Evaluation of anti-oxidant and antimelanogenic effects of the essential oil and extracts of Rosa × damascena in B16F10 murine melanoma cell line.

Objectives: Rosa × damascena Herrm. belonging to the Rosaceae family has demonstrated anti-inflammatory and anti-oxidant effects previously. Excessive production of free radicals and activation of tyrosinase enzyme caused by UV induces excessive concentration of melanin pigment and skin spots in the long term. Therefore, finding natural sources with anti-oxidant and antityrosinase effects helps to regulate the melanogenesis process. In the current research, we investigated the antimelanogenic, anti-oxidant, and anti-tyrosinase effects of its essential oil, methanol extract (MeOH), and different fractions including n-hexane, dichloromethane (CH2Cl2), n-butanol (BuOH), ethyl acetate (EtOAc), and H2O of R. × damascena in B16F10 cell line. Materials and Methods: For this purpose, impacts of extracts and essential oil of R. × damascena were investigated on cell viability, cellular tyrosinase, melanin content, mushroom tyrosinase, reactive oxygen species (ROS) production, as well as the amount of tyrosinase protein in the B16F10 murine melanoma cell line. Results: Essential oil, MeOH, and different fractions of R. × damascena were not cytotoxic on B16F10 cells. However, they had significant reducing effects on mushroom tyrosinase activity, melanin content, and ROS production. Also, there is a significant decrease in tyrosinase protein levels at 200 µg/ml but not at other concentrations. Conclusion: Therefore, the essential oil, MeOH, and different fractions of R. × damascena had promising antimelanogenic activity via repression of mushroom tyrosinase activity and ROS production.

The Cytotoxic Activity of Dammarane-Type Triterpenoids Isolated from the Stem Bark of Aglaia cucullata (Meliaceae).

The Aglaia genus, a member of the Meliaceae family, is generally recognized to include a number of secondary metabolite compounds with diverse structures and biological activities, including triterpenoids. Among the members of this genus, Aglaia cucullata has been reported to have unique properties and thrives exclusively in mangrove ecosystems. This plant is also known to contain various metabolites, such as flavaglines, bisamides, and diterpenoids, but there are limited reports on the isolation of triterpenoid compounds from its stem bark. Therefore, this research attempted to isolate and elucidate seven triterpenoids belonging to dammarane-type (1-7) from the stem bark of Aglaia cucullata. The isolated compounds included 20S,24S-epoxy-3α,25-dihydroxy-dammarane (1), dammaradienone (2), 20S-hydroxy-dammar-24-en-3-on (3), eichlerianic acid (4), (20S,24RS)-23,24-epoxy-24-methoxy-25,26,27-tris-nor dammar-3-one (5), 3α-acetyl-cabraleahydroxy lactone (6), and 3α-acetyl-20S,24S-epoxy-3α,25-dihydroxydammarane (7). Employing spectroscopic techniques, the chemical structures of the triterpenoids were identified using FTIR, NMR, and HRESITOF-MS. The cytotoxic activity of compounds 1-7 was tested with the PrestoBlue cell viability reagent against MCF-7 breast cancer, B16-F10 melanoma, and CV-1 normal kidney fibroblast cell lines. The results displayed that compound 5 had the highest level of bioactivity compared to the others. Furthermore, the IC50 values obtained were more than 100 µM, indicating the low potential of natural dammarane-type triterpenoids as anticancer agents. These findings provided opportunities for further studies aiming to increase their cytotoxic activities through semi-synthetic methods.

Dammarane-Type Triterpenoid from the Stem Bark of Aglaia elliptica (Meliaceae) and Its Cytotoxic Activities.

