chemotherapeutic agents. Ethnobotanical utilization of Bryonia species goes back to the old era, and contemporary but preliminary studies have evidenced the anticancer effects of this kind of plant. Methods. The MTT assay was used to investigate the cytotoxicity of a range of concentrations from different extracts of Bryonia aspera root in cancer and noncancer cells. The apoptosis was assessed using annexinV-FITC/PI flow cytometry assay. The expression of selected hallmark genes from different cell death modalities, including apoptosis, necroptosis, ferroptosis, and pyroptosis, was investigated using the qPCR method. The ROS production was also measured by the fluorescence technique. Results. Compared to the normal cells, all three extracts could induce significant cell death in lower doses in breast, ovarian, and glioblastoma cancer cells. Flow cytometry and gene expression studies revealed that different extracts of Bryonia aspera tend to induce different types of cell death in the selective cancer cell lines. ROS production was not impacted significantly by any of those three extracts in none of the cancer cells. Conclusion. The findings showed that all three extracts of Bryonia aspera root contain biologically active compounds that induce different types of programmed cell death in the investigated breast, ovarian, and glioblastoma cancer cells in concentrations significantly less than the doses affecting normal cells.

Abstract 1875: Enhanced lung cancer treatment using AOH1996, a potent PCNA inhibitor

Abstract AOH1996 targets a cancer-associated form of PCNA (caPCNA) by altering the protein-protein interface between caPCNA and its many binding partners. AOH1996 does this by inserting into the PCNA-interacting protein-box (PIP-box) pocket that is, in part, defined by the interdomain connecting loop (IDCL), the site of interaction between PCNA and most of its many binding partners. As a result, AOH1996 inhibits PCNA functions such as DNA replication, DNA repair, and transcription-replication conflict (TRC) resolution leading to cell cycle arrest and apoptosis. These effects are cancer specific and AOH1996 has little effect on non-cancerous cells even at 6-fold the effective dose in cancer cells. AOH1996 was developed in our lab and is currently in Phase I human clinical trials. Six patients have been enrolled in the trial and have not experienced AOH1996-related toxicities as the doses have been escalated to levels found to be efficacious in preclinical animal studies. Resistance to EGFR tyrosine kinase inhibitors (TKIs) poses a significant challenge in the treatment of non-small cell lung cancer (NSCLC) patients harboring EGFR activating mutations. Initially, EGFR TKIs like gefitinib, erlotinib, afatinib and osimertinib deliver remarkable responses, inducing tumor regression and improving overall survival. However, the emergence of resistance mechanisms, such as secondary EGFR mutations and bypass signaling pathway activation, hampers the efficacy of these drugs over time. The functional relationship between PCNA and EGFR has implications for the treatment of lung cancer patients with activating EGFR mutations. Membrane-localized EGFR modulates the PCNA-centric process of DNA replication through signaling pathways such as the Ras-Raf-Mek-Erk, PI3K/AKT, and PLCγ1/PKC signaling pathways. Nucleus-localized EGFR enhances PCNA stability by phosphorylating PCNA on tyrosine 211 (Y211), which prevents its degradation. In addition, phosphorylation at Y211 by EGFR results in decreased association of PCNA with MutS, which results in reduced mismatch repair (MMR). In this study, we combine AOH1996 and osimertinib as an innovative approach to treating NSCLC cancers with activating EGFR mutations and find that the drug combination: 1.) enhances the killing of NSCLC cell lines with activating EGFR mutations and acquired resistance to TKIs; 2.) enhances the destabilization of PCNA on chromatin; and 3.) causes changes in the localization and colocalization of PCNA and EGFR. We conclude that the combination of AOH1996 and osimertinib holds much potential as an improved treatment regimen for lung cancer patients with activating EGFR mutations. Citation Format: Robert G. Lingeman, Robert J. Hickey, Linda H. Malkas. Enhanced lung cancer treatment using AOH1996, a potent PCNA inhibitor [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 1875.

