HeLa Research Articles

In vitro antioxidant and anticancer potential of intra-cellular ethyl acetate extract of marine-derived fungus Talaromyces tratensis SS10

Marine fungi are well-known for producing structurally distinct secondary metabolites, making them potential sources of novel therapies. The present investigation aims to study the in vitro antioxidant and anticancer potential of intra-cellular crude ethyl acetate extracts of Talaromyces tratensis SS10. In the present study, qualitative and quantitative phytochemical studies of various solvent extracts of T. tratensis have been carried out using standard protocols. Later, ethyl acetate extract of T. tratensis was analyzed for phytochemicals using Gas Chromatography Mass Spectrometry (GC-MS). Further, the antioxidant properties of the T. tratensis ethyl acetate extract have been done by Ferric reducing antioxidant power assay (FRAP). Further, the anticancer potential of this extract has been carried out by MTT assay against human cancer cells such as MDA MB 231, HeLa, and HT-29. Ethyl acetate exhibited a higher yield of chemical extraction than the other solvents used. The GCMS analysis of T. tratensis ethyl acetate extract revealed major phytoconstituents such as N-(1,1-Dimethylpropyl)-2,2,3-trimethylaziridine-1-carboxamide, 1-Undecanol, 5,5 Dimethyl-3-vinyl cyclohex-2-en-1-one, 1,2-Benzenedicarboxylic acid, bis (2-methyl propyl) ester. T. tratensis ethyl acetate extract showed the highest percentage of Fe3+ reduction (48.093±1.469%) at 120 μg/mL, with an IC50 value of 157.26 μg/mL concentration. Furthermore, 100 μg/mL of the extract showed significant cell death rates in cytotoxic assays, indicating a low percentage of viable cells for all three examined cell lines. The T. tratensis ethyl acetate extract has shown a dose-dependent cytotoxic effect against all tested cancer cell lines. The better IC50 value (6.25 μg/mL) was recorded in the case of HeLa cell lines followed by 12.5 μg/mL for both MDA MB 231 and HT-29 cell lines. The presence of bioactive compounds such as Benzeneethanamine, N-[(pentafluorophenyl)methylene]-beta.,3,4-tris[(trimethylsilyl) oxy]-, 1,2-Benzenedicarboxylic acid, bis(2-methylpropyl) ester, and cyclononasiloxane, octadecamethyl- may have contributed to the ethyl acetate extracts’ strong antioxidant and anticancer properties. The current study’s findings show that T. tratensis SS10 has the potential for drug development due to its chemical constituents, which possess various biological activities.

Circ_0006789 promotes cervical cancer development via the miR-615-5p/HSF1 axis

ObjectiveCircular RNAs (circRNAs) are involved in the development of human cancers, including cervical cancer (CC). However, the role and mechanism of circ_0006789 (circSLC25A43) in CC are unclear. The purpose of this study was to investigate the functional role of circ_0006789 in CC.MethodsThe expression of circ_0006789 in CC tissues and cell lines was examined by RT-qPCR. The characterization of circ_0006789 in CC cells was verified by subcellular localisation, actinomycin D assay, and RNase R assay. After circ_0006789 was knocked down in CC cell lines, the proliferation, apoptosis, migration and invasion of CC cells were assessed by CCK-8 method, flow cytometry, and Transwell assay. RIP assay, FISH assay, dual luciferase reporter gene assay and Western blot were used to investigate the regulatory mechanism between circ_0006789, miR-615-5p and heat shock factor 1 (HSF1).Resultscirc_0006789 was upregulated in CC tissues and cell lines. CC cells were inhibited in their proliferation, migration, and invasion, as well as promoted to apoptosis when circ_0006789 was knocked down. It was found that circ_0006789 targeted miR-615-5p, and miR-615-5p expression was inversely correlated with circ_0006789 expression. Furthermore, HSF1 was a target gene of miR-615-5p. Furthermore, the suppressive effects on HeLa cells mediated by circ_0006789 knockdown were counter-balanced when miR-615-5p was knocked down and HSF1 was overexpressed. Mechanistically, circ_0006789 was found to promote CC development by reducing miR-615-5p and increasing HSF1 expressions.Conclusioncirc_0006789 accelerates CC development via the miR-615-5p/HSF1 axis.