Two new dammarane-type triterpenoid fatty acid ester derivatives, 3ß-oleate-20S-hydroxydammar-24-en (1) and 3ß-oleate-20S,24S-epoxy-25-hydroxydammarane (2) with a known dammarane-type triterpenoid compound, such as 20S-hydroxydammar-24-en-3-on (3), were isolated from the stem bark of Aglaiaelliptica (C.DC.) Blume. The chemical structures were determined by spectroscopic methods, including FTIR, NMR (one and two-dimensional), and HRESITOF-MS analysis, as well as chemical derivatization and comparison with previous literature. Furthermore, the synthetic analog resulting from transesterification of 1 and 2 also obtained 3ß,20S-dihydroxy-dammar-24-en (4) and 20S,24S-epoxy-3ß,25-dihydroxydammarane (5), respectively. The cytotoxic effect of all isolated and synthetic analog compounds was evaluated using PrestoBlue reagent against MCF-7 breast cancer cell and B16-F10 melanoma cell lines. The 20S-hydroxydammar-24-en-3-on (3) showed the strongest activity against MCF-7 breast cancer and B16-F10 melanoma cell, indicating that the ketone group at C-3 in 3 plays an essential role in the cytotoxicity of dammarane-type triterpenoid. On the other hand, compounds 1 and 2 had very weak cytotoxic activity against the two cell lines, indicating the presence of fatty acid, significantly decreasing cytotoxic activity. This showed the significance of the discovery to investigate the essential structural feature in dammarane-type triterpenoid, specifically for the future development of anticancer drugs.

Synthesis of cinnamic acid ester derivatives with antiproliferative and antimetastatic activities on murine melanoma cells.

Melanoma is the most aggressive skin cancer, and its incidence has continued to rise during the past decades. Conventional treatments present severe side effects in cancer patients, and melanoma can be refractory to commonly used anticancer drugs, which justify the efforts to find new potential anti-melanoma drugs. An alternative to promote the discovery of new pharmacological substances would be modifying chemical groups from a bioactive compound. Here we describe the synthesis of seventeen compounds derived from cinnamic acid and their bioactivity evaluation against melanoma cells. The compound phenyl 2,3-dibromo-3-phenylpropanoate (3q) was the most effective against murine B16-F10 cells, as observed in cytotoxicity and cell migration assays. Simultaneously, this compound showed low cytotoxic activity on non-tumor cells. At the highest concentration, the compound 3q was able to trigger apoptosis, whereas, at lower concentrations, it affected the cell cycle and melanoma cell proliferation. Furthermore, cinnamate 3q impaired cell invasion, adhesion, colonization, and actin polymerization. In conclusion, these results highlight the antiproliferative and antimetastatic potential of cinnamic acid derivatives on melanoma.

Antiproliferative activity of aqueous and polyphenol-rich extracts of Larrea divaricata Cav. on a melanoma cell line.

Most of the deaths from skin cancer are caused by melanoma, a malignancy in which STAT3 plays a crucial role. The inhibition of STAT3 is considered a potential target to induce cell death, tumor regression and metastasis inhibition. The objective of this work was to evaluate the activity of the aqueous extract of Larrea divaricata (Aq), a fraction rich in polyphenols (EA),and the isolated compound quercetin-3-methyl ether (Q3ME) on B16F10 melanoma cells. The effects of Aq, EA and Q3ME were assessed on B16F10 cells by determining the proliferation, viability, apoptosis induction and the expression and phosphorylation of STAT3. The phytochemical composition of the extracts was determined by High Performance Liquid Chromatography. Aq, EA and Q3ME presented antiproliferative activity on B6F10 cells through p-STAT3 inhibition and early and late apoptosis induction (EC50 EA= ≤0.1 µg/ml; Aq= 316 ± 30 µg/ml; Q3ME= <0.1 µg/ml). L. divaricata could be considered for the development of adjuvant phytotherapies in melanoma treatment.

Garcinol-loaded novel cationic nanoliposomes: in vitro and in vivo study against B16F10 melanoma tumor model.

