N-phenyl pyrazoline derivative inhibits cell aggressiveness and enhances paclitaxel sensitivity of triple negative breast cancer cells.

Protein kinase dysregulation induces cancer cell aggressiveness leading to rapid tumor progression and poor prognosis in TNBC patients. Many small-molecule kinase inhibitors have been tested in clinical trials to treat TNBC patients. In the previous study, we found that N-phenylpyrazoline small molecule acts as a protein kinase inhibitor in cervical cancer cells. However, there remains unknown about N-phenyl pyrazoline potency as a kinase inhibitor and its anti-cancer activity in TNBC cells. In this study, we investigated the activity of N-phenyl pyrazoline against TNBC cells via tyrosine kinase inhibition. Based on the MTT assay, the IC50 values for the N-phenyl pyrazoline 2, 5, A, B, C, and D against Hs578T were 12.63 µM, 3.95 µM, not available, 18.62 µM, 30.13 µM, and 26.79 µM, respectively. While only P5 exhibited the IC50 against MDA MB 231 (21.55 µM). Further, N-phenyl pyrazoline 5 treatment significantly inhibited the cell proliferation rate of Hs578T and MDA MB 231 cells. The migration assay showed that treatment with the compound N-phenyl pyrazoline 5 with 4 µM concentration significantly reduced cell migration of Hs578T cells. N-phenyl pyrazoline 5 treatment at 1 µM and 2 µM was able to reduce the tumorsphere size of Hs578t cells. A combination treatment of P5 and paclitaxel showed a synergistic effect with a combination index score > 1 in both TNBC cells. Further, the P5 predictively targeted the protein kinases that significantly correlated to breast cancer prognosis. The GSEA analysis result shows that receptor tyrosine kinase, Notch3, Notch4, and Ephrin signaling pathways were targeted by P5. The P5 treatment reduced the EGFR expression level and activation in TNBC cells.

Pyrazoline B-Paclitaxel or Doxorubicin Combination Drugs Show Synergistic Activity Against Cancer Cells: In silico Study.

Background: Multidrug resistance in various cancer types is a major obstacle in cancer treatment. The concept of a single drug molecular target often causes treatment failure due to the complexity of the cellular processes. Therefore, combination chemotherapy, in which two or more anticancer drugs are co-administered, can overcome this problem because it potentially have synergistic efficacy besides reducing resistance, and drug doses. Previously, we reported that pyrazoline B had promising anticancer activity in both in silico and in vitro studies. To increase the efficacy of this drug, co-administration with established anticancer drugs such as doxorubicin and paclitaxel is necessary. Materials and Methods: In this study, we used an in silico approach to predict the synergistic effect of pyrazoline B with paclitaxel or doxorubicin using various computational frameworks and compared the results with those of an established study on the combination of doxorubicin-cyclophosphamide and paclitaxel-ascorbic acid. Results and Discussion: Drug interaction analysis showed the combination was safe with no contraindications or side effects. Furthermore, molecular docking studies revealed that doxorubicin-pyrazoline B and doxorubicin-cyclophosphamide may synergistically inhibit cancer cell proliferation by inhibiting the binding of topoisomerase I to the DNA chain. Moreover, the combination of pyrazoline B-paclitaxel may has synergistic activity to cause apoptosis by inhibiting Bcl2 binding to the Bax fragment or inhibiting cell division by inhibiting α-ß tubulin disintegration. Paclitaxel-ascorbic acid had a synergistic effect on the inhibition of α-ß tubulin disintegration. Conclusion: The results show that this combination is promising for further in vitro and in vivo studies.

Isolation of active compounds from Streptomyces sennicomposti GMY01 and cytotoxic activity on breast cancer cells line.

The occurrence of resistance to anticancer and the emergence of serious side effects due to chemotherapy is one of the main problems in cancer treatment, including breast cancer. The need for effective anticancer with a specific target is urgently required. Streptomyces are widely known as the potential producers of new anticancer molecules. Previously reported that the methanol extract of Streptomyces sennicomposti GMY01 isolated from Krakal Coast, Gunungkidul had very strong cytotoxic activity against MCF-7 and T47D breast cancer cells with IC50 values of 0.6 and 1.3 µg/mL, respectively. The following study aimed to isolate and identify active compounds of the S. sennicomposti GMY01 and evaluate its cytotoxic activity. The study was started by re-culturing and re-fermented optimization of S. sennicomposti GMY01 in a larger volume, then the bacteria were extracted using methanol following the bioassay-guided isolation of the extract obtained. The active compounds obtained were then structurally determined using UV/Vis spectroscopy, Fourier Transform-Infrared (FT-IR), Liquid Chromatography-Mass Spectroscopy (LC-MS), 1H NMR, and 13C NMR and analyzed for their cytotoxic activity using MTT assay on MCF-7 and normal Vero cells line. The results showed that the culture of the S. sennicomposti GMY01 using Starch Nitrate Broth (SNB) media yields the best results compared to other culture media. An active anticancer compound namely mannotriose was successfully isolated from the methanol extract with an IC50 value of 5.6 µg/mL and 687 µg/mL against the MCF-7 and Vero cells lines, respectively, indicating that this compound showed strong cytotoxic activity with high selectivity.

