The cytotoxic activities of the major diterpene extracted from Salvia multicaulis (Bardakosh) are mediated by the regulation of heat-shock response and fatty acid metabolism pathways in human leukemia cells.

BACKGROUND: Leukemia is one of the most lethal cancers worldwide and represents the sixth-leading cause of cancer deaths. The results of leukemia treatment have not been as positive as desired, and recurrence is common. PURPOSE: Thus, there is an urgent requirement for the development of new therapeutic drugs. Salvia multicaulis (Bardakosh) is a widespread species that contains multiple phytochemical components with anti-cancer activities. METHODS: We isolated and characterized the major diterpene candesalvone B methyl ester from S. multicaulis and investigated its action as a cytotoxic agent towards sensitive and drug-resistant leukemia cells by the resazurin reduction assay. Additionally, the targeted genes and the affected molecular mechanisms attributed to the potent cytotoxic activities were discovered by transcriptome-wide mRNA expression profiling. The targets predicted to be regulated by candesalvone B methyl ester in each cell line were confirmed by qRT-PCR, molecular docking, microscale thermophoresis, and western blotting. Moreover, cell cycle distribution and apoptosis were analyzed by flow cytometry. RESULTS: Candesalvone B methyl ester was cytotoxic with IC50 values of 20.95 ± 0.15 µM against CCRF-CEM cells and 4.13 ± 0.10 µM against multidrug-resistant CEM/ADR5000 leukemia cells. The pathway enrichment analysis disclosed that candesalvone B methyl ester could regulate the heat-shock response signaling pathway via targeting heat shock factor 1 (HSF1) in CCRF-CEM cells and ELOVL fatty acid elongase 5 (ELOVL5) controls the fatty acid metabolism pathway in CEM/ADR5000 cells. Microscale thermophoresis showed the binding of candesalvone B methyl ester with HSF1 and ELOVL5, confirming the results of molecular docking analysis. Down-regulation of both HSF1 and ELOVL5 by candesalvone B methyl ester as detected by both western blotting and RT-qPCR was related to the reversal of drug resistance in the leukemia cells. Furthermore, candesalvone B methyl ester increased the arrest in the sub-G1 phase of the cell cycle in a dose-dependent manner from 1.3 % to 32.3 % with concomitant induction of apoptosis up to 29.0 % in CCRF-CEM leukemic cells upon inhibition of HSF1. CONCLUSION: Candesalvone B methyl ester isolated from S. multicaulis exerted cytotoxicity by affecting apoptosis, cell division, and modulation of expression levels of genes contributing to the heat stress signaling and fatty acid metabolism pathways that could relieve drug resistance of leukemia cells.

Foliar application of esculin and digitoxin improve the yield quality of salt-stressed flax by improving the antioxidant defense system.

BACKGROUND: Secondary metabolites of several plants, including esculin and digitoxin, which are cardiac glycosides, were previously employed for their therapeutic effects. The current study aims to investigate the functions of the main Na+ /K+ transport inhibitor digitoxin and the antioxidant esculin for enhancing flax plant growth and production under salinity. METHODOLOGY: Flax plants were irrigated with distilled water supplemented with 0.0 and 5000 mg/L salt solution starting from 15 DAS from sowing. Then exogenous treatment with digitoxin and esculin with 50 mg L- 1 and 100 mg L- 1 were used for this work. RESULTS: According to the results of this work, foliar spraying of esculin or digitoxin increased the salinity tolerance of flax plants.The foliar application of either esculin or digitoxin induced an elevation in the contents of photosynthetic pigments, osmolytes including soluble sugar and proline as well as the total phenols in salt-stressed flax plants. Moreover, esculin and digitoxin in particular counteract oxidative stress by increasing the activity of antioxidant enzymes including superoxide dismutase, catalase, peroxidase, phenylalanine ammonia-lyase, and tyrosine ammonia lyase, leading to a decrease in reactive oxygen species and lipid peroxidation levels and electrolyte leakage. The efficiency of esculin and digitoxin to sustain ion homeostasis by inhibiting Na+ absorption and increasing potassium, calcium, and phosphorus in flax plants may be the reason for their protective actions towards salinity.As a consequence, esculin and digitoxin increased yield quantity and quality as shown by increases in all investigated yield criteriaas shoot height, root length, their fresh and dry weights as well asseed yield/plant (g), and 1000 seeds weight, especially those that improved the desired oil properties. CONCLUSION: In conclusion, this study concluded that digitoxin was more effective in inhibiting Na+ build-up and increasing flax salinity tolerance, particularly at the high investigated dose as compared to esculin. In this study, we reported the recent findings of exogenousapplication of either digitoxin or esculin glycosides which are new investigated salt alleviators never used before for improving the salt tolerance in flax plants.