Aim: Garcinol (GAR)-loaded cationic nanoliposomes were developed to achieve potential antitumor efficacy on B16F10 melanoma cells in vitro and in vivo. Materials & methods: Two different phospholipids namely, distearoyl phosphatidylcholine (DSPC) and dipalmitoyl phosphatidylcholine (DPPC) were used in formulation to elucidate the difference in cellular uptake, cytotoxicity, in vivo tumor uptake (by scintigraphic imaging after technetium-99m radiolabeling) and therapeutic efficacy. Results: Different in vitro protocols, for example, MTT assay, apoptosis study, gene expression analysis, chromatin condensation and cytoskeleton breakdown analysis in B16F10 cell lines as well as scintigraphic analysis and tumor inhibition studies (B16F10 tumor xenograft model) revealed superiority of GAR-DPPC than GAR-DSPC and free GAR in melanoma prevention. Conclusion: Cationic nanoliposomal formulations could be a future medication for skin cancer treatment.

Evaluation of Breast Cancer and Melanoma Metastasis in Syngeneic Mouse Models.

Syngeneic mouse models enable us to study the interaction of the immune system with tumor cells during tumor development and metastasis. It further allows us to evaluate the efficacy of immunotherapies on the different stages of tumor growth and the metastatic process. Here we describe two syngeneic mouse models, a murine mammary carcinoma and a murine melanoma model, that are used to study metastasis. Metastases occur spontaneously in the murine mammary carcinoma. The presence and number of foci are evaluated by culturing the lung tissue in a colony formation assay. For the murine melanoma model, tumor cells are injected intravenously, and metastatic burden is analyzed by counting of metastatic lesions.

Inhibitory properties of aromatic thiosemicarbazones on mushroom tyrosinase: Synthesis, kinetic studies, molecular docking and effectiveness in melanogenesis inhibition.

The group of 19 thiosemicarbazones (TSCs) were synthesized and its inhibitory activity toward mushroom tyrosinase and ability to inhibition of melanogenesis in B16 cells were investigated. Moreover, molecular docking of these compounds to the active site of the enzyme was performed. The obtained results allowed to make the structure-activity relationship (SAR) analysis. Kinetic studies revealed that TSCs 1, 2, 11 and 18 have better inhibitory properties than kojic acid, a reference compound, with the best inhibitory constant (Ki) value of 0.38 µM for TSC 2. According to SAR analysis, the smaller and less branched molecules exhibit higher affinity to the enzyme. Melanin production in B16 cells was inhibited by all investigated compounds at micromolar level. Most of compounds studied in this work can be considered as potent inhibitors of tyrosinase and melanogenesis. They may have broad application in food preservatives and cosmetics. Combined results of molecular docking and SAR analysis can be helpful in designing novel tyrosinase inhibitors of desired properties.

Anti-melanogenic activity of Viola odorata different extracts on B16F10 murine melanoma cells.

OBJECTIVES: In previous studies, antioxidant activity of Viola odorata L. has been demonstrated. In this study, we have investigated the anti-melanogenic effect of extract and fractions of the plant in B16F10 cell line. MATERIALS AND METHODS: Impact of different increasing concentrations of extract and fractions of V. odorata was evaluated on cell viability, cellular tyrosinase, melanin content and mushroom tyrosinase as well as ROS production in B16F10 murine melanoma cell line. RESULTS: Viola odorata had no cytotoxicity on B16F10 cells compared to control group. Kojic acid as positive control had significant decreasing effects on cellular and mushroom tyrosinase activity, melanin content and ROS production (P<0.001, for all cases). V. odorata (1-20 µg/ml) decreased all measured parameters including cellular tyrosinase and melanin content as well as ROS production and among all extract and fractions ethyl acetate fraction had the best effect (P<0.05). CONCLUSION: Viola odorata had promising anti-melanogenic activity through inhibition of cellular tyrosinase activity and ROS production as well as melanin content. More basic and clinical studies need to aver its impact.

Differential biological effects of dehydroepiandrosterone (DHEA) between mouse (B16F10) and human melanoma (BLM) cell lines.