Marine-Derived Streptomyces sennicomposti GMY01 with Anti-Plasmodial and Anticancer Activities: Genome Analysis, In Vitro Bioassay, Metabolite Profiling, and Molecular Docking.

To discover novel antimalarial and anticancer compounds, we carried out a genome analysis, bioassay, metabolite profiling, and molecular docking of marine sediment actinobacteria strain GMY01. The whole-genome sequence analysis revealed that Streptomyces sp. GMY01 (7.9 Mbp) is most similar to Streptomyces sennicomposti strain RCPT1-4T with an average nucleotide identity (ANI) and ANI based on BLAST+ (ANIb) values of 98.09 and 97.33% (>95%). An in vitro bioassay of the GMY01 bioactive on Plasmodium falciparum FCR3, cervical carcinoma of HeLa cell and lung carcinoma of HTB cells exhibited moderate activity (IC50 value of 46.06; 27.31 and 33.75 µg/mL) with low toxicity on Vero cells as a normal cell (IC50 value of 823.3 µg/mL). Metabolite profiling by LC-MS/MS analysis revealed that the active fraction of GMY01 contained carbohydrate-based compounds, C17H29NO14 (471.15880 Da) as a major compound (97.50%) and mannotriose (C18H32O16; 504.16903 Da, 1.96%) as a minor compound. Molecular docking analysis showed that mannotriose has a binding affinity on glutathione reductase (GR) and glutathione-S-transferase (GST) of P. falciparum and on autophagy proteins (mTORC1 and mTORC2) of cancer cells. Streptomyces sennicomposti GMY01 is a potential bacterium producing carbohydrate-based bioactive compounds with anti-plasmodial and anticancer activities and with low toxicity to normal cells.

Antibiofilm Activity of Invasive Plants against Candida albicans: Focus on Baccharis halimifolia Essential Oil and Its Compounds.

The extracts of five invasive plants were investigated for antifungal and antibiofilm activities against Candida albicans, C. glabrata, C. krusei, and C. parapsilosis. The antifungal activity was evaluated using the microdilution assay and the antibiofilm effect by measurement of the metabolic activity. Ethanol and ethanol-water extracts of Reynoutria japonica leaves inhibited 50 % of planktonic cells at 250 µg mL-1 and 15.6 µg mL-1 , respectively. Ethanol and ethanol-water extracts of Baccharis halimifolia inhibited >75 % of the mature biofilm of C. albicans at 500 µg mL-1 . The essential oil (EO) of B. halimifolia leaves was the most active (50 % inhibition (IC50 ) at 4 and 74 µg mL-1 against the maturation phase and 24 h old-biofilms of C. albicans, respectively). Oxygenated sesquiterpenes were the primary contents in this EO (62.02 %), with ß-caryophyllene oxide as the major component (37 %). Aromadendrene oxide-(2), ß-caryophyllene oxide, and (±)-ß-pinene displayed significant activities against the maturation phase (IC50 =9-310 µ mol l-1 ) and preformed 24 h-biofilm (IC50 =38-630 µ mol l-1 ) of C. albicans with very low cytotoxicity for the first two compounds. C. albicans remained the most susceptible species to this EO and its components. This study highlighted for the first time the antibiofilm potential of B. halimifolia, its EO and some of its components.

Screening antibiofilm activity of invasive plants growing at the Slope Merapi Mountain, Central Java, against Candida albicans.

