Half Maximal Inhibitory Concentration Research Articles

Rigidifying aggregation-induced emission luminogens by metal–organic framework formation for sensitive lateral flow immunoassay

T-2 toxin, the strongest toxic of type A trichothecenes, widely exists in barley, wheat, corn, oats, and even human food, which is urgent to establish a rapid and sensitive detection method for T-2 toxin. Herein, we developed a fluorescent lateral flow immunoassay (LFIA) for sensitive detection of T-2 toxin. Specially, the monoclonal antibody (mAb) was prepared by immunizing mice and cell fusion, and exhibited a low half maximal inhibitory concentration (IC50) of 0.482 ng mL−1. In addition, an aggregation-induced emission (AIE)-based metal-organic framework (HfMOFs) with excellent fluorescence performance (QY: 72.5 %) was developed as reporter. Benefiting from these advantages, the limit of detection (LoD) of HfMOFs-LFIA was 0.0487 ng mL−1, which was 7.7-fold lower than traditional nanoparticles-based LFIA (0.375 ng mL−1). In addition, HfMOFs-LFIA was used for the detection of T-2 toxin in actual samples and showed satisfactory recoveries (71.3 %–123.7 %) with coefficient of variation (2.96 %–14.07 %). Overall, the dual-improvement in mAb and label might power the development in LFIA, achieving innovative prospects for LFIA in food safety fields.

Investigation of Apoptosis-mediated Cytotoxic Effects of Royal Jelly on HL-60 Cells

In recent years, the use of nontoxic natural products that can be effective on cancer cells as new agents has attracted the attention of scientists in order to reduce the negative side effects of existing cancer drugs and their toxicity to normal cells. Some in vivo and in vitro studies have shown that royal jelly (RJ) inhibits cell proliferation and induces apoptosis. In this research, we aimed to investigate the effects of RJ on proliferative and apoptotic processes in the human acute promyelocytic leukemia cell line (HL-60). The HL60 cell line was treated with different concentrations of RJ for 24, 48, and 72 hours. The half maximum inhibitory concentration (IC50) of RJ was determined using 3-(4.5 dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide test (MTT) and the proliferation activity of HL-60 cells was evaluated. Flow cytometry analysis was performed to measure apoptosis in HL-60 cells. IC50 values for RJ were calculated as 13.98, 6.45, and 2.06 mg/mL for 24, 48, and 72 hours, respectively. Flow cytometry results also showed that RJ had apoptotic effects at the concentrations found. The results showed that RJ treatment significantly induced apoptosis and reduced the proliferation of HL-60 cells. This study shows that RJ can be a complementary treatment against HL-60 acute myeloid leukemia cells due to its anticancer and antiproliferative effects.

Formulation and characterization of 5-fluorouracil and metformin biodegradable nanospheres for treating colon cancer

BackgroundCancer and diabetes mellitus are quite common diseases found together worldwide. A considerable amount of evidence is available for the rapid development and presentation of various types of cancer in the type 2 diabetes mellitus (DM2) population as compared with the population without diabetes. The objective of the study is to formulate and evaluate 5-fluorouracil (5-FU) and metformin (MET) nanoparticles (NPs) and to establish the characteristic features of the biodegradable NPs. 5-FU and MET NPs with chitosan as a biodegradable polymer were formulated by the ionotropic cross-linking method. 0.25 gm of MET and 0.25 gm of 5-FU were dissolved in 100 ml of distilled water, and stock solution was prepared. 5 ml of stock solution was slowly mixed into the chitosan solution to obtain the mixture of drugs and chitosan. The tripolyphosphate reserve liquid was dripped slowly into the chitosan solution with constant stirring until the opaline appearance was noticed in the mixture, then filtered, and dried. The NPs were evaluated for their physical properties and drug release kinetics. Cell proliferation assay was done using human colorectal carcinoma cell line (HCT 116).ResultsThe formulation composed of 1.5 w/v of chitosan, 0.25 w/v of MET, and 5-FU had an average particle diameter of 198.7 nm. The polydispersity index was 0.757. Thermal and infrared spectroscopy results indicated the drugs and polymers selected for the formulation were unique without any identifiable interaction. The NPs were spherical in appearance, with numerous pours on the surface, which was evident when microscopically examined. Uniformity in drug release was observed, and the formulation demonstrated excellent release kinetics. HTC 116 cell line confirmed that the maximum percentage of cell death and minimum viability of cells were observed while using the combination of MET and 5-FU-NPs as compared to the pure MET or 5-FU alone.ConclusionMET and 5-FU-loaded chitosan NPs were found to have excellent physiochemical properties, particle size, and drug release from the polymer in a controlled manner. Half-maximal inhibitory concentration value (IC50) of MET and 5-FU-NPs was found to be significantly less as compared to MET and 5-FU alone or the combination.