Allylthiourea‐mediated self‐healing hydrogels based on poly(vinyl alcohol): Enhanced cell viability/biocompatibility and sustained drug release

AbstractHydrogel‐based materials represent promising candidates for drug delivery. In our research, we synthesized a series of hydrogels—PVATU‐1 (2:1), PVATU‐2 (1:1), and PVATU‐3 (0.5:1)—by adjusting the poly(vinyl alcohol) (PVA) ratio while maintaining a constant ratio of allylthiourea (ATU) monomer and glutaraldehyde cross‐linker, aiming to understand their impacts on gelation. These PVATU hydrogels displayed impressive swelling in neutral conditions and exhibited visible self‐healing capabilities. Notably, PVATU‐1 shows superior mechanical strength among the variants. Surface analysis using AFM and SEM unveiled porous structures within the hydrogels, while thermal stability tests indicated their resilience up to 200°C. DSC analysis revealed minimal variation in glass transition temperatures (Tg), affirming stability. A distinct endothermic peak around 250–350°C across all hydrogels confirmed their semi‐crystalline nature, corroborated by powder XRD. Noteworthy, PVATU‐1 demonstrated optimal loading and releasing efficiency for levofloxacin, with 74.5% loading and 82.8% releasing efficiency within 18 h. Biocompatibility assessments on HeLa cells affirmed the nontoxicity of PVATU hydrogel treatments.

Behaviors of nucleosomes with mutant histone H4s in euchromatic domains of living human cells.

Since Robert Feulgen first stained DNA in the cell, visualizing genome chromatin has been a central issue in cell biology to uncover how chromatin is organized and behaves in the cell. To approach this issue, we have developed single-molecule imaging of nucleosomes, a basic unit of chromatin, to unveil local nucleosome behavior in living cells. In this study, we investigated behaviors of nucleosomes with various histone H4 mutants in living HeLa cells to address the role of H4 tail acetylation, including H4K16Ac and others, which are generally associated with more transcriptionally active chromatin regions. We ectopically expressed wild-type (wt) or mutated H4s (H4K16 point; H4K5,8,12,16 quadruple; and H4 tail deletion) fused with HaloTag in HeLa cells. Cells that expressed wtH4-Halo, H4K16-Halo mutants, and multiple H4-Halo mutants had euchromatin-concentrated distribution. Consistently, the genomic regions of the wtH4-Halo nucleosomes corresponded to Hi-C contact domains (or topologically associating domains, TADs)with active chromatin marks (A-compartment). Utilizing single-nucleosome imaging, we found that none of the H4 deacetylation or acetylation mimicked H4 mutants altered the overall local nucleosome motion. This finding suggests that H4 mutant nucleosomes embedded in the condensed euchromatic domains with excess endogenous H4 nucleosomes cannot cause an observable change in the local motion. Interestingly, H4 with four lysine-to-arginine mutations displayed a substantial freely diffusing fraction in the nucleoplasm, whereas H4 with a truncated N-terminal tail was incorporated in heterochromatic regions as well as euchromatin. Our study indicates the power of single-nucleosome imaging to understand individual histone/nucleosome behavior reflecting chromatin environments in living cells.