BACKGROUND: Candida albicans causes high-mortality candidiasis. Antifungal drug resistance demands the development of virulence factor-targeting drugs, particularly antibiofilm. This study screened the effects of five invasive plants growing in Indonesia (Mimosa pudica, Lantana camara, Acacia mangium, Ageratina riparia, and Mikania micrantha) against C. albicans biofilms. Antifungal activity, antiphospholipase activity, biofilm morphology of C. albicans, and cytotoxic capacity were also evaluated. METHODS: Maceration was used to extract the plants, and the most active extract inhibiting the biofilms was fractionated using liquid-liquid fractionation. Antibiofilm activity was determined by a colorimetric assay, MTT. Antifungal activity was tested using the broth microdilution method. A phospholipase assay was performed using the egg-yolk agar method. Influence on the C. albicans morphology was assessed using scanning electron microscopy (SEM). The cytotoxic effect was carried out against Vero and HeLa cell lines. RESULTS: M. pudica extracts showed the most potent antifungal efficacy with minimum inhibitory concentration (MIC) of 15.62 µg/mL and 7.81 µg/mL for aerial parts and roots, respectively. At high concentrations (500 µg/mL and 250 µg/mL), ethanol extract of M. pudica aerial parts strongly inhibited the phospholipase activity. Ethyl-acetate fraction of M. pudica aerial parts demonstrated the most potent antibiofilm activity against 24 h old biofilm of C. albicans with an inhibitory concentration (53.89%) of 62.5 µg/mL showed no cytotoxicity in both Vero and HeLa cells. This fraction affected the morphology of C. albicans and contained promising compounds for inhibiting the 24 h old biofilm of C. albicans. CONCLUSIONS: Invasive M. pudica plant inhibited the growth of planktonic C. albicans cells and its ethyl acetate fraction decreased the metabolic activity of C. albicans biofilms. This result demonstrates the potential of invasive M. pudica plant to reduce biofilm-associated candida infection.

Chalcone-3 Inhibits the Proliferation of Human Breast Cancer MDA-MB-231 Cell Line.

OBJECTIVE: Chalcone-3 has been shown to be cytotoxic and selective against luminal subtype breast cancer cell lines, which are suspected to occur through the mechanism of epidermal growth factor receptors (EGFR) inhibition. However, the cytotoxic effect has never been tested on cell strains from patients with triple negative breast cancer (TNBC), where EGFR expression is known to increase. This study aimed to identify the role of chalcone-3 in one of the downstream targets of EGFR as an antiproliferative agent. METHODS: Chalcone-3 was examined for its effect on proliferation in human breast cancer MDA-MB-231 cell lines. The percentage of proliferation inhibition was analyzed using methyl-thiazol tetrazolium assay. Flow cytometry was used to analyze the population of cell cycle distribution and the expression of cyclin-D1 and pEGFR. RESULTS: Chalcone-3 inhibited the proliferation of MDA-MB-231 cells in a dose and time-dependent manner with an IC50 value of 17.98±6.36 µg/mL by inducing cell cycle arrest at the G2/M phase. Flow cytometry assays showed that chalcone-3 significantly reduced the expression of pEGFR and cyclin-D1, contributing to cell cycle arrest. CONCLUSION: Chalcone-3 might have potential as an anti-proliferative drug to treat TNBC.

In Vitro Insulin Resistance Model: A Recent Update.

Insulin resistance, which affects insulin-sensitive tissues, including adipose tissues, skeletal muscle, and the liver, is the central pathophysiological mechanism underlying type 2 diabetes progression. Decreased glucose uptake in insulin-sensitive tissues disrupts insulin signaling pathways, particularly the PI3K/Akt pathway. An in vitro model is appropriate for studying the cellular and molecular mechanisms underlying insulin resistance because it is easy to maintain and the results can be easily reproduced. The application of cell-based models for exploring the pathogenesis of diabetes and insulin resistance as well as for developing drugs for these conditions is well known. However, a comprehensive review of in vitro insulin resistance models is lacking. Therefore, this review was conducted to provide a comprehensive overview and summary of the latest in vitro insulin resistance models, particularly 3T3-L1 (preadipocyte), C2C12 (skeletal muscle), and HepG2 (liver) cell lines induced with palmitic acid, high glucose, or chronic exposure to insulin.

Metabologenomics approach to the discovery of novel compounds from Streptomyces sp. GMR22 as anti-SARS-CoV-2 drugs.