Streptomyces sp. from desert soil as a biofactory for antioxidants with radical scavenging and iron chelating potential

Iron homeostasis is vital for normal physiology, but in the majority of circumstances, like iron overload, this equilibrium is upset leading to free iron in the plasma. This condition with excess iron is known as hemochromatosis, which has been linked to many side effects, including cancer and liver cirrhosis. The current research aimed to investigate active molecules from Streptomyces sp. isolated from the extreme environment of Bahawalpur deserts. The strain was characterized using 16 S rRNA sequencing. Chemical analysis of the ethyl acetate cure extract revealed the presence of phenols, flavonoids, alkaloids, and tannins. Multiple ultraviolet (UV) active metabolites that were essential for the stated pharmacological activities were also demonstrated by thin layer chromatography (TLC) and high-performance liquid chromatography (HPLC). Additionally, Gas chromatography/mass spectrometry (GC-MS) analysis revealed the primary constituents of the extract to compose of phenol and ester compounds. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay was used to assess the extract’s antioxidant capacity, and the results showed a good half-maximal inhibitory concentration (IC50) value of 0.034 µg/mL in comparison to the positive control ascorbic acid’s 0.12 µg/mL. In addition, iron chelation activity of extract showed significant chelation potential at 250 and 125 µg/mL, while 62.5 µg/mL showed only mild chelation of the ferrous ion using ethylene diamine tetra acetic acid (EDTA) as a positive control. Likewise, the extract’s cytotoxicity was analyzed through 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay using varying concentrations of the extract and showed 51% cytotoxicity at 350 µg/mL and 65% inhibition of cell growth at 700 µg/mL, respectively. The bioactive compounds from Streptomyces sp. demonstrated strong antioxidant and iron chelating potentials and can prolong the cell survival in extreme environment.

Different Doxorubicin Sensitivity Across Various Human Cancer Cell Lines

Doxorubicin (Dox) is a highly potent chemotherapeutic agent, approved by FDA since 1974, for treating a broad spectrum of cancers. However, development of severe side effects and drug resistance are main issues that limit the clinical use of Dox. Herein, we aimed to investigate the sensitivity of Dox in a variety of human cancer cell lines. Eleven cell lines were tested in this study including 2 hepatocellular carcinoma (HCC) cell lines (HepG2 and Huh7), 4 bladder cancer (BlCa) cell lines (UMUC-3, VMCUB-1, TCCSUP and BFTC-905), a lung cancer cell line (A549), a cervical carcinoma cell line (HeLa), a breast cancer cell line (MCF-7), a skin melanoma cell line (M21) and a noncancer human kidney cell line (HK-2). The MTT assay was employed for the determination of cytotoxicity. Cells were treated with various concentrations of Dox for 24 h. The half-maximal inhibitory concentration (IC50) of Dox was calculated to compare the Dox sensitivity among tested cell lines. The result showed that IC50 of Dox in HepG2, Huh7, UMUC-3, VMCUB-1, TCCSUP, BFTC-905, A549, HeLa, MCF-7, M21 and HK-2 cells were 12.2, > 20, 5.1, > 20, 12.6, 2.3, > 20, 2.9, 2.5, 2.8 and > 20 mM, respectively. BFTC-905 had the lowest IC50 value, therefore, it was the most sensitive to Dox. In contrast, Huh7, VMCUB-1 and A549 cells were resistant to Dox with IC50 values > 20 mM. Conclusions, we demonstrated that different human malignant cell lines had different sensitivity to Dox. BFTC-905 BlCa cell line had the highest sensitivity to Dox. Huh7, VMCUB-1 and A549 cell lines were resistant to Dox. Perhaps, the different Dox sensitivity in different cell lines was due to the different acquisition of Dox resistance mechanisms. Huh7, VMCUB-1 and A549 cell lines could be suitable cell models to investigate the molecular mechanism of Dox resistance in different cancers. HIGHLIGHTS Different types of cancer cell lines exhibited different sensitivity to doxorubicin. BFTC-905, MCF-7 and M21 cell lines were sensitive to doxorubicin. HepG2, UMUC-3, TCCSUP and HeLa cells were moderately sensitive to doxorubicin. Huh7, VMCUB-1, A549 and HK-2 cells were resistant to doxorubicin. Huh7, VMCUB-1 and A549 cell lines could be suitable cell models for investigating the molecular pathways of Dox resistance in different types of cancers. GRAPHICAL ABSTRACT

Pharmacological blockade of the mast cell MRGPRX2 receptor supports investigation of its relevance in skin disorders.