Screening Wheat Genotypes for Specific Genes Linked to Drought Tolerance.

Drought stress, which significantly affects growth and reduces grain yield, is one of the main problems for wheat crops. Producing promising drought-tolerant wheat cultivars with high yields is one of the main targets for wheat breeders. In this study, a total of seven drought-tolerant wheat genotypes were screened for the presence of 19 specific drought tolerance genes. The genotypes were tested under normal and drought conditions for two growing seasons. Four spike traits, namely, spike length (SPL), grain number per spike (GNPS), number of spikelets per spike (NSPS), and grain yield per spike (GYPS), were scored. The results revealed that drought stress decreased the SPL, GNPS, NSPS, and GYPS, with ranges ranging from 2.14 (NSPS) to 13.92% (GNPS) and from 2.40 (NSPS) to 11.09% (GYPS) in the first and second seasons, respectively. ANOVA revealed high genetic variation among the genotypes for each trait under each treatment. According to the drought tolerance indices, Omara 007 presented the highest level of drought tolerance (average of sum ranks = 3), whereas both Giza-36 genotypes presented the lowest level of drought tolerance (average of sum ranks = 4.8) among the genotypes tested. Among the 19 genes tested, 11 were polymorphic among the selected genotypes. Omara 007 and Omara 002 presented the greatest number of specific drought tolerance genes (nine) tested in this study, whereas Sohag-5, Giza-36, and PI469072 presented the lowest number of drought tolerance genes (four). The number of different genes between each pair of genotypes was calculated. Seven different genes were found between Omara 007 and Giza-36, Omara 007 and Sohag-5, and Omara 002 and PI469072. The results of this study may help to identify the best genotypes for crossing candidate genotypes, and not merely to genetically improve drought tolerance in wheat.

The application of PAHs-Degrading Pseudomonas aeruginosa to mitigate the phytotoxic impact of pyrene on barley (Hordeum vulgare L.) and broad bean (Vicia faba L.) plants.

Mitigating the negative impacts of polycyclic aromatic hydrocarbons (PAHs) is an urgent need due to their toxicity and persistence in the environment. This study investigated the use of Pseudomonas aeruginosa ASU-B6 to detoxify pyrene (PY). The bacterium P. aeruginosa ASU-B6 is capable of degrading PY by 92% as a sole carbon source after 15 days of incubation with phthalate being the major metabolic product. In this regard, the impact of pyrene (PY), P. aeruginosa ASU-B6 (ASU-B6), the bacterial strain combined with pyrene (ASU-B6/PY) and the metabolites produced after pyrene degradation (PY-metabolites) on the germination and physiological attributes of Hordeum vulgare and Vicia faba seedlings were studied. A single application of PY or ASU-B6 showed a toxic effect on the germination of both tested seeds. Interestingly, broad bean seedlings exhibited less sensitivity to PY stress in terms of growth and metabolism compared to barley. Notably, ASU-B6 inhibited fresh and dry weight of shoots and roots of barley and, to a lesser extent, reduced the germination of broad beans compared to the control. However, the combined PY-metabolites and ASU-B6/PY showed a mutual ameliorative effect on seedlings growth, alleviating the phytotoxic impact of each component. Pyrene reduced the virulence of ASU-B6 by inhibiting the production of pyocyanin pigment, while bacteria ameliorated pyrene toxicity through its degradation. Heatmap and principal component analyses highlighted that increasing the contents of hydrogen peroxide, superoxide anion, hydroxyl radical, and lipid peroxidation positively correlated to the toxicity of PY or ASU-B6. However, improving the antioxidant system which buffers the oxidative stress induced by different combinations of PY and ASU-B6 enhanced the growth of germinated seedlings corresponding to PY or ASU-B6. This study reflected the role of ASU-B6 in ameliorating PY-phytotoxicity. In addition, the application of ASU-B6 strain is recommended as a prospective candidate for remediation of PAHs-contaminated environment with a positive impact on the plant growth and metabolic products.

Identification of Cytisine Derivatives as Agonists of the Human Delta Opioid Receptor by Supercomputer-Based Virtual Drug Screening and Transcriptomics.

Delta opioid receptors (DORs) are rising as therapeutic targets, not only for the treatment of pain but also other neurological disorders (e.g., Parkinson's disease). The advantage of DOR agonists compared to µ-opioid receptor agonists is that they have fewer side effects and a lower potential to induce tolerance. However, although multiple candidates have been tested in the past few decades, none have been approved for clinical use. The current study focused on searching for new DOR agonists by screening a chemical library containing 40,000 natural and natural-derived products. The functional activity of the top molecules was evaluated in vitro through the cyclic adenosine monophosphate accumulation assay. Compound 3 showed promising results, and its activity was further investigated through transcriptomic methods. Compound 3 inhibited the expression of TNF-α, prevented NF-κB translocation to the nucleus, and activated the G-protein-mediated ERK1/2 pathway. Additionally, compound 3 is structurally different from known DOR agonists, making it a valuable candidate for further investigation for its anti-inflammatory and analgesic potential.