COVID-19 is spreading rapidly yet there is no clinically proven drug available now. Soil-derived Streptomyces sp. GMR22 has a large genome size (11.4 Mbp) and a huge BGCs (Biosynthetic Gene Clusters) encoding secondary metabolites. This bacterium is a potential source for producing a wide variety of compounds which are able to block SARS-CoV-2, the causative agent of COVID-19. This study aimed to predict the secondary metabolites of Streptomyces sp. GMR22 and to evaluate the ability as SARS-CoV-2 inhibitor. The AntiSMASH 5.0 was used for genome mining analysis and targeted liquid chromatography-high resolution mass spectrometry (LC-HRMS) was used for metabolite analysis. In silico molecular docking was performed on important target proteins of SARS-CoV-2 i.e., spike protein (PDB ID: 6LXT), Receptor Binding Domain (RBD)-ACE2 (Angiotensin-Converting Enzyme 2) (PDB ID: 6VW1), 3CLpro (3-chymotrypsin-like protease) (PDB ID: 6M2N), and RdRp (RNA-dependent RNA polymerase) (PDB ID: 6M71). Two compounds from GMR22 extract, echoside A and echoside B were confirmed by targeted LC-HRMS and potential as SARS-CoV-2 inhibitor. Echoside A and echoside B showed higher docking score than remdesivir as COVID-19 drug on four target proteins, i.e., spike protein (-7.9 kcal/mol and -7.8 kcal/mol), RBD-ACE2 (-7.5 kcal/mol and -8.2 kcal/mol), 3CLpro (-8.4 kcal/mol and -9.4 kcal/mol) and RdRp (-7.3 kcal/mol and -8.0 kcal/mol). A combination of genome mining and metabolomic approaches can be used as integrated strategy to elucidate the potential of GMR22 as a resource in the discovery of anti-COVID -19 compound.

Two strains of airborne Nocardiopsis alba producing different volatile organic compounds (VOCs) as biofungicide for Ganoderma boninense.

Nocardiopsis are actinobacteria which produce active compounds, such as antifungals and volatile compounds. Ganoderma boninense is a pathogenic and aggressive fungus that decreases palm oil yield during production. In this study, we isolated two strains of Nocardia (GME01 and GME22) from airborne contaminants on the actinobacteria culture collection in the laboratory. The aim of this study is to identify two strains of Nocardiopsis and to obtain the antifungal potency of volatile organic compounds (VOCs) against G. boninese. We characterized the morphology using Scanning Electrone Microscope (SEM), molecular properties and whole-cell protein spectra using Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS), antifungal assay on G. boninense and VOCs analysis of Nocardia using solid phase micro extraction/gas chromatography (SPME/GC). The two Nocardiopsis strains had the similar characteristic such as white aerial mycelium and spores, aerobic, grow well on ISP-2, TSA and NA medium without diffusible pigment and had the highest similarity with Nocardiopsis alba DSM 43377 (99.63% and 99.55% similarity for GME01 and GME22, respectively), Different morphological feature was found in aerial mycelium and spores. GME22 has a clearly fragmented mycelium whereas GME01 has none. Other features also showed different on the whole-cell protein spectra, antifungal activity and VOCs profiles. Antifungal activity assay on G. boninense showed that N. alba GME22 has higher antifungal activity than GME01 related with the VOCs abundance in two strains. Almost 38.3% (18 VOCs) of N. alba GME22 and 25.5% (12 VOCs) of N. alba GME01 were found specifically in each strain, and 36.2% (the 17 same VOCs) produced by both. The known volatile antifungal compounds S-methyl ethanethioate, 1,2-dimethyldisulfane, acetic acid, 2-methyl propanoic acid, 3-methyl-butanoic acid, nonan-2-one, undecan-2-one and 2-isopropyl-5-methylcyclohexan-1-ol only produced by N. alba GME22 and 1,3-dimethyltrisulfane only produced by N. alba GME01. A total of two known antifungal compounds 1,2-dimethyldisulfane and 6-methylheptan-2-one were produced by both N. alba. The abundance of antifungal VOCs produced by these bacteria is potentially to be used as biocontrol agent for pathogenic fungi in plants.

Folic Acid Treatment for Patients with Vascular Cognitive Impairment: A Systematic Review and Meta-Analysis.