Because MRGPRX2 is now recognized as the mast cell receptor for basic secretagogues, there is currently a tremendous interest in whether MRGRPX2 could play an important role in various pruritic dermatoses such as chronic spontaneous urticaria. Therefore, we sought to identify new potent and selective antagonists to pharmacologically characterize the biological role of MRGPRX2. Various relevant in vitro, ex vivo, and in vivo model systems were used to investigate the role of MRGPRX2. This included the study of freshly isolated human skin mast cells and human basophils as well as an ex vivo human skin microdialysis preparation. The additivity of MRGPRX2 and FcεR1-mediated degranulation was also investigated. Human MRGPRX2 knock-in mice were generated to interrogate pharmacokinetic/pharmacodynamic relationships because both antagonists studied were shown to be human specific. Two novel and structurally distinct MRGPRX2 antagonists were identified with one, Compound B, being orally active and demonstrating high potency in blocking Substance P-mediated degranulation using freshly isolated human skin mast cells with half maximal inhibitory concentration (IC50) at 0.42 nM. Compound B also potently blocked Substance P-stimulated histamine release from resident mast cells in a human skin explant setup as well as blocking itch in an established behavioral scratching model using MRGPRX2 knock-in mice. Unlike human mast cells, Substance P failed to elicit a functional response in human basophils. These data fully support the investigation of MRGPRX2 receptor antagonists in mast cell-driven allergic skin disorders such as chronic spontaneous urticaria.

In vitro effects of l-kynurenine and quinolinic acid on adhesion, migration and apoptosis in B16 F10 melanoma cells

IntroductionThe inhibition of melanoma adhesion through adhesion molecules, such as integrins and E-cadherin, may represent a promising strategy for managing melanoma metastasis. Compounds, namely l-kynurenine (L-kyn) and quinolinic acid (Quin), previously displayed anti-cancer effects at half-maximal inhibitory concentration (IC50) against B16 F10 melanoma cells in vitro. However, the role of these compounds in B16 F10 melanoma cell adhesion, migration and apoptosis remain unknown. MethodsPost-exposure to the compounds, flow cytometry was used to analyse the expression of very late antigen-5 (VLA-5), E-cadherin and cleaved caspase-3 in B16 F10 melanoma and RAW 264.7 murine macrophage cells. An adhesion assay was used to quantify the adhesion of both cell lines to vitronectin. A scratch migration assay was used to measure the possible inhibition of cell migration in B16 F10 cells in response to L-kyn and Quin. ResultsIn both B16 F10 and RAW 264.7 cells, neither L-kyn nor Quin induced significant effects on VLA-5 expression or cell adhesion to vitronectin. In B16 F10 cells, both L-kyn and Quin elevated E-cadherin expression and displayed a trend of suppressed migration. However, only L-kyn elevated E-cadherin in RAW 264.7 cells. L-kyn induced apoptosis by elevating cleaved caspase-3 expression in both cell lines. ConclusionL-kyn and Quin demonstrated promising antimetastatic effects in their ability to elevate E-cadherin expression and induce apoptosis in B16 F10 melanoma cells. However, these effects did not occur in response to vitronectin or VLA-5 integrin alterations. Furthermore, it cannot be excluded that L-kyn also induced apoptosis in RAW 264.7 cells. As such, these effects should be confirmed in additional control cell lines and substantiated with in vivo models.