Investigating the Endophyte Actinomycetota sp. JW0824 Strain as a Potential Bioinoculant to Enhance the Yield, Nutritive Value, and Chemical Composition of Different Cultivars of Anise (Pimpinella anisum L.) Seeds.

Anise (Pimpinella anisum L.) seeds have various nutritional and therapeutic benefits and are thus considered a valuable addition to animal and human health. Hence, in this study, we aimed to induce the nutritive and biological value of anise seeds. To this end, the potential biofortification effect of the endophytic Actinomycetota sp. JW0824 strain, isolated during the fall of 2023 from the medicinal plant Achyranthes aspera, exhibiting natural distribution in the Jazan region of Saudi Arabia, was investigated in four varieties of anise seeds from Egypt, Tunisia, Syria, and Morocco. Results revealed significant increments (p < 0.05) in the seed dry weight percentage (DW%) and oil yields. In line with increased biomass accumulation, the metabolism of the primary and secondary metabolites was increased. There were differential increases in proteins, sugars, flavonoids, alkaloids, phenols, vitamins (e.g., ß-carotene, ascorbic acid), and essential oil components (e.g., phenylpropanoids and monoterpenes), along with their precursor phenylalanine. Consistently, the activity of L-phenylalanine aminolyase (PAL) was increased in the Egyptian and Tunisian varieties at 83.88% and 77.19%, respectively, while 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase (DAHPS) activity increased in all varieties, with a significant 179.31% rise in the Egyptian variety. These findings highlight the beneficial effects of Actinomycetota sp. JW0824 as a bioinoculant for anise seeds, suggesting its potential application in agricultural practices to improve seed yield and quality. Further field trials are recommended to assess the commercial viability of this endophyte for enhancing anise seed production and potentially benefiting other plant species.

Toxicity of bisphenol A and p-nitrophenol on tomato plants: Morpho-physiological, ionomic profile, and antioxidants/defense-related gene expression studies.

Bisphenol A (BPA) and p-nitrophenol (PNP) are emerging contaminants of soils due to their wide presence in agricultural and industrial products. Thus, the present study aimed to integrate morpho-physiological, ionic homeostasis, and defense- and antioxidant-related genes in the response of tomato plants to BPA or PNP stress, an area of research that has been scarcely studied. In this work, increasing the levels of BPA and PNP in the soil intensified their drastic effects on the biomass and photosynthetic pigments of tomato plants. Moreover, BPA and PNP induced osmotic stress on tomato plants by reducing soluble sugars and soluble proteins relative to control. The soil contamination with BPA and PNP treatments caused a decline in the levels of macro- and micro-elements in the foliar tissues of tomatoes while simultaneously increasing the contents of non-essential micronutrients. The Fourier transform infrared analysis of the active components in tomato leaves revealed that BPA influenced the presence of certain functional groups, resulting in the absence of some functional groups, while on PNP treatment, there was a shift observed in certain functional groups compared to the control. At the molecular level, BPA and PNP induced an increase in the gene expression of polyphenol oxidase and peroxidase, with the exception of POD gene expression under BPA stress. The expression of the thaumatin-like protein gene increased at the highest level of PNP and a moderate level of BPA without any significant effect of both pollutants on the expression of the tubulin (TUB) gene. The comprehensive analysis of biochemical responses in tomato plants subjected to BPA and PNP stress illustrates valuable insights into the mechanisms underlying tolerance to these pollutants.

The Histogenetic Origin of Malignant Cells Predicts Their Susceptibility towards Synthetic Lethality Utilizing the TK.007 System.

BACKGROUND: Remarkable differences exist in the outcome of systemic cancer therapies. Lymphomas and leukemias generally respond well to systemic chemotherapies, while solid cancers often fail. We engineered different human cancer cells lines to uniformly express a modified herpes simplex virus thymidine kinase TK.007 as a suicide gene when ganciclovir (GCV) is applied, thus in theory achieving a similar response in all cell lines. METHODS: Fifteen different cell lines were engineered to express the TK.007 gene. XTT-cell proliferation assays were performed and the IC50-values were calculated. Functional kinome profiling, mRNA sequencing, and bottom-up proteomics analysis with Ingenuity pathway analysis were performed. RESULTS: GCV potency varied among cell lines, with lymphoma and leukemia cells showing higher susceptibility than solid cancer cells. Functional kinome profiling implies a contribution of the SRC family kinases and decreased overall kinase activity. mRNA sequencing highlighted alterations in the MAPK pathways and bottom-up proteomics showed differences in apoptotic and epithelial junction signaling proteins. CONCLUSIONS: The histogenetic origin of cells influenced the susceptibility of human malignant cells towards cytotoxic agents with leukemias and lymphomas being more sensitive than solid cancer cells.