Hypoxia-activated prodrug combining site-specific chemotherapy and light-driven photothermal therapy

The development of a highly effective and low toxic drug is very beneficial for precise cancer therapy. Herein, taking advantage of the hypoxic nature of tumors, we carefully designed and synthesized the first hypoxia-activated single-molecule prodrug BDP-CPT-NO2 for fluorescence imaging-guided chemo-photothermal dual-mode therapy. The prodrug contains the chemotherapeutic drug camptothecin (CPT) and a boron dipyrromethene (BDP)-based photothermal agent, which are connected through a nitroreductase (NTR)-responsive linker (a nitro unit), and shows no fluorescence and extremely low cytotoxicity under normoxia. However, the nitro in the prodrug could be reduced to amino by exogenous NTR with nicotinamide adenine dinucleotide (NADH) in PBS buffer solution and endogenous NTR in hypoxic cancer cells, which is followed by a self-immolative reaction that leads to the release of CPT and BDP-COOH. This process is accompanied by significantly enhanced BDP and CPT fluorescence, which are due to NTR-induced ester-to-carboxylate conversion and fluorescence resonance energy transfer (FRET) interruption, respectively. Concomitantly, BDP-CPT-NO2 displays powerful combined chemo-PTT therapeutic efficacy, as evidenced in human cervical cancer HeLa and human liver cancer HepG2 cells, as well as in mice bearing H22 tumors. This study provides a promising strategy for effective and selective cancer treatment by hypoxia-activated prodrugs caged within organic photothermal agents.

Cytocompatible hydrogel derived from dextrin and Poly(N-Vinyl acetamide) toward controlled antimicrobial release

The work aims to develop a biopolymer-dextrin-based three-dimensional crosslinked hydrophilic material for antimicrobial delivery. The copolymer (Dxt-PNVA) has been prepared by in-situ grafting of poly(N-vinylacetamide), followed by cross-linking with N,N'-methylenebisacrylamide with dextrin. By varying the concentrations of the reactants, an optimized copolymer (Dxt-PNVA 5) with the lowest % of crosslinking has been chosen for biomedical applications. The copolymeric gel has been characterized in detail using various techniques, followed by its cytocompatibility study using HeLa Cell lines that confirm the cytocompatible nature of the material. Copolymeric gel is a suitable material for the effective release of two antimicrobials (amoxicillin trihydrate and ornidazole) from tablet formulation for a prolonged period of 24 h. The obtained data demonstrates that the cytocompatible hydrogel has the potential characteristics of a daily dose administration for amoxicillin trihydrate and ornidazole.

New Carbothioamide and Carboxamide Derivatives of 3-Phenoxybenzoic Acid as Potent VEGFR-2 Inhibitors: Synthesis, Molecular Docking, and Cytotoxicity Assessment.

Because of the well-established link between angiogenesis and tumor development, the use of antiangiogenic therapeutics, such as those targeting VEGFR-2, presents a promising approach to cancer treatment. In the current study, a set of five hydrazine-1-- carbothioamide (compounds 3a-e) and three hydrazine-1-carboxamide derivatives (compounds 4a-c) were successfully synthesized from 3-phenoxybenzoic acid. These compounds were specially created as antiproliferative agents with the goal of targeting cancer cells by inhibiting VEGFR-2 tyrosine kinase. The new derivatives were synthesized by conventional organic methods, and their structure was versified by IR, 1HNMR, 13CNMR, and mass spectroscopy. In silico investigation was carried out to identify the compounds' target, molecular similarity, ADMET, and toxicity profile. The cytotoxic activity of the prepared compounds was evaluated in vitro against three human cancer cell lines (DLD1 colorectal adenocarcinoma, HeLa cervical cancer, and HepG2 hepatocellular carcinoma). The effects of the leading compound on cell cycle progression and apoptosis induction were investigated by flow cytometry, and the specific apoptotic pathway triggered by the treatment was evaluated by RT-PCR and immunoblotting. Finally, the inhibitory activities of the new compounds against VEGFR-2 was measured. The designed derivatives exhibited comparable binding positions and interactions to the VEGFR-2 binding site to that of sorafenib (a standard VEGFR-2 tyrosine kinase inhibitor), as determined by molecular docking analysis. Compound 4b was the most cytotoxic compound, achieving the lowest IC50 against HeLa cells. Compound 4b, a strong representative of the synthesized series, induced cell cycle arrest at the G2/M phase, increased the proportion of necrotic and apoptotic HeLa cells, and activated caspase 3. The EC50 value of compound 4b against VEGFR-2 kinase activity was comparable to sorafenib's. Overall, the findings suggest that compound 4b has a promising future as a starting point for the development of new anticancer drugs.