Chemical characterization, antimicrobial and mosquito larvicidal activities of the essential oil of Camellia pleurocarpa (Gagnep.) Sealy from Vietnam

Camellia pleurocarpa (Gagnep.) Sealy, a species of yellow camellia indigenous to Vietnam, was investigated for its essential oil properties in this study. Hydrodistillation of its leaves yielded a complex mixture rich in sesquiterpenes and diterpenes. Chemical analysis identified 50 constituents, with notable compounds including spathulenol (13.26%), phytol (9.94%), and α-selinene (5.34%). The essential oil exhibited significant antimicrobial activity against three Gram-positive bacteria Enterococcus faecalis, Staphylococcus aureus, and Bacillus cereus; one Gram-negative bacterium Pseudomonas aeruginosa; and one yeast Candida albicans, with minimum inhibitory concentration (MIC) values ranging from 16 to 32 μg/mL and half-maximal inhibitory concentration (IC50) values ranging from 7.26 to 16.23 μg/mL. Furthermore, it demonstrated larvicidal potential against Aedes aegypti and Culex quinquefasciatus mosquitoes, with LC50 values ranging from 13.49 to 28.95 μg/mL and LC90 values ranging from 28.29 to 53.24 μg/mL. These findings underscore the promising antimicrobial and mosquito larvicidal properties of C. pleurocarpa essential oil, suggesting its potential as a natural alternative in combating microbial infections and controlling mosquito-borne diseases. This study represents the first comprehensive report on the chemical compositions and bioactivities of C. pleurocarpa essential oil.

Characterization of chemical properties of fish-flavoured spicy tamarind cube

Fish-flavoured spicy tamarind cube is categorized as a type of bouillon cube. It is a processed food that has been developed and benefits the consumers due to low cost, being ready to use, reduced cooking time and giving intense flavour to the cooking dish. Food processing improves the stability, safety, taste and makes the food healthier. While there are various advantages to processing, the chemical food characteristics might be adversely affected. The study aimed to evaluate the amount of chemical composition, macromolecule, phytochemical content, peroxide value and antioxidant activity present in the fish-flavoured spicy tamarind cube. The analysis was performed using a different methodology. The content of mineral and ascorbic acid of the cube was evaluated by using scanning electron microscopy-energy dispersive X-ray (SEM-EDX) and titration methods respectively. Carbohydrate was estimated by difference using equation and protein by micro Kjeldahl method. For both phytochemical compounds, total phenolic and flavonoid are measured spectrophotometrically in this study. In addition, peroxide value was determined by titration method and antioxidant activity was assayed using 2,2- diphenyl-1-picrylhydrazyl (DPPH) free radical. The result showed the cube contained 2.12% sodium, 1.22% copper and 0.43% potassium while the ascorbic acid value was 66.00 mg/100 g. The macromolecule content recorded was 40.15 g/100 g for carbohydrates and 7.54 g/100 g for protein. The cube had 10.77 mg GAE/g and 21.72 mg QE/g for total phenolic and flavonoid respectively. The peroxide value of the cube was 9.52 meq/kg and classified in a moderate oxidation state. Furthermore, the half-maximal inhibitory concentration (IC50) for antioxidant activity analysed was 89.04 µg/mL which means the cube shows intermediate antioxidant activity. Based on the result, the cube is safe to consume and can provide health benefits to consumers.

Albendazole nanosuspension coated granules for the rapid localized release and treatment of colorectal cancer

Albendazole (ABZ), an anthelmintic drug, has been repurposed to treat various types of cancers. However, poor solubility of ABZ, resulting in low bioavailability, limits its application. Nanosuspension is a versatile method for enhancing the dissolution of hydrophobic molecules, but a successful drying has been the biggest challenge in the field. The objective of this research is to formulate and optimize ABZ nanosuspension (NS) coated granules for rapid delivery of ABZ for the treatment of colorectal cancer. ABZ NS was prepared by dual centrifugation method using Kollidon® VA64 and sodium lauryl sulphate (SLS) as stabilizers. The processing method was optimized to obtain a stable nanosuspension with particle size < 300 nm. The optimized ABZ NS was coated on microcrystalline cellulose (MCC) to form the nano-coated granules (NCG) and filled in EUDRACAP® for colon targeted delivery. The ABZ NS and NCG achieved ∼ 60 % and ∼55 % drug release, respectively, in presence of bile salt at colonic pH. Half-maximal inhibitory concentration (IC50) of ABZ NS was found to be 1.18 ± 0.081 µM and 3.59 ± 0.080 µM in two colorectal cancer cell lines: HCT 116 and HT-29, respectively. In addition, In vitro 3D tumor assay revealed that ABZ NS has superior tumor growth inhibition activity compared to the control and pure ABZ. The preparation of ABZ NCG in EUDRACAP® could be a promising approach to achieve colon targeted delivery and to repurpose ABZ for the treatment of colorectal cancer.