Aniquinazoline B, a Fungal Natural Product, Activates the µ-Opioid Receptor.

The development of new µ-opioid receptor (MOR) agonists without the undesirable side effects, such as addiction or respiratory depression, has been a difficult challenge over the years. In the search for new compounds, we screened our chemical database of over 40.000 substances and further assessed the best 100 through molecular docking. We selected the top 10 compounds and evaluated them for their biological activity and potential to influence cyclic adenosine monophosphate (cAMP) levels. From the tested compounds, compound 7, called aniquinazoline B, belonging to the quinazolinone alkaloids class and isolated from the marine fungus Aspergillus nidulans, showed promising results, by inhibiting cAMP levels and in vitro binding to MOR, verified through microscale thermophoresis. Transcriptomic data investigation profiled the genes affected by compound 7 and discovered activation of different pathways compared to opioids. The western blot analysis revealed compound 7 as a balanced ligand, activating both p-ERK1/2 and ß-arrestin1/2 pathways, showing this is a favorable candidate to be further tested.

Jozimine A2, a Dimeric Naphthylisoquinoline (NIQ) Alkaloid, Shows In Vitro Cytotoxic Effects against Leukemia Cells through NF-κB Inhibition.

Naphthylisoquinoline (NIQ) alkaloids are rising as a promising class of secondary metabolites with pharmaceutical potential. NF-κB has already been recognized as a significant modulator of cancer proliferation and drug resistance. We have previously reported the mechanisms behind the cytotoxic effect of dioncophylline A, an NIQ monomer, in leukemia cells. In the current study, we have investigated the cytotoxic effect of jozimine A2, an NIQ dimer, on leukemia cells in comparison to a second, structurally unsymmetric dimer, michellamine B. To this end, molecular docking was applied to predict the binding affinity of the dimers towards NF-κB, which was then validated through microscale thermophoresis. Next, cytotoxicity assays were performed on CCRF-CEM cells and multidrug-resistant CEM/ADR5000 cells following treatment. Transcriptome analysis uncovered the molecular networks affected by jozimine A2 and identified the cell cycle as one of the major affected processes. Cell death modes were evaluated through flow cytometry, while angiogenesis was measured with the endothelial cell tube formation assay on human umbilical vein endothelial cells (HUVECs). The results indicated that jozimine A2 bound to NF-κB, inhibited its activity and prevented its translocation to the nucleus. In addition, jozimine A2 induced cell death through apoptosis and prevented angiogenesis. Our study describes the cytotoxic effect of jozimine A2 on leukemia cells and explains the interactions with the NF-κB signaling pathway and the anticancer activity.

Genome-wide scanning to identify and validate single nucleotide polymorphism markers associated with drought tolerance in spring wheat seedlings.

Unlike other growth stages of wheat, very few studies on drought tolerance have been done at the seedling stage, and this is due to the complexity and sensitivity of this stage to drought stress resulting from climate change. As a result, the drought tolerance of wheat seedlings is poorly understood and very few genes associated with drought tolerance at this stage were identified. To address this challenge, a set of 172 spring wheat genotypes representing 20 different countries was evaluated under drought stress at the seedling stage. Drought stress was applied on all tested genotypes by water withholding for 13 days. Two types of traits, namely morphological and physiological traits were scored on the leaves of all tested genotypes. Genome-wide association study (GWAS) is one of the effective genetic analysis methods that was used to identify target single nucleotide polymorphism (SNP) markers and candidate genes for later use in marker-assisted selection. The tested plant materials were genotyped using 25k Infinium iSelect array (25K) (herein after it will be identified as 25K) (for 172 genotypes) and genotyping-by-sequencing (GBS) (for 103 genotypes), respectively. The results of genotyping revealed 21,093 25K and 11,362 GBS-SNPs, which were used to perform GWAS analysis for all scored traits. The results of GWAS revealed that 131 and 55 significant SNPs were controlling morphological and physiological traits, respectively. Moreover, a total of eight and seven SNP markers were found to be associated with more than one morphological and physiological trait under drought stress, respectively. Remarkably, 10 significant SNPs found in this study were previously reported for their association with drought tolerance in wheat. Out of the 10 validated SNP markers, four SNPs were associated with drought at the seedling stage, while the remaining six SNPs were associated with drought stress at the reproductive stage. Moreover, the results of gene enrichment revealed 18 and six pathways as highly significant biological and molecular pathways, respectively. The selection based on drought-tolerant alleles revealed 15 genotypes with the highest number of different drought-tolerant alleles. These genotypes can be used as candidate parents in future breeding programs to produce highly drought-tolerant genotypes with high genetic diversity. Our findings in this study provide novel markers and useful information on the genetic basis of drought tolerance at early growth stages.