BACKGROUND: As the life expectancy of elderly people has drastically increased, the incidence of cardiovascular and cerebrovascular diseases in this population has proportionally grown. Vascular cognitive impairment (VCI) refers to all forms of cognitive disorder associated with cerebrovascular disease. Homocysteine has recently been recognized as a contributor to the pathomechanisms involved in cognitive impairment. B vitamins, such as folic acid, are known to be effective in lowering homocysteine levels. AIM OF THE STUDY: To evaluate the efficacy of folic acid in patients with VCI. METHODS: We conducted a systematic review and meta-analysis of research on folic acid treatments for VCI. Only randomized controlled trials studies that compared the efficacy of folic acid to placebo or other interventions were considered, irrespective of publication status, year of publication, and languages. Two independent reviewers searched the Medline via Ovid, EMBASE and Cochrane Central Register of Controlled Trials (Central) journal databases up to July 2021 and independently appraised the included studies. We used mean difference outcome with 95% confidence intervals (CI) to calculate the change of Mini-Mental State Examination (MMSE), cognitive function domain, and concentration of homocysteine. RESULTS: We found three studies comparing folic acid with placebo and one study comparing folic acid with other interventions. There is only slight evidence that the MMSE score in patients who received Folic Acid increased 0.3 point higher compared to the placebo group after 24 months (95% CI:-0.12-0.37; p=0.31). There is very strong evidence that the concentration of Homocysteine in the Folic Acid group became 6.16 µmol/L lower compared to the placebo group after 6 months (95% CI:2.32-8.21 lower; p<0.001). CONCLUSIONS: Our review shows the effectiveness of folic acid in lowering plasma homocysteine concentration after 6 months period compared to placebo. However, this effect is not accompanied by improvement in cognitive function.

In Vitro Antiplasmodial, Heme Polymerization, and Cytotoxicity of Hydroxyxanthone Derivatives.

The previous study showed that xanthone had antiplasmodial activity. Xanthone, with additional hydroxyl groups, was synthesized to increase its antiplasmodial activity. One of the strategies to evaluate a compound that can be developed into an antimalarial drug is by testing its mechanism in inhibiting heme polymerization. In acidic condition, hematin can be polymerized to ß-hematin in vitro, which is analog with hemozoin in Plasmodium. This study was conducted to evaluate the antiplasmodial activity of hydroxyxanthone derivative compounds on two strains of Plasmodium falciparum 3D-7 and FCR-3, to assess inhibition of heme polymerization activity and determine the selectivity of hydroxyxanthone derivative compounds. The antiplasmodial activity of each compound was tested on Plasmodium falciparum 3D-7 and FCR-3 with 72 hours incubation period, triplicated in three replications with the microscopic method. The compound that showed the best antiplasmodial activity underwent flow cytometry assay. Heme polymerization inhibition test was performed using the in vitro heme polymerization inhibition activity (HPIA) assay. The antiplasmodial activity and heme polymerization inhibition activity were expressed as the 50% inhibitory concentration (IC50). In vitro cytotoxicity was tested using the MTT assay method on Vero cell lines to determine its selectivity index. The results showed that among 5-hydroxyxanthone derivative compounds, the 1,6,8-trihydroxyxanthone had the best in vitro antiplasmodial activity on both 3D-7 and FCR-3 Plasmodium falciparum strains with IC50 values of 6.10 ± 2.01 and 6.76 ± 2.38 µM, respectively. The 1,6,8-trihydroxyxanthone showed inhibition activity of heme polymerization with IC50 value of 2.854 mM and showed the high selectivity with selectivity index of 502.2-556.54. In conclusion, among 5-hydroxyxanthone derivatives tested, the 1,6,8-trihydroxyxantone showed the best antiplasmodial activity and has heme polymerization inhibition activity and high selectivity.

Screening of Indonesian peat soil bacteria producing antimicrobial compounds.

The development and world-wide spread of multidrug-resistant (MDR) bacteria have a high concern in the medicine, especially the extended-spectrum of beta-lactamase (ESBL) producing Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA). There are currently very limited effective antibiotics to treat infections caused by MDR bacteria. Peat-soil is a unique environment in which bacteria have to compete each other to survive, for instance, by producing antimicrobial substances. This study aimed to isolate bacteria from peat soils from South Kalimantan Indonesia, which capable of inhibiting the growth of Gram-positive and Gram-negative bacteria. Isolates from peat soil were grown and identified phenotypically. The cell-free supernatant was obtained from broth culture by centrifugation and was tested by agar well-diffusion technique against non ESBL-producing E. coli ATCC 25922, ESBL-producing E. coli ATCC 35218, methicillin susceptible Staphylococcus aureus (MSSA) ATCC 29,213 and MRSA ATCC 43300. Putative antimicrobial compounds were separated using SDS-PAGE electrophoresis and purified using electroelution method. Antimicrobial properties of the purified compounds were confirmed by measuring the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). In total 28 isolated colonies were recovered; three (25PS, 26PS, and 27PS) isolates produced proteins with strong antimicrobial activities against both reference strains. The substance of proteins from three isolates exerted strong antimicrobial activity against ESBL-producing E. coli ATCC 35,218 (MIC = 2,80 µg/mL (25PS), 3,76 µg/mL (26PS), and 2,41 µg/mL (27PS), and MRSA ATCC 43,300 (MIC = 4,20 µg/mL (25PS), 5,65 µg/mL (26PS), and 3,62 µg/mL (27PS), and also had the ability bactericidal properties against the reference strains. There were isolates from Indonesian peat which were potentials sources of new antimicrobials.