Ras-Targeting Stabilized Peptide Increases Radiation Sensitivity of Cancer Cells.

Radiation therapy is one of the most common treatments for cancer. However, enhancing tumors' radiation sensitivity and overcoming tolerance remain a challenge. Previous studies have shown that the Ras signaling pathway directly influences tumor radiation sensitivity. Herein, we designed a series of Ras-targeting stabilized peptides, with satisfactory binding affinity (KD = 0.13 μM with HRas) and good cellular uptake. Peptide H5 inhibited downstream phosphorylation of ERK and increased radio-sensitivity in HeLa cells, resulting in significantly reduced clonogenic survival. The stabilized peptides, designed with an N-terminal nucleation strategy, acted as potential radio-sensitizers and broadened the applications of this kind of molecule. This is the first report of using stabilized peptides as radio-sensitizers, broadening the applications of this kind of molecule.

DNA Walker-Driven Mass Nanotag Assembly System for Simultaneously Profiling Dual Markers of Oxidative Stress at Different Cellular Locations.

Simultaneous profiling of redox-regulated markers at different cellular sublocations is of great significance for unraveling the upstream and downstream molecular mechanisms of oxidative stress in living cells. Herein, by synchronizing dual target-triggered DNA machineries in one nanoentity, we engineered a DNA walker-driven mass nanotag (MNT) assembly system (w-MNT-AS) that can be sequentially activated by oxidative stress-associated mucin 1 (MUC1) and apurinic/apyrimidinic endonuclease 1 (APE1) from plasma membrane to cytoplasm and induce recycled assembly of MNTs for multiplex detection of the two markers by matrix-assisted laser desorption ionization mass spectrometry (MALDI MS). In the working cascade, the sensing process governs the separate activation of w-MNT-AS by MUC1 and APE1 in diverse locations, while the assembly process contributes to the parallel amplification of the ion signal of the characteristic mass tags. In this manner, the differences between MCF-7, HeLa, HepG2, and L02 cells in membrane MUC1 expression and cytoplasmic APE1 activation were fully characterized. Furthermore, the oxidative stress level and dynamics caused by exogenous H2O2, doxorubicin, and simvastatin were comprehensively demonstrated by tracking the fate of the two markers across different cellular locations. The proposed w-MNT-AS coupled MS method provides an effective route to probe multiple functional molecules that lie at different locations while participating in the same cellular event, facilitating the mechanistic studies on cellular response to oxidative stress and other disease-related cellular processes.

Structurally diverse isoquinoline alkaloids from the barks of Alangium salviifolium (L. f.) Wangerin and their cytotoxicity

Eleven undescribed isoquinoline alkaloids (1−8, 14, 15, and 24), along with 19 analogues (9−13, 16−23, and 25−30) were isolated from the barks of Alangium salviifolium. The structures of the undescribed compounds were elucidated through the analysis of their HR-ESI-MS, 1D and 2D NMR, IR, UV, and X-ray diffraction. The absolute configuration of 8 was established via the ECD calculation. Notably, compounds 1/2 and 3/4 were two pairs of C-14 epimers. The isolated alkaloids were evaluated for their cytotoxicity against various cancer cell lines, including SGC-7901, HeLa, K562, A549, BEL-7402, HepG2, and B16, β-carboline-benzoquinolizidine (14−22) and cepheline-type (24−28) alkaloids exhibited remarkable cytotoxicity, with IC50 values ranging from 0.01 to 48.12 μM. Remarkably, compounds 17 and 21 demonstrated greater cytotoxicity than the positive control doxorubicin hydrochloride. Furthermore, a significant proportion of these bioactive alkaloids possess a C-1′ epimer configuration. The exploration of their structure-activity relationship holds promise for directing future investigations into alkaloids derived from Alangium, potentially leading to novel insights and therapeutic advancements.