Cytotoxicity of dioncophylline A and related naphthylisoquinolines in leukemia cells, mediated by NF-κB inhibition, angiogenesis suppression, G2/M cell cycle arrest, and autophagy induction.

BACKGROUND: Inhibition of NF-κB activity represents a strategy to treat acute myeloid leukemia, one of the most lethal leukemia types. Naphthylisoquinolines (NIQs) are cytotoxic alkaloids from lianas of the families Ancistrocladaceae and Dioncophyllaceae, which are indigenous to tropical rainforests. PURPOSE: Uncovering therapeutic possibilities and underlying molecular mechanisms of dioncophylline A and its derivatives towards NF-κB related cellular processes. METHODS: Resazurin-based cell viability assay was performed for dioncophylline A and three derivatives on wild-type CCRF-CEM and multidrug-resistant CEM/ADR5000 cells. Transcriptome analysis was executed to discover cellular functions and molecular networks associated with dioncophylline A treatment. Expression changes obtained by mRNA microarray hybridization were confirmed using qRT-PCR. Molecular docking was applied to predict the affinity of the NIQs with NF-κB. To validate the in silico approach, NF-κB reporter assays were conducted on HEK-Blue™ Null1 cells. Cell death mechanisms and cell cycle arrest were studied using flow cytometry. The potential activity on angiogenesis was evaluated with the endothelial cell tube formation assay on HUVECs using fluorescence microscopy. Intracellular NF-κB location in HEK-Blue™ Null1 cells was visualized with immunofluorescence. Finally, the anti-tumor activity of dioncophylline A was studied by a xenograft zebrafish model in vivo. RESULTS: Our study demonstrated that dioncophylline A and its derivatives exerted potent cytotoxicity on leukemia cells. Using Ingenuity Pathway Analysis, we identified the NF-κB network as the top network, and docking experiments predicted dioncophylline A and two of its derivatives sharing the same binding pocket with the positive control compound, triptolide. Dioncophylline A showed the best inhibitory activity in NF-κB reporter assays compared to its derivatives, caused autophagy rather than apoptosis, and induced G2/M arrest. It also prevented NF-κB translocation from the cytoplasm to the nucleus. Tube formation as an angiogenesis marker was significantly suppressed by dioncophylline A treatment. Finally, the remarkable anti-tumor activity of dioncophylline A was proven in zebrafish in vivo. CONCLUSION: Taken together, we report for the first time the molecular mechanism behind the cytotoxic effect of dioncophylline A on leukemia cells. Dioncophylline A showed strong cytotoxic activity, inhibited NF-κB translocation, significantly affected the NF-κB in silico and in vitro, subdued tube formation, induced autophagy, and exerted antitumor activity in vivo. Our findings enlighten both the cellular functions including the NF-κB signaling pathway and the cytotoxic mechanism affected by dioncophylline A.

Elevated CO2 differentially attenuates beryllium-induced oxidative stress in oat and alfalfa.

Elevated CO2 (eCO2 ) is one of the climate changes that may benefit plant growth under emerging soil contaminants such as heavy metals. In this regard, the morpho-physiological mechanisms underlying the mitigating impact of eCO2 on beryllium (Be) phytotoxicity are poorly known. Hence, we investigated eCO2 and Be interactive effects on the growth and metabolism of two species from different groups: cereal (oat) and legume (alfalfa). Be stress significantly reduced the growth and photosynthetic attributes in both species, but alfalfa was more susceptible to Be toxicity. Be stress induced reactive oxygen species (ROS) accumulation by increasing photorespiration, subsequently resulting in increased lipid and protein oxidation. However, the growth inhibition and oxidative stress induced by Be stress were mitigated by eCO2 . This could be explained, at least partially, by the increase in organic acids (e.g., citric acid) released into the soil, which subsequently reduced Be uptake. Additionally, eCO2 reduced cellular oxidative damage by reducing photorespiration, which was more significant in alfalfa plants. Furthermore, eCO2 improved the redox status and detoxification processes, including phytochelatins, total glutathione and metallothioneins levels, and glutathione-S-transferase activity in both species, but to a greater extend in alfalfa. In this context, eCO2 also stimulated anthocyanin biosynthesis by accumulating its precursors (phenylalanine, coumaric acid, cinnamic acid, and naringenin) and key biosynthetic enzymes (phenylalanine ammonia-lyase, cinnamate hydroxylase, and coumarate:CoA ligase) mainly in alfalfa plants. Overall, this study explored the mechanistic approach by which eCO2 alleviates the harmful effects of Be. Alfalfa was more sensitive to Be stress than oats; however, the alleviating impact of eCO2 on Be stress was more pronounced in alfalfa.