The Effect of (E)-1-(4'-aminophenyl)-3-phenylprop-2-en-1-one on MicroRNA-18a, Dicer1, and MMP-9 Expressions against DMBA-Induced Breast Cancer.

BACKGROUND: Most of breast cancer patients are estrogen receptor alpha-positive and have high resistance and side effect of chemotherapeutic drug. Therefore, discovering an effective anticancer agent is needed. This research explored the effect of (E)-1-(4'-aminophenyl)-3-phenylprop-2-en-1-one (APE) on miR-18a, Dicer1, and MMP-9 expressions. METHODS: Twenty four female Sprague-Dawley rats were invetigated in this study. The rats were divided into 6 groups of 4. G1 was considered as normal rat. G2, G3, T1, T2, and T3 were given DMBA 20 mg/kgBW twice a week for 5 weeks to induce mammary cancer. After being affiliated with cancer, G2 was given vehicle and G3 was treated with tamoxifen. T1, T2, and T3 were treated with APE intraperitoneally everyday for 21 days at doses of 5, 15, and 45 mg/kgBW/day, respectively. Blood plasma was collected to measure miR-18a expression using qRT-PCR. Mammary tissues were also collected to determine Dicer1 and MMP-9 expressions by using  immunohistochemistry. RESULTS: The results showed significant down-regulation of miR-18a relative expression and up-regulation of Dicer1 expression in G3 and T1 compared to G2 (P<0.05). MMP-9 expression has significant decrease in T1 compared to G2 (P<0.05). CONCLUSION: APE can decrease miR-18a and MMP-9 expressions and increase Dicer1 expression in rat mammary cancer. Therefore, this compound could be a candidate of novel anticancer.

Synergistic Effect of 1,3,6-Trihydroxy-4,5,7-Trichloroxanthone in Combination with Doxorubicin on B-Cell Lymphoma Cells and Its Mechanism of Action Through Molecular Docking.

BACKGROUND: The increasing rate of cancer chemoresistance and adverse side effects of therapy have led to the wide use of various chemotherapeutic combinations in cancer management, including lymphoid malignancy. OBJECTIVE: We investigated the effects of a combination of 1,3,6-trihydroxy-4,5,7-trichloroxanthone (TTX) and doxorubicin on the Raji lymphoma cell line. METHODS: Raji cells were treated with different concentrations of TTX, doxorubicin, or combinations thereof. Cancer cell growth inhibition was evaluated using 3-(4,5-dimethyltiazol-2-yl)-2,5- diphenyltetrazolium bromide/MTT assay to determine the half-maximal inhibitory concentration. Combination index values were calculated using CompuSyn (ComboSyn, Inc, Paramus, NJ). Molecular docking was conducted using a Protein-Ligand ANT System. RESULTS: The mean (SD) half-maximal inhibitory concentration values of TTX and doxorubicin were 15.948 (3.101) µM and 25.432 (1.417) µM, respectively. The combination index values of the different combinations ranged from 0.057 to 0.285, indicating strong to very strong synergistic effects. The docking study results reveal that TTX docks at the active site of Raf-1 and c-Jun N-kinase receptors with predicted free energies of binding of -79.37 and -75.42 kcal/mol, respectively. CONCLUSIONS: The xanthone-doxorubicin combination showed promising in vitro activity against lymphoma cells. The results also indicate that the TTX and doxorubicin combination's effect was due to the interaction between TTX with Raf-1 and c-Jun N-kinase receptors, 2 determinants of doxorubicin resistance progression.

Predictors of COVID-19 severity: a systematic review and meta-analysis.