Design and synthesis of N-aryl-2-trifluoromethyl-quinazoline-4-amine derivatives as potential Werner-dependent antiproliferative agents.

To discover new Werner (WRN) helicase inhibitors, a series of N-aryl-2-trifluoromethyl-quinazoline-4-amine derivatives were designed and synthesized through a structural optimization strategy, and the anticancer activities of 25 new target compounds against PC3, K562, and HeLa cell lines were evaluated by the MTT assay. Some of these compounds exhibited excellent inhibitory activity against three different cancer cell lines. Compounds 6a, 8i, and 13a showed better antiproliferative activity against K562 cells, with IC50 values of 3871.5, 613.6 and 134.7nM, respectively, than did paclitaxel (35.6nM), doxorubicin (2689.0nM), and NSC 617145 (20.3nM). To further verify whether the antiproliferative activity of these compounds is dependent on WRN, PC3 cells overexpressing WRN (PC3-WRN) were constructed to further study their antiproliferative potency in vitro, and the inhibition ratio and IC20 values showed that compounds 6a, 8i, and 13a were more sensitive to PC3-WRN than were the control group cells (PC3-NC). The IC20 ratios of compounds 6a, 8i, and 13a to PC3-NC and PC3-WRN were 94.3, 153.4 and 505.5, respectively. According to the docking results, the compounds 6a, 8i, and 13a overlapped well with the binding pocket of 6YHR. Further study demonstrated that among the tested compounds, 13a was the most sensitive to PC3-WRN. In summary, our research identified a series of N-aryl-2-trifluoromethyl-quinazoline-4-amine derivatives as potential WRN-dependent anticancer agents.

Mesencephalic astrocyte-derived neurotrophic factor inhibits cervical cancer progression via regulating macrophage phenotype.

Cervical cancer is a common gynecologic malignant tumor, but the critical factors affecting cervical cancer progression are still not well demonstrated. Mesencephalic astrocyte-derived neurotrophic factor (MANF) has been widely recognized as an anti-inflammatory factor to regulate macrophage polarization. In this study, the effect and mechanism of MANF on cervical cancer were preliminarily explored. Kaplan-Meier curve was used to show the overall survival time of the involved cervical cancer patients with high and low MANF expression in cervical cancer tissues. MANF was highly expressed in peritumoral tissues of cervical carcinoma by using immunohistochemistry and western blot. MANF mRNA level was detected by using qRT-PCR. Dual-labeled immunofluorescence showed MANF was mainly expressed in macrophages of cervical peritumoral tissues. Moreover, MANF-silenced macrophages promoted HeLa and SiHa cells survival, migration, invasion and EMT via NF-κB signaling activation. The results of tumor formation in nude mice indicated MANF-silenced macrophages promoted cervical tumor formation in vivo. Our study reveals an inhibitory role of MANF in cervical cancer progression, indicating MANF as a new and valuable therapeutic target for cervical cancer treatment.

Design, Synthesis and Biological Evaluation of Novel Gypsogenin Derivatives as Potential Anticancer and Antimicrobial Agents.