Molecular determinants of the response of cancer cells towards geldanamycin and its derivatives.

Geldanamycin is an ansamycin-derivative of a benzoquinone isolated from Streptomyces hygroscopicus. It inhibits tyrosine kinases and heat shock protein 90 (HSP90). Geldanamycin and 11 derivatives were subjected to molecular docking to HSP90, and 17-desmethoxy-17-N,N-dimethylamino-geldanamycin (17-DMAG) was the compound with the highest binding affinity (-7.73 ± 0.12 kcal/mol) and the lowest inhibition constant (2.16 ± 0.49 µM). Therefore, 17-DMAG was selected for further experiments in comparison to geldanamycin. Multidrug resistance (MDR) represents a major problem for successful cancer therapy. We tested geldanamycin and 17-DMAG against various drug-resistant cancer cell lines. Although geldanamycin and 17-DMAG inhibited the proliferation in all cell lines tested, multidrug-resistant P-glycoprotein-overexpressing CEM/ADR5000 cells were cross-resistant, ΔEGFR-overexpressing tumor cells and p53 knockout cells were sensitive to these two compounds. COMPARE and hierarchical cluster analyses were performed, and 60 genes were identified to predict the sensitivity or resistance of 59 NCI tumor cell lines towards geldanamycin and 17-DMAG. The distribution of cell lines according to their mRNA expression profiles indicated sensitivity or resistance to both compounds with statistical significance. Moreover, bioinformatic tools were used to study possible mechanisms of action of geldanamycin and 17-DMAG. Galaxy Cistrome analyses were carried out to predict transcription factor binding motifs in the promoter regions of the candidate genes. Interestingly, the NF-ĸB DNA binding motif (Rel) was identified as the top transcription factor. Furthermore, these 60 genes were subjected to Ingenuity Pathway Analysis (IPA) to study the signaling pathway interactions of these genes. Interestingly, IPA also revealed the NF-ĸB pathway as the top network among these genes. Finally, NF-ĸB reporter assays confirmed the bioinformatic prediction, and both geldanamycin and 17-DMAG significantly inhibited NF-κB activity after exposure for 24 h. In conclusion, geldanamycin and 17-DMAG exhibited cytotoxic activity against different tumor cell lines. Their activity was not restricted to HSP90 but indicated an involvement of the NF-KB pathway.

The Novel Artemisinin Dimer Isoniazide ELI-XXIII-98-2 Induces c-MYC Inhibition, DNA Damage, and Autophagy in Leukemia Cells.

The proto-oncogenic transcription factor c-MYC plays a pivotal role in the development of tumorigenesis, cellular proliferation, and the control of cell death. Its expression is frequently altered in many cancer types, including hematological malignancies such as leukemia. The dimer isoniazide ELI-XXIII-98-2 is a derivative of the natural product artemisinin, with two artemisinin molecules and an isoniazide moiety as a linker in between them. In this study, we aimed to study the anticancer activity and the molecular mechanisms of this dimer molecule in drug-sensitive CCRF-CEM leukemia cells and their corresponding multidrug-resistant CEM/ADR5000 sub-line. The growth inhibitory activity was studied using the resazurin assay. To reveal the molecular mechanisms underlying the growth inhibitory activity, we performed in silico molecular docking, followed by several in vitro approaches such as the MYC reporter assay, microscale thermophoresis, microarray analyses, immunoblotting, qPCR, and comet assay. The artemisinin dimer isoniazide showed a potent growth inhibitory activity in CCRF-CEM but a 12-fold cross-resistance in multidrug-resistant CEM/ADR5000 cells. The molecular docking of artemisinin dimer isoniazide with c-MYC revealed a good binding (lowest binding energy of -9.84 ± 0.3 kcal/mol) and a predicted inhibition constant (pKi) of 66.46 ± 29.5 nM, which was confirmed by microscale thermophoresis and MYC reporter cell assays. Furthermore, c-MYC expression was downregulated by this compound in microarray hybridization and Western blotting analyses. Finally, the artemisinin dimer isoniazide modulated the expression of autophagy markers (LC3B and p62) and the DNA damage marker pH2AX, indicating the stimulation of both autophagy and DNA damage, respectively. Additionally, DNA double-strand breaks were observed in the alkaline comet assay. DNA damage, apoptosis, and autophagy induction could be attributed to the inhibition of c-MYC by ELI-XXIII-98-2.