Assessing Radiation Effects on Chemo-Treated BT20 and 4T1 Breast Cancer, and Neuroblastoma Cell Lines: A Study of Single and Multiple-Cell Ionization via Infrared Laser Trapping

Background: Our study aimed to assess the radiation sensitivity of BT20, a human breast tumor cell line, using the laser-trapping technique and compare it with N2a and 4T1 cells. Additionally, we investigated the impact of the antitumor compound 2-Dodecyl-6-methoxycyclohexa-2,5-diene-1,4-dione (DMDD) on radiation sensitivity. Methods and Materials: We employed laser trapping to calculate both the threshold ionization energy (TIE) and threshold radiation dose (TRD) for BT20, N2a, and 4T1 cells. We assessed the effect of DMDD on BT20 cells’ radiosensitivity and conducted comparisons across these cell lines. Results: Our findings reveal that DMDD significantly enhances the radiosensitivity of BT20 breast carcinoma cells. Moreover, we observed distinct trends in TIE and TRD across the three cell lines, with differences attributed to variations in cell size and composition. When multiple cell ionizations were considered, a notable reduction in TRD was observed, implicating factors such as the chain effect of ionizing radiation and the influence of DMDD. The study found that TIE increased with the number of cells in the trap while TRD consistently decreased across all three cell lines, suggesting comparable radiation sensitivity, and oligostilbene treatment further reduced TRD, presenting the potential for enhancing therapeutic ratios in cancer treatment. Conclusion: The antitumor compound DMDD enhances the radiosensitivity of BT20 breast carcinoma cells, highlighting its potential in cancer treatment. Furthermore, our study underscores the impact of cell size and multiple-cell ionizations on TRD. Leveraging laser trapping techniques, biocompatible nanoparticles, and advanced optical tweezers opens promising avenues for personalized and effective cancer therapy approaches.

Palladium-103 (103Pd/103mRh), a promising Auger-electron emitter for targeted radionuclide therapy of disseminated tumor cells - absorbed doses in single cells and clusters, with comparison to 177Lu and 161Tb.

Early use of targeted radionuclide therapy (TRT) to eradicate disseminated tumor cells (DTCs) might offer cure. Selection of appropriate radionuclides is required. This work highlights the potential of 103Pd (T1/2 = 16.991 d) which decays to 103mRh (T1/2 = 56.12 min) then to stable 103Rh with emission of Auger and conversion electrons. Methods: The Monte Carlo track structure code CELLDOSE was used to assess absorbed doses in single cells (14-μm diameter; 10-μm nucleus) and clusters of 19 cells. The radionuclide was distributed on the cell surface, within the cytoplasm, or in the nucleus. Absorbed doses from 103Pd, 177Lu and 161Tb were compared after energy normalization. The impact of non-uniform cell targeting, and the potential benefit from dual-targeting was investigated. Additional results related to 103mRh, if used directly, are provided. Results: In the single cell, and depending on radionuclide distribution, 103Pd delivered 7- to 10-fold higher nuclear absorbed dose and 9- to 25-fold higher membrane dose than 177Lu. In the 19-cell clusters, 103Pd absorbed doses also largely exceeded 177Lu. In both situations, 161Tb stood in-between 103Pd and 177Lu. Non-uniform targeting, considering four unlabeled cells within the cluster, resulted in moderate-to-severe dose heterogeneity. For example, with intranuclear 103Pd, unlabeled cells received only 14% of the expected nuclear dose. Targeting with two 103Pd-labeled radiopharmaceuticals minimized dose heterogeneity. Conclusion: 103Pd, a next-generation Auger emitter, can deliver substantially higher absorbed doses than 177Lu to single tumor cells and cell clusters. This may open new horizons for the use of TRT in adjuvant or neoadjuvant settings, or for targeting minimal residual disease.

Assessment of the In Vivo Reprotoxicity of Isotretinoin in Sprague-Dawley Male Rat.

Isotretinoin (ISO) belongs to a family of drugs called retinoids. It is the most effective drug prescribed by dermatologists for the treatment of the inflammatory disease, acne vulgaris. A significant barrier to the use of ISO has worries regarding its adverse effect profile. Despite the well-recognized reproductive toxicity and teratogenicity in females, there is no warning related to the use by male patients in the medication prospectus. Current data on the effects on human male fertility is contradictory and inconclusive. This study was undertaken to investigate the potential effects of ISO oral doses in the Sprague-Dawley male rat germ cells using the sperm morphology assay. Also, the serum levels of the follicle-stimulating hormone (FSH), luteinizing hormone (LH), and testosterone were measured. The rat groups were given varying ISO doses via gastric gavage for seven consecutive days. The epididymis sperm specimens were microscopically examined for the following reproductive toxicity parameters: sperm concentration, examined viability, motility, and morphology. The serum FSH, LH, and testosterone levels were measured by using the corresponding enzyme-linked immunosorbent assay (ELISA) kit. The data were analyzed statistically by one-way analysis of variance (ANOVA) followed by the Tukey test at P ≤ 0.05 significance level. The results indicated that the drug did not significantly increase the sex hormone levels but notably affected both the sperm quantity and quality. These observations suggest that ISO was reprotoxic, and future therapies should be further reassessed.