Natural compounds are important sources for the treatment of chronic disorders such as cancer and microbial infectious disorders. In this research, Gypsogenin and its derivatives (2 a-2 f) have been tested against different cancer cell lines (MCF-7, HeLa, Jurkat and K562 cell lines) and further analyzed for cell proliferation, cell death type, and for act of the mechanism. Cell proliferation was determined by the MTT method and cell death types were analyzed with HO/PI staining. Fibroblast Growth Factor 1 (FGF-1), Interleukin 1 (IL-1), Interleukin 6 (IL-6), and Tumor Necrosis Factor Alpha (TNF-α), key players in breast cancer development and progression, were determined by Elisa kits. Results showed that compound 2 e inhibited the MCF-7 cell line proliferation with an IC50 value of 0.66±0.17 μM with 93.38 % apoptosis rate. Compound 2 e also decreased FGF-1, IL-1, IL-6, and TNF-α levels. Molecular docking studies performed in the binding site of FGFR-1 indicated that compound 2 e formed key hydrogen bonding with Arg627 and Asn568. Besides, compounds 2 a-2 f were evaluated for their antimicrobial activities against gram-negative and gram-positive bacteria and C. albicans via the microdilution method. Overall, compound 2 e stands out as a potential anticancer agent for future studies.

Evaluation of the potential of Stichopus Herrmanni extract in inhibiting cervical cancer cell proliferation

BackgroundSea cucumbers, particularly Stichopus herrmanni, are known for their medicinal value in Asian traditional medicine owing to the abundant presence of saponins, terpenoids, and phenols. Investigations on these bioactive compounds have revealed their cytotoxic propensity against various cancer cell lines, indicating their therapeutic potential. However, limited research exists on the potential application of sea cucumber extracts specifically in cervical cancer. PurposeThis study aimed to identify the bioactive components of Stichopus herrmanni extract, determine their association with potential targets in cervical cancer, and evaluate their cytotoxic effects on cervical cancer cells. Study DesignThis experimental study employed both in vitro and in silico methods to evaluate the potential cytotoxic effects of Stichopus herrmanni extracts on cervical cancer cells. MethodsLiquid Chromatography-Mass Spectrometry (LC-MS) was used to identify and quantify the bioactive constituents of Stichopus herrmanni crude extract. An in silico approach was used to identify the active components, potential targets, and signaling pathways of the extract. Furthermore, the MTT assay was used to determine the cytotoxicity of the extract. ResultsLC-MS analysis identified the presence of rengyol, eucommiol, ganoderic acid, and 6-isoinosine in the extracts. An in silico study based on structural analysis identified ganoderic acid and isoinosine as crucial active components capable of regulating majority of the targets associated with cervical cancer. Overlapping targets, namely, CASP3, CAT, FASLG, IL24, TP53, TP53BP1, ALB, BDNF, and COX2 between cervical cancer and the extract highlighted new therapeutic prospects for cervical cancer following protein-protein interaction network screening. The in vitro cytotoxic effects of the extracts were established in HeLa cells. ConclusionResearch indicates that Stichopus herrmanni may be a valuable source of bioactive compounds with potential applications in the treatment of cancer and the induction of apoptosis

Effect of RNF113A deficiency on oxidative stress-induced NRF2 pathway

ABSTRACT The ring finger protein 113A (RNF113A) serves as an E3 ubiquitin ligase and a subunit of the spliceosome. Mutations in the RNF113A gene are associated with X-linked trichothiodystrophy (TTD). However, the cellular roles of RNF113A remain largely unknown. In this study, we performed transcriptome profiling of RNF113A knockout (KO) HeLa cells using RNA sequencing and revealed the upregulation of NRF2 pathway-associated genes. Further analysis confirmed that the KO of RNF113A promotes nuclear localization of the NRF2 protein and elevates the mRNA levels of NRF2 target genes. RNF113A KO cells showed high levels of intracellular reactive oxygen species (ROS) and decreased resistance to cell death following H2O2 treatment. Additionally, RNF113A KO cells more sensitively formed stress granules (SGs) under arsenite-induced oxidative stress. Moreover, RNF113A KO cells exhibited a decrease in glutathione levels, which could be attributed to a reduction in GLUT1 expression levels, leading to decreased glucose uptake reactions and lower intracellular glucose levels. These alterations potentially caused a reduction in ROS scavenging activity. Taken together, our findings suggest that the loss of RNF113A promotes oxidative stress-mediated activation of the NRF2 pathway, providing novel insights into RNF113A-associated human diseases.