Background: The unpredictability of the progression of coronavirus disease 2019 (COVID-19) may be attributed to the low precision of the tools used to predict the prognosis of this disease. Objective: To identify the predictors associated with poor clinical outcomes in patients with COVID-19. Methods: Relevant articles from PubMed, Embase, Cochrane, and Web of Science were searched as of April 5, 2020. The quality of the included papers was appraised using the Newcastle-Ottawa scale (NOS). Data of interest were collected and evaluated for their compatibility for the meta-analysis. Cumulative calculations to determine the correlation and effect estimates were performed using the Z test. Results: In total, 19 papers recording 1,934 mild and 1,644 severe cases of COVID-19 were included. Based on the initial evaluation, 62 potential risk factors were identified for the meta-analysis. Several comorbidities, including chronic respiratory disease, cardiovascular disease, diabetes mellitus, and hypertension were observed more frequent among patients with severe COVID-19 than with the mild ones. Compared to the mild form, severe COVID-19 was associated with symptoms such as dyspnea, anorexia, fatigue, increased respiratory rate, and high systolic blood pressure. Lower levels of lymphocytes and hemoglobin; elevated levels of leukocytes, aspartate aminotransferase, alanine aminotransferase, blood creatinine, blood urea nitrogen, high-sensitivity troponin, creatine kinase, high-sensitivity C-reactive protein, interleukin 6, D-dimer, ferritin, lactate dehydrogenase, and procalcitonin; and a high erythrocyte sedimentation rate were also associated with severe COVID-19. Conclusion: More than 30 risk factors are associated with a higher risk of severe COVID-19. These may serve as useful baseline parameters in the development of prediction tools for COVID-19 prognosis.

The effects of soursop supplementation on blood pressure, serum uric acid, and kidney function in a prehypertensive population in accordance with the 2017 ACC/AHA guideline.

Soursop consumption is beneficial to health, but there have been few clinical studies observing its benefit in human subjects. We investigated the effects of soursop supplementation on blood pressure (BP), serum uric acid (SUA), and kidney function. A total of 143 subjects were included in this randomized controlled trial. Subjects were selected from a prehypertension population dataset (n = 4190) in the "Mlati Study Database" in 2007 (using the Joint National Committee (JNC) 7 guideline). After 10 years, 143 samples showed essential prehypertension combined with high-normal SUA levels. Subjects were randomly allocated into two groups, i.e., the treatment and control group. For a 3-month period, the treatment group was given 2 × 100 g soursop fruit juice per day and the control group was not treated. Using the JNC 7 guideline, the treatment group showed a significantly lower mean systolic BP after being adjusted by three times of examinations (baseline, week 6 and 12) compared with the control group. Furthermore, the control group was more likely to have prehypertension, hypertension, and high-normal and high SUA levels after 6 weeks, as well as after 12 weeks, compared with the treatment group. An additional analysis using the 2017 ACC/AHA guideline for subjects with stage 1 hypertension showed results similar to that using the JNC 7 guideline. Moreover, it indicated that mean of both systolic and diastolic BP of the treatment group was significantly lower compared with the control group after 12 weeks of treatment. We conclude that soursop supplementation can lower BP and SUA levels.

Chlorogenic acid ameliorates memory loss and hippocampal cell death after transient global ischemia.

Chlorogenic acid (CGA) is known to have antioxidant potentials, yet the effect of CGA on brain ischemia has not been sufficiently understood. Brain ischemia such as transient global ischemia disrupts many areas of the brain of rats, including the hippocampus. Male Wistar rats were randomly assigned into five groups, that is, sham-operated (SO), bilateral common carotid occlusion (BCCO), and BCCO+ 15, 30, and 60 mg/kg bw CGA groups (CGA15, CGA30, and CGA60, respectively). Brain ischemia was induced in Wistar rats with BCCO for 20 min followed by intraperitoneal injection of CGA. The rats were examined for the spatial memory in a Morris water maze test on the 3rd day and were euthanized on the 10th day after BCCO. The total number of pyramidal cells was estimated, and the mRNA expressions of Bcl2, Bax, caspase-3, SOD2, SOD1, GPx, ET-1, eNOS, CD31, and VEGF-A were measured. The BCCO group spent less time and distance in the target quadrant than any other group in the spatial memory retention test. The CA1 pyramidal cell numbers in the BCCO and CGA15 groups were lower than in the CGA30 and CGA60 groups. The mRNA expressions of Bcl2, SOD2, and CD31 in the BCCO group were lower than in the CGA15, CGA30, and CGA60 groups. The ET-1 expression was higher in the BCCO and CGA15 groups than in the SO, CGA30, and CGA60 groups. CGA improves the spatial memory and prevents the CA1 pyramidal cell death after BCCO by increasing Bcl2, SOD2, and CD31 expressions and decreasing ET-1 expression.

