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.

Mitochondrial ROS induced by ML385, an Nrf2 inhibitor aggravates the ferroptosis induced by RSL3 in human lung epithelial BEAS-2B cells.

Ferroptosis is a new type of cell death marked by iron and lipid ROS accumulation. GPX4 is one of the glutathione peroxidases known to regulate ferroptosis tightly. On the other hand, Nrf2 also plays a vital role in ferroptosis as it targets genes related to oxidant defense. Herein, we employed beas-2 human epithelial cells treated with a low concentration of RSL3 to induce ferroptosis. To study the protective role of Nrf2, we used ML385 as its specific inhibitor. A combination of ML385 and a low concentration of RSL3 synergistically induced more toxicity to RSL3. Furthermore, we found that mitochondrial ROS is elevated in ML385 and RSL3 combination group. In addition, Mito TEMPOL application successfully prevents the upregulation of mitochondrial ROS, lipid ROS, reduces the toxicity of RSL3, restores the antioxidant capacity of the cells, and mitochondrial functions reflected by mitochondrial membrane potential and mitochondrial oxidative phosphorylation system (OXPHOS) expression. Altogether, our study demonstrated that Nrf2 inhibition by ML385 induces more toxicity when combined with RSL3 through the elevation of mitochondrial ROS and disruption of mitochondrial function.

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.

Bioguided Fractionation of Local Plants against Matrix Metalloproteinase9 and Its Cytotoxicity against Breast Cancer Cell Models: In Silico and In Vitro Study (Part II).

In our previous work, the partitions (1 mg/mL) of Ageratum conyzoides (AC) aerial parts and Ixora coccinea (IC) leaves showed inhibitions of 94% and 96%, respectively, whereas their fractions showed IC50 43 and 116 µg/mL, respectively, toward Matrix Metalloproteinase9 (MMP9), an enzyme that catalyzes a proteolysis of extracellular matrix. In this present study, we performed IC50 determinations for AC n-hexane, IC n-hexane, and IC ethylacetate partitions, followed by the cytotoxicity study of individual partitions against MDA-MB-231, 4T1, T47D, MCF7, and Vero cell lines. Successive fractionations from AC n-hexane and IC ethylacetate partitions led to the isolation of two compounds, oxytetracycline (OTC) and dioctyl phthalate (DOP). The result showed that AC n-hexane, IC n-hexane, and IC ethylacetate partitions inhibit MMP9 with their respective IC50 as follows: 246.1 µg/mL, 5.66 µg/mL, and 2.75 × 10-2 µg/mL. Toward MDA-MB-231, 4T1, T47D, and MCF7, AC n-hexane demonstrated IC50 2.05, 265, 109.70, and 2.11 µg/mL, respectively, whereas IC ethylacetate showed IC50 1.92, 57.5, 371.5, and 2.01 µg/mL, respectively. The inhibitions toward MMP9 by OTC were indicated by its IC50 18.69 µM, whereas DOP was inactive. A molecular docking study suggested that OTC prefers to bind to PEX9 rather than its catalytic domain. Against 4T1, OTC showed inhibition with IC50 414.20 µM. In conclusion, this study furtherly supports the previous finding that AC and IC are two herbals with potential to be developed as triple-negative anti-breast cancer agents.

Hesperidin as a preventive resistance agent in MCF-7 breast cancer cells line resistance to doxorubicin.

OBJECTIVE: To evaluate of hesperidin to overcome resistance of doxorubicin in MCF-7 resistant doxorubicin cells (MCF-7/Dox) in cytotoxicity apoptosis and P-glycoprotein (Pgp) expression in combination with doxorubicin. METHODS: The cytotoxic properties, 50% inhibition concentration (IC50) and its combination with doxorubicin in MCF-7 cell lines resistant to doxorubicin (MCF-7/Dox) cells were determined using MTT assay. Apoptosis induction was examined by double staining assay using ethidium bromide-acridine orange. Immunocytochemistry assay was performed to determine the level and localization of Pgp. RESULTS: Single treatment of hesperidin showed cytotoxic activity on MCF-7/Dox cells with IC50 value of 11 µmol/L. Thus, combination treatment from hesperidin and doxorubicin showed addictive and antagonist effect (CI>1.0). Hesperidin did not increase the apoptotic induction, but decreased the Pgp expressions level when combined with doxorubicin in low concentration. CONCLUSIONS: Hesperidin has cytotoxic effect on MCF-7/Dox cells with IC50 of 11 µmol/L. Hesperidin did not increased the apoptotic induction combined with doxorubicin. Co-chemotherapy application of doxorubicin and hesperidin on MCF-7/Dox cells showed synergism effect through inhibition of Pgp expression.

Hesperidin as a preventive resistance agent in MCF-7 breast cancer cells line resistance to doxorubicin

Objective:To evaluate of hesperidin to overcome resistance of doxorubicin in MCF-7 resistant doxorubicin cells (MCF-7/Dox) in cytotoxicity apoptosis and P-glycoprotein (Pgp) expression in combination with doxorubicin. Methods:The cytotoxic properties, 50%inhibition concentration (IC50) and its combination with doxorubicin in MCF-7 cell lines resistant to doxorubicin (MCF-7/Dox) cells were determined using MTT assay. Apoptosis induction was examined by double staining assay using ethidium bromide-acridine orange. Immunocytochemistry assay was performed to determine the level and localization of Pgp. Results: Single treatment of hesperidin showed cytotoxic activity on MCF-7/Dox cells with IC50 value of 11 μmol/L. Thus, combination treatment from hesperidin and doxorubicin showed addictive and antagonist effect (CI>1.0). Hesperidin did not increase the apoptotic induction, but decreased the Pgp expressions level when combined with doxorubicin in low concentration. Conclusions: Hesperidin has cytotoxic effect on MCF-7/Dox cells with IC50 of 11 μmol/L. Hesperidin did not increased the apoptotic induction combined with doxorubicin. Co-chemotherapy application of doxorubicin and hesperidin on MCF-7/Dox cells showed synergism effect through inhibition of Pgp expression.