Antiviral activity of Polygonum aviculare extract against murine norovirus as norovirus surrogate and its application in model food

Norovirus is a major cause of acute gastroenteritis, with vaccines or antiviral therapy yet to be approved. Polygonum aviculare is a medicinal plant with various physiological properties. In this study, the antiviral activity of Polygonum aviculare extract (PAE) was examined against murine norovirus (MNV-1), a human norovirus surrogate. Pre-treatment with PAE suppressed MNV-1, achieving a half-maximal inhibitory concentration (IC50) of 78.4 μg/mL. TEM images revealed viral morphological changes following PAE treatment, indicating the antiviral effect of PAE. Treatment with 100 and 200 μg/mL PAE for 1 h significantly reduced the temperature-dependent antiviral PAE activity at all temperatures. Time-of-addition test results also demonstrated PAE antiviral activity in cell pre-, co-, and post-treatments, with slightly higher activity detected upon cell pre-treatment. HPLC analysis identified avicularin and myricitrin peaks, and LC-QTOF/MS determined phenolic acids and flavonoids as the main constituents. Our findings confirmed the antiviral efficacy of myricetin and quercetin at concentrations of 50 and 100 μM in co-treatment experiments. Upon inoculating the cabbage surface with MNV-1, PAE treatment at 4 or 25 °C reduced MNV-1 plaque formation. Collectively, our findings suggest that PAE could effectively inactivate norovirus and demonstrate promising potential as a natural antiviral agent for food safety.

Traditional Medicinal Ranunculaceae Species from Romania and Their In Vitro Antioxidant, Antiproliferative, and Antiparasitic Potential

Several Ranunculaceae species are used in folk medicine to eliminate pathologies associated with oxidative stress as well as parasitic infections; however, a number of studies confirming their pharmacological properties is limited. In this study, 19 ethanolic extracts obtained from 16 Ranunculaceae species were assayed for in vitro antioxidant, antiproliferative, and antiparasitic potential. The maximum antioxidant potential in both oxygen radical absorbance capacity (ORAC) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) assays was observed for Aconitum toxicum extract [half-maximal inhibitory concentration (IC50) 18.7 and 92.6 μg/mL]. Likewise, Anemone transsilvanica extract exerted the most promising antiproliferative activity against Caco-2 (IC50 46.9 μg/mL) and HT29 (IC50 70.2 μg/mL) cell lines in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Additionally, a dual antioxidant and cytotoxicity effect was demonstrated for Aconitum moldavicum and Caltha palustris extracts. Whilst the efficacy of extracts was modest against Trypanosoma brucei (IC50 ranging from 88.8 to 269.3 µg/mL), several extracts exhibited high potency against Leishmania infantum promastigotes (Aconitum vulparia IC50 18.8 µg/mL). We also tested them against the clinically relevant intracellular stage and found extract of A. vulparia to be the most effective (IC50 29.0 ± 1.1 µg/mL). All tested extracts showed no or low toxicity against FHs 74Int normal cell line (IC50 ranging from 152.9 to &gt;512 µg/mL). In conclusion, we suggest the above-mentioned plant extracts as potential candidates for development of novel plant-based antioxidant and/or antiproliferative and/or antileishmanial compounds.

Design, Synthesis, and Biological Activity Evaluation of Eco-Friendly Rosin-Diamide-Based Fungicides as Potential Succinate Dehydrogenase Inhibitors for Sustainable Crop Protection.

To develop novel succinate dehydrogenase (SDH) inhibitors for sustainable crop protection, a series of dehydroabietyl-diamide-based fungicides (a total of 21) were designed. In vitro fungicidal activity measurement showed that compound 3u exhibited excellent fungicidal activity against Valsa mali (half-maximal effective concentration, EC50 = 0.195 μg/mL), surpassing that of the positive control carbendazim (EC50 = 1.35 μg/mL). The in vivo fungicidal activity assessment suggested that 3u exhibited a protective effect on apple branches (69.7-48.1%) and apples (94.6-56.6%). Furthermore, biosafety evaluation indicated that 3u was significantly environmentally friendly toward zebrafish. Subsequently, morphology, physiology, and molecular docking were investigated to elucidate the mode of action of 3u against V. mali. Results demonstrated a strong binding between 3u and SDH, resulting in decreased SDH activity (half-maximal inhibitory concentration, IC50 = 11.7 μg/mL). Moreover, 3u disrupted the mycelial cell membrane and accelerated electrolyte leakage, ultimately resulting in the death of V. mali. These findings suggest that 3u could serve as a potent SDH inhibitor for sustainable crop protection.