Metformin Modulates Cyclin D1 and P53 Expression to Inhibit Cell Proliferation and to Induce Apoptosis in Cervical Cancer Cell Lines.

Background: Cervical cancer is one of the most prevalent gynecological cancers worldwide and contributes in high mortality of Indonesian women. The efficacy of chemotherapy as a standart therapy for cervical cancer decreases because it frequenly rises adverse effects. Recent studies have found that metformin has a potential anticancer effect mostly through reduction of cyclin expression and activation of Activated Adenosine Monophosphate Kinase (AMPK). This study aimed to investigate the effect of metfomin on expression of cyclin D1 and p53 and apoptosis in HeLa cancer cell line. Methods: HeLa cells were treated with various doses of metformin and doxorubicin as a positive control. Cytotoxic effect of metformin was determined using the MTT assay. Immunocytochemistry was used to assess cyclin D1 and p53 expression and apoptosis levels of treated HeLa cells were analyzed using flowcytometry. Data of cyclin D1 expression was statistically analyzed using the Kruskal-Wallis test followed by the Tamhane test, whilst ANOVA and Tukey post Hoc tests were used to analyze data of p53 and apoptosis level. The significant value was p< 0.05. Results: Metformin was able to inhibit proliferation of HeLa cells with IC50 60 mM. HeLa cells treated with 60 and 120 mM metformin had lower cyclin D1 expression than HeLa cells treated without metformin and reached a significant difference (p= 0.001). Moreover, 30 mM or higher doses of metformin increase significantly p53 expression (p< 0.001). Induction of apoptosis was observed in HeLa cells treated with all doses of metformin and reached statistically difference (p= 0.04 and p < 0.001). Conclusion: Metformin can modulate cyclin D1 and p53 expression in HeLa cancer cell line, leading to inhibition of cell proliferation and induction of apoptosis. Other cyclin family members, CDK inhibitors and AMPK signaling should be further investigated in order to know mechanism of metformin action.

Synergistic Action of 1,2-Epoxy-3 (3- (3,4-dimethoxyphenyl)- 4H-1-benzopiyran-4-on) Propane with Doxorubicin and Cisplatin through Increasing of p53, TIMP-3, and MicroRNA-34a in Cervical Cancer Cell Line (HeLa)

Objective: Cervical cancer is the second most common cancer among women worldwide, with a high mortality rate especially in developing countries. Insufficient treatment for cervical cancer, multiple side effects, and high drug prices encourage researchers to look for effective and selective cancer drugs with appropriate molecular targets. This study explored the cytotoxicity of (1,2-epoxy-3(3-(3,4-dimethoxyphenyl)-4H-1-benzopyran-4-on) propane (EPI) synthesized from clove leaves oil on HeLa cells, its combination with doxorubicin (DOX) and cisplatin (CIS), and also their influence on p53, TIMP-3, and miR-34a as therapeutic targets. Materials and Methods: This research was an experimental in vitro study on cervical cancer uteri culture. The cytotoxicity was analyzed by MTT assay. The drug combination synergisms were indicated by the combination index (CI) (using CompuSyn 1.4). HeLa cells in 32 wells were divided into eight groups as negative control, which were given EPI ½IC50, EPI IC50, EPI 2IC50, DOX IC50, combination of EPI+DOX, CIS, and the combination of EPI+CIS. The p53 and TIMP-3 concentrations were measured using ELISA, and expressions of miR-34a with qRT-PCR. One-way ANOVA and post hoc Tukey tests were performed to determine the mean difference of all variables between the study groups. Results: IC50 for EPI was 33.24 (±3.01) µg/ml, while DOX and CIS were 4.8 µg/ml (±0.1), and 23.34 µg/ml (±3.01), respectively, while CI values for EPI-DOX were <0.1 and for EPI-CIS <0.9. Expression of p53 in group 6 (1.67±0.31) µg/ml and 8 (1.18±0.18) µg/ml, TIMP-3 6 (3.81±0.49) µg/ml and 8 (2.93±0.42) µg/ml were significantly higher compared to the control group (p<0.05). All treatment groups showed significantly increased miR-34a expressions compared to the control group (p<0.05). Conclusion: The combinations showed a very strong synergism and a moderate slight synergism for EPI-DOX and EPI-CIS. Both combinations were able to increase the expressions of p53, TIMP-3 proteins, and MiR-34a in the HeLa cells.