Dehydroepiandrosterone (DHEA) is a weak androgen and had been shown to have anti-cancer, anti-adipogenic and anti-inflammatory effects on mouse and other rodent models, but not on humans, suggesting a systemic level difference between mouse and human. Our previous study on DHEA biological functions involving a variety of cell lines, suggested that the functional differences between mouse and human existed even at the cellular level. Hence, using mouse and human melanoma cell models, in-vitro effects of DHEA on cell growth, mechanism of cell death and mechanism of DHEA action were studied. Results indicated a differential biological effects of DHEA between mouse and human melanoma cell lines. These in-vitro studies also suggested that the differential biological effects observed between these two cell lines could be due to the difference in the way DHEA was processed or metabolized inside the cell.

In vivo antitumor potential of extracts from different parts of Bauhinia variegata linn. Against b16f10 melanoma tumour model in c57bl/6 mice

Background: Melanoma is a metastatic type of skin cancer that is difficult to treat and the majority of efforts are directed to the design of new drugs. Medicinal Plants have been the primary source of medicines since life on earth; more than 50% of existing cancer treatments is derived from plants. Bauhinia variegata is well-known medicinal plant used from the ancient era to till date for their medicinal values. Scientific literatures have not documented any evidence of the antitumour potential of Bauhinia variegata against B16F10 melanoma tumor model in C57BL mice. The present investigation was undertaken to explore the antitumour activity of Leaf, stem bark and flower extract of Bauhinia variegata against B16F10 melanoma tumour model in C57BL mice. Methods: Hydro-methanolic extract prepared from the leaf, stem bark and flower of Bauhinia variegata were assessed for their antitumor activity. The extracts at doses of 500 and 750 mg/kg b.wt. were given orally along with cyclophosphamide (chemotherapeutic drug) for 40 days for exploring antitumor activity against melanoma tumor (B16F10) in C57BL mice. Inhibition of tumor growth, increase in survival time of animal with treatment, histopathological studies and antioxidant parameter were determined. Results: The Present investigation showed significant effect of the B. variegata L. in preventing melanoma tumor by B16F10 cell line in C57BL/6 mice. As compared with the tumour control group, the remarkable results especially in the group which received B. variegata extract and cyclophosphamide together were obtained for all of the measured parameters. Dose dependent response was observed in tumor volume, inhibition rate, life span time and antioxidant parameter of extracts. Combination treatment of cyclophosphamide and B. variegata extracts showed more pronounced effect. Conclusions: These findings suggest that B. variegata hydromethanolic extract may contain bioactive compounds of potential therapeutic significance which are relatively safe from toxic effects, and can compromise the medicinal use of this plant in folk medicine (US)

Interaction between omega 3 PUFA and UVB radiation: Photoprotective effect in normal and tumoral murine melanocytes?

Omega 3 polyunsaturated fatty acids (omega 3 PUFA) are attracting a growing interest as potential adjuvants for cancer prevention and treatment. There is evidence about photoprotection in normal cells, but few previous studies have evaluated it in tumoral cells. Therefore, this study investigated the effect of α-linolenic acid (ALA) in normal murine melanocytic cells (Melan-a) and in tumoral murine melanocytic cells (B16F10) exposed to UVB radiation. Our results showed that ALA exhibited an antiproliferative effect in B16F10 cells, and had minimal effect in Melan-a cells, as demonstrated by MTT assay. On the other hand, the combination of ALA (7.5µM) and UVB (0.01J/cm2) showed a protective effect for both cell lines, Melan-a and B16F10. ALA and UVB combined or UVB alone induced an accumulation of cell lines at the S/G2/M phase. In addition, the combination of ALA and UVB, and UVB alone, both induced cell death in 24h; and in 48h, ALA attenuated this effect in both cells. Further to these findings, it was demonstrated that ALA did not alter ROS levels in both cells exposed to UVB radiation. The effect of an omega 6 PUFA, linoleic acid, under the same conditions of ALA were tested. It was not protective in either cell line. Therefore, our results can be very important since it was shown another role to an omega 3 PUFA as a photoprotective agent in a melanoma cell.