Assessment of cytotoxicity of 5-arylaminouracil derivatives

We have previously shown that 5-arylaminouracil derivatives can inhibit HIV-1, herpesviruses, mycobacteria and other pathogens through various mechanisms. The purpose of this study was to evaluate the potential of 5-arylaminouracils and their derivatives against leukemia, neuroblastoma and glial brain tumors. The cytotoxicity of 5-aminouracils with various substituents, as well as their 5’-norcabocyclic and ribo derivatives, was screened against two neuroblastoma cell lines (SH-SY5Y and IMR-32), lymphoblastic cells K-562, promyeoloblastic cells HL-60 and low-passage variants of well-differentiated glioblastoma multiforme (GBM5522 and GBM6138). As a result of assessing the cytotoxicity of the resulting compounds on the above cell lines using the standard MTT test, it was revealed that most of the compounds do not have significant toxicity. However, in the GBM-6138 cell line, 5-(4-isopropylphenylamine)uracil and 5-(4-tert-butylphenylamine)uracil exhibited a dose-dependent toxic effect, with half-maximal inhibition concentrations IC50 of 9 μM and 2.3 μM, respectively. The antitumor activity of compounds of this type has been demonstrated for the first time and can serve as a starting point for further research.

Study of mRNA expression of thirteen genes of Trypanosoma evansi in response to diminazene aceturate and isometamidium chloride

The monomorphic, non-cyclic, extracellular haemoprotozoan parasite, Trypanosoma evansi leads to Surra disease in domesticated animals. Currently, diminazene aceturate (DA) and isometamidium chloride (ISM) are the most used chemotherapeutic agents for the treatment of Surra in animals. There is still little knowledge on the anti- trypanosomal mechanism of action of DA and ISM. The work addresses a significant gap in the understanding of the anti-typanosomal mechanism of DA and ISM by investigating their effects on mRNA expression profiles of 13 genes of T. evansi. The half maximal inhibitory concentration (IC50) of DA and ISM for a pony isolate of T. evansi was estimated as 335.3 nM and 308.6 nM, respectively. Transcript analysis of DA and ISM exposed T. evansi population showed its effects on the metabolic machinery of T. evansi by down-regulating the mRNA expression of all the 13 targeted genes. However, ISM exposure did not affect mRNA expression of Expression site-associated genes 8 (ESAG8), oligopeptidase B and ornithine decarboxylase genes. The finding provides valuable insights into the molecular action of these drugs, which is crucial for developing more effective treatment of Surra disease. Further, comprehensive transcriptome and proteomic analysis could provide a deeper insight into precise molecular pathway of these medications against T. evansi.

Onametostat, a PfPRMT5 inhibitor, exhibits antimalarial activity to Plasmodium falciparum.

Protein arginine methyltransferases (PRMTs) play critical roles in Plasmodium falciparum, a protozoan causing the deadliest form of malaria, making them potential targets for novel antimalarial drugs. Here, we screened 11 novel PRMT inhibitors against P. falciparum asexual growth and found that onametostat, an inhibitor for type II PRMTs, exhibited strong antimalarial activity with a half-maximal inhibitory concentration (IC50) value of 1.69 ± 0.04 µM. In vitro methyltransferase activities of purified PfPRMT5 were inhibited by onametostat, and a shift of IC50 to onametostat was found in the PfPRTM5 disruptant parasite line, indicating that PfPRTM5 is the primary target of onametostat. Consistent with the function of PfPRMT5 in mediating symmetric dimethylation of histone H3R2 (H3R2me2s) and in regulating invasion-related genes, onametostat treatment led to the reduction of H3R2me2s level in P. falciparum and caused the defects on the parasite's invasion of red blood cells. This study provides a starting point for identifying specific PRMT inhibitors with the potential to serve as novel antimalarial drugs.