An automated approach for hemocytometer cell counting based on image-processing method

Evaluating cell viability plays an important role in routine cell culture, toxicology, and drug development. Trypan blue stain is one of the simplest assays that are used to determine the number of viable cells in a cell suspension. The stained cell suspension can then be counted under a microscope using a hemocytometer. Hemocytometers have a fixed volume, making it possible to calculate the concentration of cells in the sample and measure viability. Most of the related studies are conducted using manual counting methods. However, manual counting is time-consuming and laborious. While various automated cell counting devices have been proposed, they tend to be too expensive for widespread deployment. Although low-cost automated cell counting methods presented in the literatures provide high accuracy and cost-effective alternatives, only cells in the field of view of microscope with unknown volume are counted. This will make the concentration inestimable. Accordingly, the present study proposes a low-cost but accurate method for hemocytometer cell counting using an image-processing approach. The accuracy of the proposed method is demonstrated by comparing the cell counting results obtained for human cervical carcinoma (HeLa) cells with the manual counts. It is shown that the proposed approach has a mean absolute percentage error of less than 6.26 % for samples containing a mixture of live and dead cells. Moreover, the proposed system is equipped with a simple and intuitive human machine interface such that the counting task can be performed by individuals with no prior experience in the image-processing field.

Conditional Cell Penetration of Masked CPPs by an ADEPT-like Approach.

The intracellular delivery of cargos via cell penetrating peptides (CPPs) holds significant promise as a drug delivery vehicle, but a major issue is their lack of cell type specificity, which can lead to detrimental off-target effects. We use an ADEPT-like concept to introduce conditional and selective activation of cellular uptake by using the lysine-rich, cationic, and amphiphilic L17E peptide as a model CPP. By masking the lysine residues of the L17E peptide with enzyme-cleavable acetyl protecting groups, the delivery of the covalently conjugated fluorophore TAMRA to HeLa cells was diminished. Recovery of cellular uptake could be achieved by deacetylation of the masked acetylated L17E peptide using the NAD-dependent sirtuin 2 (SirT2) deacetylase in vitro. Finally, trastuzumab-SirT2 and anti-B7H3-SirT2 antibody-enzyme conjugates were generated for the conditional and selective delivery of a cryptophycin cytotoxin by the L17E peptide. While the masked peptide still demonstrated some cytotoxicity, selective cell killing mediated by the antibody-enzyme conjugates was observed.

The influence of an innovative antibacterial drug of the thiadiazinone class on the virulence factors of bacteria of the phylum Pseudomonadota, which chronically infect patients with cystic fibrosis

Introduction. Infections of the lower respiratory tract by bacteria of the Pseudomonadota phylum: Pseudomonas aeruginosa, Burkholderia spp., Achromobacter spp. are critical to the quality and life expectancy of patients with cystic fibrosis (CF). When the infection is chronic, eradication of bacteria with existing antibacterial drugs is practically impossible. To explore alternative drugs, trials are needed on bacteria isolated from CF patients and characterized using genomic approaches. The objective of our study was a comparative analysis of virulence factors of 6 isolates of bacteria of the Pseudomonadota phylum and testing the efficacy of the innovative drug Fluorothiazinone (FT) in suppressing the pathogenicity of bacteria in vitro. Materials and methods. Isolates of A. ruhlandii ST36, A. xylosoxidans ST555, B. cepacia ST2140, B. gladioli ST2141, P. aeruginosa ST859 and ST198 were examined using whole-genome sequencing and bioinformatics analysis to search for resistance and virulence determinants. The FT drug was tested for its effect on bacteria in in vitro experiments on cytotoxicity on HeLa cells, motility and biofilm formation. Results. Genomic studies have confirmed the arsenal of resistance determinants, especially the efflux systems of bacteria isolated from patients with CF, and the diversity of virulence factors, among which we identified factors in the categories of motility, signals of quorum-sensing systems, secretion systems, exotoxins, as the most essential for the adaptation of bacteria to conditions of the lower respiratory tract. In vitro tests of the FT drug showed its effectiveness in suppressing cytotoxicity (2.6–4.0 times), motility (2.0–3.6 times) and the process of biofilm formation (2.0–7.7 times). Conclusion. For the first time, the effectiveness of the innovative antibacterial drug Fluorothiazinone has been shown against bacteria of the Pseudomonadota phylum, isolated from chronically infected patients with CF, with the described potential of virulence factors.