Comparative Study of Three Biological Control Agents and Two Conventional Fungicides against Coriander Damping-off and Root Rot Caused by Rhizoctonia solani.

The in vitro and in vivo efficacy of three biocontrol agents, Trichoderma viride, Pseudomonas fluorescence, and Bacillus subtilis, were tested against Rhizoctonia solani (AG-4) infection compared to two conventional fungicides (Rizolex-T 50%wettable powder and Amistar 25%). Antifungal enzyme activity was assayed in the culture filtrate of the biocontrol agents. The impact of the tested biocontrol agents on the induction of the coriander immune system was investigated against R. solani by assessing the resistance-related enzymes and compounds in biocontrol agent-treated plants compared with the control. The obtained results revealed that all tested biocontrol agents significantly reduced the linear growth of R. solani, and T. viride recorded the highest inhibition percentage. This could be linked to the ability of T. viride to produce higher activities of antimicrobial enzymes, i.e., cellulase, chitinase, and protease, compared to P. fluorescence and B. subtilis. Applying the tested biocontrol agents significantly alleviated pre- and post-emergence damping-off and root rot/wilt diseases of infected coriander compared with untreated plants. The tested biocontrol agents exhibited significantly higher germination percentage and vigor index of the coriander than the tested fungicides. The tested biocontrol agents significantly minimized the reduction of photosynthetic pigments induced by R. solani. In addition, the results showed a significant increase in enzymes/molecules (i.e., phenylalanine, catalase, peroxidase, catalase, superoxide dismutase, phenylalanine ammonia-lyase, phenolics, ascorbic acids, and salicylic acid) involved directly and indirectly in coriander resistance to R. solani. The principal component analysis of the recorded data recommended the role of the high accumulation of oxidative parameters (hydrogen peroxide and lipid peroxidation) and the inhibition of phenolic compounds in the downregulation of coriander resistance against R. solani. The heatmap analysis results revealed that biocontrol agents, especially Trichoderma, enhanced the resistance against R. solani via the stimulation of salicylic acid, phenolics, and antioxidant enzymes. Overall, the data recommended the efficacy of biocontrol agents, especially T. viride, against R. solani infecting coriander plants, which could be an efficient and a safer alternative to conventional fungicides.

Effects of microbial biostimulants (Trichoderma album and Bacillus megaterium) on growth, quality attributes, and yield of onion under field conditions.

Microbial biostimulants (MBs) promote plant growth and stress tolerance in a sustainable manner. However, precise field trials of MBs are required in natural setting with a range of crop varieties to harness the benefits of biostimulants on crop yield improvement. This study investigated the effects of two MBs, Trichoderma album and Bacillus megaterium, on an onion cultivar's growth, nutritional qualities, antioxidant properties, and yield potentials under field conditions for two successive years. Before transplantation, onion bulbs were gelatin-coated with 2.0 and 4.0 g L-1 of each of the MB. Results revealed that MBs-pretreated onion plants exhibited better growth indices, photosynthetic pigment contents, and yield-attributing features like bulb weight than control plants. Nutraceutical analysis demonstrated that T. album-pretreated (by 2.0 g L-1) onion cultivar enhanced the level of K+ (by 105.79%), Ca2+ (by 37.77%), proline (by 34.21%), and total free amino acids (by 144.58%) in bulb tissues over the control plants. Intriguingly, the pretreatment with both T. album and B. megaterium (by 2.0 g L-1) increased the levels of total soluble carbohydrates (by 19.10 and 84.02%), as well as antioxidant properties, including increased activities of superoxide dismutase (by 58.52 and 31.34%), catalase (by 164.71 and 232%), ascorbate peroxidase (by 175.35 and 212.69%), and glutathione-S-transferase (by 31.99 and 9.34%) and improved the contents of ascorbic acid (by 19.1 and 44.05%), glutathione (by 6.22 and 33.82%), and total flavonoids (by 171.98 and 56.24%, respectively) in the bulb tissues than control plants. Although both MBs promoted the growth and nutraceutical qualities of onion bulbs, T. album pretreatment showed better effects than that of B. megaterium in the field settings. Based on the morphophysiological attributes and biochemical properties, a low dose (2.0 g L-1) was more effective than a high dose (4.0 g L-1) of T. album in promoting onion growth. Overall, the current research findings imply that T. album might be a potential MB in improving growth and quality attributes, and hence the productivity of onion cultivars under field circumstances.

A drug repurposing approach for individualized cancer therapy based on transcriptome sequencing and virtual drug screening.