Tyrosinase Inhibitory Activity of Crude Procyanidin Extract from Green Soybean Seed and the Stability of Bioactive Compounds in an Anti-Aging Skin Care Formulation

Green soybean (Glycine max L.) seed contains a high procyanidin content and high antioxidant activity. Moreover, ultrasound-assisted extraction (UAE) has proved to be advantageous in providing high extraction efficiency. Hence, this study aimed to extract procyanidins from green soybean seeds (GSSs) using UAE. This study also evaluated the inhibitory activities of tyrosinase and the cytotoxic effects of crude procyanidin extract. The extract exhibited maximum levels of bioactive components and antioxidant capacity when subjected to a temperature of 15 °C and an extraction time of 20 min. The crude procyanidin extract at a concentration of 10 mg/mL inhibited the tyrosinase enzyme by more than 60%, and the half-maximal inhibitory concentration (IC50) value obtained for the extract was 6.85 ± 0.81 mg/mL. This result was much greater than the IC50 value obtained for kojic acid (0.089 ± 0.08 mg/mL), which was used as a positive control. For the cytotoxicity assessment, the results indicated that the crude procyanidin extract showed no cytotoxicity and actually stimulated the growth of human skin fibroblast cells. More than 80% of the bioactive compounds (total phenolic content (TPC), total flavonoid content (TFC), procyanidin content (PC)) and antioxidant activities (DPPH and FRAP) of the crude extract powder were retained at 38.68 ± 0.01 mg GAE/g, 16.07 ± 0.01 mg CAE/g, 9.24 ± 0.01 mg PC/g, 359.8 ± 0.72 μM Trolox eq/g, and 1640 ± 2.86 μM Trolox eq/g, respectively, after 12 weeks of storage at 25 °C. The crude procyanidin extract powder was then included in a facial serum formulation and tested for pH value and physical evaluation. The stability of the crude procyanidin extract facial serum was shown to be greater for bioactive compounds and antioxidant activity when stored at a temperature of 4 °C than when stored at a temperature of 25 °C. These results suggest that the GSS extracts obtained via ultrasonication show promise for use in cosmeceutical formulations for whitening skincare products.

Methyl jasmonate effects on Lactuca serriola L.: Antioxidant defense and bioactive compound changes.

The effect of methyl jasmonate (MeJA) foliar spray on the activity of antioxidant enzymes-Superoxide dismutase (SOD), Catalase (CAT), Ascorbate peroxidase (APX), and Guaiacol peroxidase (GPX)-along with assessments of total phenolic and flavonoid contents and antioxidant activity (IC50), was examined in Prickly lettuce (Lactuca serriola L.). The study involved treating plants with three MeJA solutions (0, 200, and 400 µM) and harvesting samples at four distinct time intervals. Varied MeJA concentrations and time intervals resulted in a substantial increase in the activity of all the antioxidant enzymes investigated in this study. Both concentration levels and time courses exhibited progressive outcomes. Moreover, MeJA treatment led to elevated levels of total phenolic and flavonoid contents, reaching peaks of 17.02 (mg GAL/g DW) and 8.3 (mg QUE/g DW), respectively, particularly in response to the 400 µM concentration. However, the total flavonoid content did not show any significant variation between the two concentrations. Based on the half-maximal inhibitory concentration (IC50) values, the antioxidant activity in MeJA-treated plants was found to be lower compared to the controls. However, our findings suggest that, under specific conditions discussed in this study, MeJA has the potential to enhance the nutritional value of L. serriola.