Monte Carlo investigation of S-values for 111In radionuclide therapy

Novel therapeutic strategy in radionuclide therapy use cell-penetrating monoclonal antibodies to carry Auger-emitting radionuclides into the cells. Estimation of dose in normal and tumor cells are important to investigate the efficacy and toxicity of treatment. Monte Carlo simulation is the most suitable method for estimation of absorbed dose at microscopic level. It is therefore useful to carry out Monte Carlo simulation of Auger emitting radionuclides in order to assess the sensitivity of the results with respect to transport approximations generally used in Monte Carlo codes. There are several Auger emitting radionuclides with potential clinical applications, however, based on their half-life 111In is the most suitable for Auger therapeutic purposes and was considered in the present investigation. Geant4 Monte Carlo simulation was performed and specific absorbed dose fraction (or S-values) for 111In were calculated by using different physics model (Standard, Livermore, Penelope and Geant4-DNA) and compared with Medical Internal Radiation Dosimetry (MIRD) S-values. Source was distributed in the cytoplasm (Cy), surface (Cs) and nucleus (N). Average of relative differences (RD) (%) were calculated for self and cross absorbed dose. RD(%) for self-absorption (NßN) were 4.4, 2.36, 6.21 and 1.1 for Standard, Penelope, Livermore and Geant4-DNA respectively. For cross-absorption these values were higher (e.g. for NßCy 15.4, 18.36, 19.21 and 24.8 for Standard, Penelope, Livermore and Geant4-DNA respectively). Cutoff energy considered for electrons and gamma photons affect the results in dose estimation for Auger electrons in Monte Carlo simulation.

Bioactive compound D-Pinitol-loaded graphene oxide-chitosan-folic acid nanocomposite induced apoptosis in human hepatoma HepG-2 cells

In the present study, we formulated a nanocomposite—comprising graphene oxide, chitosan, and folic acid—as a nanocarrier loaded with the phytochemical d-Pinitol and then subjected to spectroscopic and microscopic analyses to assess the shape, size, and structure of the nanocomposite. The nanocomposite was further subjected to cytotoxic analysis against hepatoma HepG-2 and non-malignant Vero cells using the MTT assay. The generation of ROS, disruption of mitochondrial membrane permeability, and induction of cell death were assessed with corresponding fluorescent staining. Finally, the apoptotic proteins Bax, Bcl2, and caspases were quantified using commercial kits. The characterization analysis confirmed the synthesis of the agglomerated spherical GO Chitosan D-Pinitol FA nanocomposites with an average particle size of 161.50 nm. The nanocomposite remarkably inhibited the HepG-2 cell viability with an IC50 concentration of 7.5 μg/mL but did not disturb the growth of non-malignant Vero cells. It effectively promoted the generation of ROS, disrupted MMP, and induced apoptosis in hepatoma HepG-2 cells, whereas it rendered minimal cytotoxicity even at high doses in mammalian Vero cells. It also promoted the caspase-3, -9, and Bax expressions but inhibited the Bcl-2 expression in the HepG-2 cells. Our findings suggest that the GO Chitosan D-Pinitol FA nanocomposite is a potent drug-loaded nanocarrier that can effectively induce an anticancer effect in hepatoma HepG-2 cells.

Abstract C108: Enhanced growth inhibition in lung cancer cells through NBF-006 combination therapy with chemotherapy or KRAS inhibitors