Synthesis and biological evaluation of novel 2-imino-4-thiazolidinone derivatives as potent anti-cancer agents.

A new series of 2-imino-4-thiazolidinone derivatives (7a-7t) has been synthesised and screened for their cytotoxicity against three cancer cell lines (B16F10, A549, PANC-1) and normal cell line (CHO). Among the compounds tested, compounds 7k, 7m, 7n showed potent cytotoxicity against B16F10 cell line with IC50 between 3.4 and 7µM. Interestingly these three compounds are non toxic to non cancerous CHO cells and induced apoptosis in B16F10 cells observed by DNA damage analysis through PI/Hoechst double staining method. Compounds 7k and 7n induced G0/G1 cell cycle arrest while compound 7m induced G2/M cell cycle arrest in B16F10 cells which confirms that these compounds have role in cancer cell cycle regulation. Additionally, compound 7m showed generation of intracellular reactive oxygen species (ROS) in B16F10 cells that may contribute in the cell cycle arrest whereas compounds 7k and 7n show anti-cancer activity through independent of ROS formation. Induction of apoptosis, cell cycle arrest in B16F10 cells are found to be the anti-cancer mechanism of these three compounds. The results all together demonstrate the potent cytotoxic nature of these compounds in cancer cells could be considered as new class of chemotherapeutic agents in near future.

Ursolic Acid Loaded PLGA Nanoparticles: in vitro and in vivo Evaluation to Explore Tumor Targeting Ability on B16F10 Melanoma Cell Lines.

PURPOSE: Ursolic acid (UA), a pentacyclic triterpenoid extracted from plants, shows promising inhibitory effect in different tumor bearing cell lines. In the present study we fabricated UA loaded PLGA nanoparticles (UA-NPs) as the drug carrier and thoroughly evaluated in vitro and in vivo the differential tumor targeting effects of UA and UA-NPs in B16F10 melanoma cells. METHODS: Ursolic acid loaded PLGA nanoparticles were prepared by emulsion solvent evaporation technique and evaluated for particle size, polydispersity, zeta potential and drug release potency. MTT assay as well as flow cytometric and confocal microscopic analyses were done in B16F10 mouse melanoma cell lines. Formulations were labeled with technetium-99m to evaluate the biodistribution and perform scintigraphic imaging studies following intravenous administration in tumor bearing mice model. RESULTS: Single emulsification technique produced smooth spherical nanoparticles of small size with relatively narrow size distribution (154 ± 4.56 nm). On B16F10 cell line, the formulation showed higher cytotoxicity compared to the free drug due to increased in vitro cellular uptake. The formulation was successfully radiolabeled and remained substantially (>90%) stable when incubated (37°C, 6 h) separately in normal saline or freshly collected rat serum or histidine solution. The radiolabeled UA-NPs exhibited slower blood clearance and comparatively high uptake in tumor region as evidenced by biodistribution and scintigraphic studies. CONCLUSIONS: The in vitro and in vivo studies have proved the tumor targeting potential of UA-NPs in B16F10 melanoma cell lines.

Resveratrol-loaded nanocapsules inhibit murine melanoma tumor growth.

In this study, resveratrol-loaded nanocapsules were developed and its antitumor activity tested on a melanoma mice model. These nanocapsules were spherically-shaped and presented suitable size, negative charge and high encapsulation efficiency for their use as a modified-release system of resveratrol. Nanoencapsulation leads to the drug amorphization. Resveratrol-loaded nanoparticles reduced cell viability of murine melanoma cells. There was a decrease in tumor volume, an increase in the necrotic area and inflammatory infiltrate of melanoma when resveratrol-loaded nanocapsules were compared to free resveratrol in treated mice. Nanoencapsulation of resveratrol also prevented metastasis and pulmonary hemorrhage. This modified-release technology containing resveratrol can be used as a feasible approach in order to inhibit murine melanoma tumor growth.