12422 Linsitinib Inhibits Proliferation And Induces Apoptosis Of IGF-1R And TSH-R Expressing Cells

Abstract Disclosure: M. Luffy: None. A. Ganz: None. J. Ropertz: None. R. Douglas: Employee; Self; Sling Therapeutics. R. Zeidan: Employee; Self; Sling Therapeutics. J. Kent: Employee; Self; Sling Therapeutics. J. Wolf: None. G.J. Kahaly: Grant Recipient; Self; Sling Therapeutics. Background: The insulin-like growth factor 1 receptor (IGF-1R) and the thyrotropin receptor (TSH-R) are expressed on orbital target cells and thyrocytes. Targeting these pathways effectively improves thyroid eye disease (TED). Methods: The inhibitory effect of Linsitinib, a small molecule targeting the IGF-1R (Sling Therapeutics, Ann Arbor, MI, USA), on cell proliferation was investigated in an IGF-1R expressing cell line, NCI-H295R (ATCC, Manassas, VA, USA) and a Chinese Hamster Ovary (CHO) cell line overexpressing the TSH-R (QuidelOrtho, San Diego, CA, USA). An IGF-1R antagonist, Teprotumumab (BioVision, Milpitas, CA, USA) served as control. Both cell lines were plated in a 96-well format and treated with four concentrations of both compounds for 24 hours. After addition of tetrazolium, absorbance was measured (GloMax luminometer, Promega, Madison, WI, USA). Furthermore, the half-maximal inhibitory concentration (IC50) of TSH-R-Ab induced stimulation (stimulatory monoclonal antibody, mAb, M22, RSR, Cardiff, UK) of the TSH-R cell line was evaluated with a cell-based bioassay for blocking TSH-R-Ab (Thyretain TBI, QuidelOrtho). Cells were treated with ten rising concentrations of either Linsitinib, Linsitinib + Metformin, Teprotumumab, or a blocking TSH-R mAb (K1-70, RSR). The Kruskal-Wallis and/or the two-tailed Mann-Whitney tests were used for statistical comparisons. Results: Linsitinib strongly inhibited the proliferation of both cell lines at the following concentrations: 31,612.5 ng/mL (-78%, P=0.0031, IGF-1R cell line and -75%, P=0.0059, TSH-R cell line), as well as at 63,225 ng/mL (-73%, P=0.0073, IGF-1R cell line and -73%, P=0.0108, TSH-R cell line). Linsitinib induced morphologically confirmed apoptosis of both cell lines across all tested concentrations (Luna-FL cell counter, Logos Biosystems, Annandale, VA, USA). Compared to Teprotumumab, Linsitinib inhibited markedly more the proliferation of the IGF-1R cell line at all concentrations (Linsitinib versus Teprotumumab, P=0.0286). Teprotumumab inhibition ranged from -21 to -31% of the IGF-1R cell line across all concentrations, while it was significant only at 15,806.25 ng/mL with the TSH-R cell line (-15%, P=0.0396). In addition, in the TSH-R-Ab blocking bioassay, all tested compounds demonstrated strong inhibition across all ten dilutions (100%). The IC50 of Linsitinib was lower than Teprotumumab (147.1 ng/mL versus 160.6 ng/mL). Combining Metformin to Linsitinib did not further lower the IC50. Not surprisingly, the TSH-R blocking mAb K1-70 had the lowest IC50 of 20.57 ng/mL. Conclusions: Linsitinib effectively induces apoptosis and inhibits proliferation of both IGF-1R and TSH-R expressing target cells, also displaying a lower IC50 in the blocking TSH-R-Ab bioassay compared to Teprotumumab, hence demonstrating its therapeutic potential for TED. Presentation: 6/2/2024

Liposomal AZD5363 Displays Antiproliferation Activities and Induces Apoptosis on Y79 Retinoblastoma Cancer Cells

Objectives Retinoblastoma (RB) is an aggressive intraocular cancer that usually develops during infancy and childhood. As an Akt kinase inhibitor, AZD5363 is a novel drug whose encapsulation into liposomes enhances its bioavailability and biomedical potential. In the present study, a liposomal membrane was created around AZD5363 to assess its efficacy on the Y79 cancer cell line. Materials and Methods AZD5363 nanoparticles were synthesized by the thin film hydration method. Dynamic light scattering (DLS) and field emission scanning electron microscopy (FESEM) techniques were applied to evaluate the particle size, and the morphology of the liposomal AZD5363 (Lipo-AZD5363). The MTT test was used to assess the half maximal inhibitory concentration (IC50) of Lipo-AZD5363, and the cytotoxic effects of Lipo-AZD5363 and doxorubicin (Dox) were investigated on the Y79 cell line. Flow cytometry was used to study apoptotic induction in selected groups. Also, the PTEN/AKT/FOXO1 gene expression level was measured using quantitative reverse transcription polymerase chain reaction (qRT-PCR) assay. Results Treatment with Lipo-AZD5363 inhibited the proliferation of Y79 RB cancer cell line in a dose-dependent manner. Lipo-AZD5363, at a lower concentration, was significantly more cytotoxic than Dox in terms of enhanced cell death (p &lt; 0.05).Furthermore, flow cytometry showed that Lipo-AZD5363 and Dox induce apoptosis in these cells. However, the number of apoptotic cells in the Lipo-AZD5363 group was clearly higher than that in the Dox group (p &lt; 0.001). Real-time PCR analysis indicated that Lipo-AZD5363 treatment resulted in an increase in PTEN and FOXO1 gene expression and a decrease in AKT gene expression. Our study revealed that all results were statistically more significant in the Lipo-AZD5363 group than in the Dox group (p &lt; 0.01, &lt;0.01, and &lt;0.001, respectively). Conclusion Lipo-AZD5363 inhibits proliferation and promotes apoptosis of RB cells by inhibiting the PI3K/AKT signaling pathway. Thus, Lipo-AZD5363 may be a promising candidate for cancer therapy. However, more experimental evidence is needed for its use in the pharmacological treatment of RB.