Abstract Background: Glutathione-S-transferase P (GSTP) is strongly up-regulated in many cancer types, specifically KRAS-mutant lung and is also a known regulator of oncogenic MAPK and PI3K pathways. In cancer cells, overexpression of GSTP regulates apoptosis by inhibiting Jun-p53 complex in JNK pathway. NBF-006 is a novel lyophilized lipid nanoparticle formulation, encapsulating a small interfering ribonucleic acid (siRNA) designed to inhibit the expression of GSTP in lung tumors. NBF-006 has been very well tolerated with early signs of efficacy in NSCLC patients in a phase 1 dose escalation and expansion study. As NBF-006 is well tolerated and unlikely to have drug-drug interactions, combination strategies are currently being developed. Due to the detoxifying role of GSTP, combination with chemo agents used to treat NSCLC may increase the sensitivity of cancer cells resulting in apoptosis and/or reduced resistance. As GSTP and KRAS enhance tumor growth and survival, inhibition of GSTP and KRAS may be synergistic. Methods: Anti-tumor activity of GSTP siRNA (NDT-05-1040), when combined with chemo agents, KRAS inhibitors or immune cells was assessed in a panel of KRAS mutant NSCLC lines. For chemo agents and KRAS inhibitors, NDT-05-1040 combinations was evaluated using Combenefit software and Loewe model. Results: Combination of NDT-05-1040 with chemotherapy drugs including paclitaxel, gemcitabine, cisplatin, and pemetrexed controlled tumor cell growth significantly better than either NBF-006 alone or the chemo treatment alone. In A549, H1373, H358, and H1792 lines, at low doses paclitaxel combined with NDT-05-1040 and resulted in synergistic cell killing (Loewe scores >20). Gemcitabine & Cisplatin combination with NDT-05-1040 in H1373 cells further improved cell killing by 5-fold (Loewe scores >20). Pemetrexed combined with NDT-05-1040 and led to additive cell killing in A549 (Loewe score >10). Combination of NDT-05-1040 with KRASG12C inhibitors, Sotorasib & Adagrasib were conducted in H358, H1222, and MiaPaca-2. Cell viability assays demonstrated that the combination resulted in strong synergistic, anti-proliferative effects at low doses in H358 cells (the greatest Loewe scores for Adagrasib and Sotorasib: 53 & 26, respectively). Conclusion: Results from the current studies suggest that NBF-006, a very well tolerated monotherapy, can be used in combination with other drugs currently indicated for KRAS-mutant NSCLC to achieve more potent antitumor effects. Citation Format: Cima Cina, Akinori Sugiyama, Murali Kuracha, Nazia Abbasi, Dominic Medina, Hiroyuki Tanaka, Sonya Zabludoff. Enhanced growth inhibition in lung cancer cells through NBF-006 combination therapy with chemotherapy or KRAS inhibitors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C108.

#236 : Alleviating Effects of EGCG on Ovarian Hyperstimulation Syndrome and the Potential Modulating Mechanism

Background and Aims: Ovarian hyperstimulation syndrome (OHSS) is one of the most severe complications of COH during IVF treatment. Increased capillary permeability is a hallmark of the pathophysiology of OHSS, vascular endothelial growth factor (VEGF) is an important mediator in OHSS, and studies have established a correlation between serum VEGF levels and the severity of the OHSS. (-)-epigallocatechin-3-gallate (EGCG), the most prevalent and biologically active polyphenolic catechin in green tea, has been reported to produce a number of beneficial effects on humans. Numerous studies have shown that in many pathological processes, EGCG could inhibit VEGF and its receptor expression and have an angiogenesis effect. Herein, EGCG might have a therapeutic effect on OHSS. Methods: We investigated the role of EGCG in OHSS both in vitro and in vivo. Primary human granulosa-lutein (hGL) cells and a human granulosa-like tumor (KGN) cell line were cultured and treated for 24 hours with various concentrations of EGCG. In SD rats, an animal OHSS model was established by injecting pregnant mare serum gonadotropin (PMSG) and randomly assigning them to receive vehicle only or EGCG for three days. For comparison, all experiments were repeated 3-8 times. The effect of EGCG on KGN and hGL cells was determined by MTT assay. The body weight of rats was measured every day, and the ovary size and weight were measured after removal. Evans-blue was used to determine permeability. ELISA was used to measure serum estrogen and VEGF levels in rats. RNA and protein expressions of VEGF, VEGFR-2, TGF-[Formula: see text]1, and T[Formula: see text]R II were detected by qPCR and Western-blotting and immunostaining in vitro and in vivo experiments. Results: Our research found that giving rats EGCG prevented the development of OHSS, as evidenced by histological examination, ovarian weight, and morphology. When compared to the OHSS group, the EGCG treatment group had significantly lower ovary weight (147.9 vs 206.5 mg) (Fig. 1). Furthermore, when compared to OHSS rats, EGCG-treated rats have lower ovarian VEGF expression as measured by IHC and RT-qPCR. VEGF and E2 protein levels in the serum of the EGCG treatment group are significantly lower. EGCG exerted inhibitory effects on cell growth only in high dose (50 uM) and longtime (48 h) treatment in KGN and hGL cells. In KGN cells and hGL cells, EGCG significantly reduces the expression of VEGF and TGF-[Formula: see text] at the RNA and protein levels. Furthermore, EGCG inhibits TGF-[Formula: see text]1-induced VEGF production and secretion in KGN cells by suppressing TGF-[Formula: see text] expression and its traditional Smad signaling pathway. EGCG also downregulates VEGF expression through the 67-kDa laminin receptor-mediated PKA-CREB pathway (Fig. 2). Conclusions: EGCG inhibited VEGF via the TGF-[Formula: see text]1 classical-SMAD pathway and the 67LR-mediated CREB pathway, reducing ovarian inflammation and slowing the progression of OHSS in a rat model.