RNA-sequencing has been proposed as a valuable technique to develop individualized therapy concepts for cancer patients based on their tumor-specific mutational profiles. Here, we aimed to identify drugs and inhibitors in an individualized therapy-based drug repurposing approach focusing on missense mutations for 35 biopsies of cancer patients. The missense mutations belonged to 9 categories (ABC transporter, apoptosis, angiogenesis, cell cycle, DNA damage, kinase, protease, transcription factor, tumor suppressor). The highest percentages of missense mutations were observed in transcription factor genes. The mutational profiles of all 35 tumors were subjected to hierarchical heatmap clustering. All 7 leukemia biopsies clustered together and were separated from solid tumors. Based on these individual mutation profiles, two strategies for the identification of possible drug candidates were applied: Firstly, virtual screening of FDA-approved drugs based on the protein structures carrying particular missense mutations. Secondly, we mined the Drug Gene Interaction (DGI) database (https://www.dgidb.org/) to identify approved or experimental inhibitors for missense mutated proteins in our dataset of 35 tumors. In conclusion, our approach based on virtual drug screening of FDA-approved drugs and DGI-based inhibitor selection may provide new, individual treatment options for patients with otherwise refractory tumors that do not respond anymore to standard chemotherapy.

Molecular Modes of Action of an Aqueous Nerium oleander Extract in Cancer Cells In Vitro and In Vivo.

Cancer drug resistance remains a major obstacle in clinical oncology. As most anticancer drugs are of natural origin, we investigated the anticancer potential of a standardized cold-water leaf extract from Nerium oleander L., termed Breastin. The phytochemical characterization by nuclear magnetic resonance spectroscopy (NMR) and low- and high-resolution mass spectrometry revealed several monoglycosidic cardenolides as major constituents (adynerin, neritaloside, odoroside A, odoroside H, oleandrin, and vanderoside). Breastin inhibited the growth of 14 cell lines from hematopoietic tumors and 5 of 6 carcinomas. Remarkably, the cellular responsiveness of odoroside H and neritaloside was not correlated with all other classical drug resistance mechanisms, i.e., ATP-binding cassette transporters (ABCB1, ABCB5, ABCC1, ABCG2), oncogenes (EGFR, RAS), tumor suppressors (TP53, WT1), and others (GSTP1, HSP90, proliferation rate), in 59 tumor cell lines of the National Cancer Institute (NCI, USA), indicating that Breastin may indeed bypass drug resistance. COMPARE analyses with 153 anticancer agents in 74 tumor cell lines of the Oncotest panel revealed frequent correlations of Breastin with mitosis-inhibiting drugs. Using tubulin-GFP-transfected U2OS cells and confocal microscopy, it was found that the microtubule-disturbing effect of Breastin was comparable to that of the tubulin-depolymerizing drug paclitaxel. This result was verified by a tubulin polymerization assay in vitro and molecular docking in silico. Proteome profiling of 3171 proteins in the NCI panel revealed protein subsets whose expression significantly correlated with cellular responsiveness to odoroside H and neritaloside, indicating that protein expression profiles can be identified to predict the sensitivity or resistance of tumor cells to Breastin constituents. Breastin moderately inhibited breast cancer xenograft tumors in vivo. Remarkably, in contrast to what was observed with paclitaxel monotherapy, the combination of paclitaxel and Breastin prevented tumor relapse, indicating Breastin's potential for drug combination regimens.

Role of Acetic Acid and Nitric Oxide against Salinity and Lithium Stress in Canola (Brassica napus L.).

In this study, canola (Brassica napus L.) seedlings were treated with individual and combined salinity and lithium (Li) stress, with and without acetic acid (AA) or nitric acid (NO), to investigate their possible roles against these stresses. Salinity intensified Li-induced damage, and the principal component analysis revealed that this was primarily driven by increased oxidative stress, deregulation of sodium and potassium accumulation, and an imbalance in tissue water content. However, pretreatment with AA and NO prompted growth, re-established sodium and potassium homeostasis, and enhanced the defense system against oxidative and nitrosative damage by triggering the antioxidant capacity. Combined stress negatively impacted phenylalanine ammonia lyase activity, affecting flavonoids, carotenoids, and anthocyanin levels, which were then restored in canola plants primed with AA and NO. Additionally, AA and NO helped to maintain osmotic balance by increasing trehalose and proline levels and upregulating signaling molecules such as hydrogen sulfide, γ-aminobutyric acid, and salicylic acid. Both AA and NO improved Li detoxification by increasing phytochelatins and metallothioneins, and reducing glutathione contents. Comparatively, AA exerted more effective protection against the detrimental effects of combined stress than NO. Our findings offer novel perspectives on the impacts of combining salt and Li stress.