DNA-based FeCuAg nanoclusters with peroxidase-like and GSH depletion activities for toxicity of in vitro cancer cells

Developing the efficient nanozymes for reactive oxygen species (ROS)-mediated highly potent tumor catalytic therapy has become a great challenge. In this study, we prepared the DNA-Fe, −FeAg, and −FeCuAg nanocluster (NCs) using the G-/C-rich single-stranded DNA (ssDNA) templates. The steady-state kinetic and the catalytic performances and mechanisms of DNA-metal NCs were first systematically investigated. The results indicated that c-kit-TBA-Fe, c-kit-TBA-FeAg, and c-kit-TBA-FeCuAg NCs exhibited the high peroxidase-like activity. All of three types of NCs presented the higher affinity to the substrate TMB and better storage stability at 4 °C than horseradish peroxidase (HRP). Moreover, c-kit-TBA-FeAg and c-kit-TBA-FeCuAg NCs presented the 6.7- and 4.7-fold stronger affinity to TMB than c-kit-TBA-Fe, respectively. However, the maximum reaction rate (Vmax) of c-kit-TBA-FeCuAg NCs with H2O2 was the largest, which promoted the generation of much more •OH in the reaction system. More importantly, c-kit-TBA-FeCuAg NCs were able to deplete largely the intracellular GSH and thus generate lots of endogenous ROS in HeLa cells, thereby exhibiting the significant and specific in vitro cancer cells toxicity. Therefore, c-kit-TBA-FeCuAg NCs, with peroxidase-like activity and glutathione (GSH) consumption ability, hold the ROS-based promising therapeutic effects for cancer.

Transferrin Immobilized Graphene Oxide Nanocomposite for Targeted Cancer Chemodynamic Therapy via Increasing Intracellular Labile Fe2+ Concentration.

Recently, different alternative regulated cell death (RCD) pathways, viz., necroptosis, pyroptosis, ferroptosis, cuproptosis etc., have been explored as important targets for the development of cancer medications in recent years, as these can change the immunogenicity of the tumor microenvironment (TME) and will finally lead to the inhibition of cancer progression and metastasis. Here, we report the development of transferrin immobilized graphene oxide (Tfn@GOAPTES) nanocomposite as a therapeutic strategy toward cancer cell killing. The electrostatic immobilization of Tfn on the GOAPTES surface was confirmed by different spectroscopy and microscopy techniques. The Tfn immobilization was found to be ∼74 ± 4%, whereas the stability of the protein on the GO surface suggested a robust nature of the nanocomposite. The MTT assay suggested that Tfn@GOAPTES exhibited cytotoxicity toward HeLa cells via increased lipid peroxidation and DNA damage. Western blot studies resulted in decreased expression of acetylation on lysine 40 of α-tubulin and increased expression of LC3a/b for Tfn@GOAPTES treated HeLa cells, suggesting autophagy to be the main cause of the cell death mechanism. Overall, we predict that the present approach can be used as a therapeutic strategy for cancer cell killing via selective induction of a high concentration of intracellular iron.