Nonivamide inhibits proliferation of human corneal epithelial cells by inducing cell cycle arrest and oxidative stress

Nonivamide, an agonist of transient receptor potential vanilloid type 1 (TRPV1), is widely used as a riot control agent, police incapacitant spray and pesticide. Although generally considered non-fatal, eye discomfort and even ocular injuries caused by such products are common. Little research has been conducted on the effects of nonivamide on corneal epithelial cells. Cell viability, impedance, flow cytometry, western blotting, and real-time fluorescence analyses were performed to investigate the effects of nonivamide on human corneal epithelial cells (HCE-T cells). We found that nonivamide impaired proliferation at subtoxic doses (100 μM and 200 μM) in HCE-T cells. Next, we described the mechanisms of action of nonivamide. Nonivamide caused cell cycle arrest by increasing p21 and decreasing cyclin D1. TRPV1 was activated by nonivamide, leading to an influx of Ca2+. Enhanced Ca2+ influx partially contributed to oxidative stress. Mitochondrial membrane potential (MMP) also decreased. All combined stress resulted in the inhibition of cell proliferation in HCE-T cells. In summary, nonivamide inhibited the proliferation of HCE-T cells at sub-toxic doses by inducing cell cycle arrest and oxidative stress. Our data demonstrate the corneal toxicity of nonivamide and explain the mechanisms underlying nonivamide-induced corneal injury.

Cholesterol deficiency as a mechanism for autism: A valproic acid model.

Dysregulated cholesterol metabolism represents an increasingly recognized feature of autism spectrum disorder (ASD). Children with fetal valproate syndrome caused by prenatal exposure to valproic acid (VPA), an anti-epileptic and mood-stabilizing drug, have a higher incidence of developing ASD. However, the role of VPA in cholesterol homeostasis in neurons and microglial cells remains unclear. Therefore, we examined the effect of VPA exposure on regulation of cholesterol homeostasis in the human microglial clone 3 (HMC3) cell line and the human neuroblastoma cell line SH-SY5Y. HMC3 and SH-SY5Y cells were each incubated in increasing concentrations of VPA, followed by quantification of mRNA and protein expression of cholesterol transporters and cholesterol metabolizing enzymes. Cholesterol efflux was evaluated using colorimetric assays. We found that VPA treatment in HMC3 cells significantly reduced ABCA1 mRNA, but increased ABCG1 and CD36 mRNA levels in a dose-dependent manner. However, ABCA1 and ABCG1 protein levels were reduced by VPA in HMC3. Furthermore, similar experiments in SH-SY5Y cells showed increased mRNA levels for ABCA1, ABCG1, CD36, and 27-hydroxylase with VPA treatment. VPA exposure significantly reduced protein levels of ABCA1 in a dose-dependent manner, but increased the ABCG1 protein level at the highest dose in SH-SY5Y cells. In addition, VPA treatment significantly increased cholesterol efflux in SH-SY5Y, but had no impact on efflux in HMC3. VPA differentially controls the expression of ABCA1 and ABCG1, but regulation at the transcriptional and translational levels are not consistent and changes in the expression of these genes do not correlate with cholesterol efflux in vitro.

Evaluate the effects of kilovoltage transmission target X-ray equipment combined with radiosensitizers for Auger electron radiotherapy

Auger electrons are well-suited for targeted radiotherapy due to their ability to transfer high energy over short distances. Auger radiotherapy can be achieved by utilizing the characteristic radiation of transmission X-ray tubes and radiosensitizers to generate numerous Auger electrons. In this study, the Monte Carlo N-Particle (MCNP6) code was employed to simulate the deposition of dose in cells when combining the transmission X-ray tube with various concentrations of radiosensitizers. The results reveal that when the transmission X-ray tube is combined with 70 mg of gold nanoparticles (GNPs), the photoelectron flux is 1.16 × 103 (particles/cm2), which is 3.86 times higher than that observed with 0 mg of GNPs. Furthermore, the Auger electron flux is 3.51 × 102 (particles/cm2), representing a 15.51-fold increase compared to 0 mg of GNPs. The corresponding dose enhancement factor (DEF) is 3.14, representing the most substantial proportional increase observed. This study suggests that the combination of a transmission X-ray tube with appropriate radiosensitizers and concentrations can generate a significant number of photoelectrons and Auger electrons, resulting in a notable radio-enhancing effect. This innovative approach holds promise for the development of Auger radiotherapy as a potential therapeutic process. Future investigations could explore the optimal combination of transmission X-ray tubes and radiosensitizers to further advance the applications of Auger radiotherapy.