B16F10 Murine Melanoma Cell Line Research Articles

Abstract 3032: Generation of a FOXC2-deficient melanoma cell line using CRISPR-Cas9 gene editing

Abstract Melanoma, a cancer derived from pigment-producing melanocytes of the skin, is a highly aggressive cancer characterized by rapid growth and metastasis. FOXC2 is a forkhead box transcription factor family member that is often dysregulated in various cancers, including melanoma, where its expression correlates with metastatic progression and drug resistance. To gain insight into the oncogenic activity of FOXC2 in melanoma, we have employed CRISPR-Cas9 gene editing technology to generate a novel FOXC2-deficient B16-F10 murine melanoma cell line. This model will be a useful tool for studying the tumor-promoting functions of FOXC2. Citation Format: Devanand M. Clark, Patrick D. Conde, Kristian M. Hargadon. Generation of a FOXC2-deficient melanoma cell line using CRISPR-Cas9 gene editing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3032.

The Antimelanogenic Activity of the Extract of Heukharang Lettuce (Lactuca sativa L.) Leaf

Heukharang (Lactuca sativa L.), the artificially developed lettuce cultivar for enhancement of lactucin content, exhibits antioxidant activities and sleep-promoting effects. However, potential of Heukharang as a raw material for the skin-whitening agent has not been investigated yet. This study evaluated the effects of Heukharang extract (HHE) on α-melanocyte-stimulating hormone (α-MSH)-induced melanogenesis in vitro and in vivo models. Our findings revealed that HHE (25–100 μg/mL) effectively inhibited α-MSH-induced melanin synthesis in both 2D and 3D cell culture environments without compromising the viability of the B16F10 murine melanoma cell line. Furthermore, HHE suppressed intracellular tyrosinase activity, along with both gene and protein expression of tyrosinase (TYR) and tyrosinase-related protein (TRP)-1 and TRP-2. HHE downregulated the mRNA level of microphthalmia-associated transcription factor (MITF) and the phosphorylation of cAMP response element-binding (CREB) protein. The antimelanogenic activity of HHE (50 and 100 μg/mL) was also confirmed using a zebrafish embryo model, highlighting its efficacy in inhibiting α-MSH-induced melanogenesis through the downregulating CREB/MITF/TYR signaling pathways. Moreover, our study demonstrated that a corresponding amount of lactucin (5–20 μM) to the HHE inhibited and regulated melanogenesis. These results collectively suggest that HHE, enriched with lactucin, holds promise as a potential skin-whitening agent for nutricosmetic industries.

Photodynamic Therapy Directed to Melanoma Skin Cancer by Thermosensitive Hydrogel Containing Chlorophyll A.

Melanoma, a severe form of skin cancer intricately linked to genetic and environmental factors, is predicted to reach 100,000 new cases worldwide by 2040, underscoring the need for effective and safe treatment options. In this study, we assessed the efficacy of a photosensitizer called Chlorophyll A (Chl-A) incorporated into hydrogels (HGs) made of chitosan (CS) and poloxamer 407 (P407) for Photodynamic Therapy (PDT) against the murine melanoma cell line B16-F10. The HG was evaluated through various tests, including rheological studies, SEM, and ATR-FTIR, along with cell viability assays. The CS- and P407-based HGs effectively released Chl-A and possessed the necessary properties for topical application. The photodynamic activity of the HG containing Chl-A was evaluated in vitro, demonstrating high therapeutic potential, with an IC50 of 25.99 µM-an appealing result when compared to studies in the literature reporting an IC50 of 173.8 µM for cisplatin, used as a positive control drug. The developed formulation of CS and P407-based HG, serving as a thermosensitive system for topical applications, successfully controlled the release of Chl-A. In vitro cell studies associated with PDT exhibited potential against the melanoma cell line.

Discovery of amphotericin B, an antifungal drug as tyrosinase inhibitor with potent anti-melanogenic activity

Tyrosinase, a rate-limiting enzyme for melanin production, has been the most efficient target for the development of depigmenting agents. Although hydroquinone, kojic acid, and arbutin are the most well-known tyrosinase inhibitors, their adverse effects are inevitable. In the present study, an in silico drug repositioning combined with experimental validation was performed to search for novel potent tyrosinase inhibitors. Docking-based virtual screening results revealed that, among the 3210 FDA-approved drugs available in the ZINC database, amphotericin B, an antifungal drug exhibited the highest binding efficiency against human tyrosinase. Results from tyrosinase inhibition assay demonstrated that amphotericin B could inhibit the activity of mushroom and cellular tyrosinases, especially from MNT-1 human melanoma cells. Molecular modeling results revealed that amphotericin B/human tyrosinase complex exhibited high stability in an aqueous environment. Melanin assay results demonstrated that amphotericin B significantly suppressed melanin production in α-MSH-induced B16F10 murine melanoma and MNT-1 human melanoma cell lines better than the known inhibitor, kojic acid. Mechanistically, amphotericin B treatment significantly activated ERK and Akt signaling pathways, resulting in the decreased expression of MITF and tyrosinase. The obtained results may pursue pre-clinical and clinical studies to examine the possibility of using amphotericin B as an alternative treatment for hyperpigmentation disorders.

Effects of redox modulation on quiescin/sulfhydryl oxidase activity of melanoma cells.

Secreted quiescin/sulfhydryl oxidase (QSOX) is overexpressed in many tumor cell lines, including melanoma, and is usually associated with a pro-invasive phenotype. Our previous work described that B16-F10 cells enter in a quiescent state as a protective mechanism against damage generated by reactive oxygen species (ROS) during melanogenesis stimulation. Ourpresent results show that QSOX activity was two-fold higher in cells with stimulated melanogenesis when compared to control cells. Considering that glutathione (GSH) is one of the main factor responsible for controlling redox homeostasis in cells, this work also aimed to investigate the relationship between QSOX activity, GSH levels and melanogenesis stimulation in B16-F10 murine melanoma cell line. The redox homeostasis was impaired by treating cells with GSH in excess or depleting its intracellular levels through BSO treatment. Interestingly, GSH-depleted cells without stimulation of melanogenesis kept high levels of viability, suggesting a possible adaptive mechanism of survival even under low GSH levels. They also showed lower extracellular activity of QSOX, and higher QSOX intracellular immunostaining, suggesting that this enzyme was less excreted from cells and corroborating with a diminished extracellular QSOX activity. On the other hand, cells under melanogenesis stimulation showed a lower GSH/GSSG ratio (8:1) in comparison with control (non-stimulated) cells (20:1), indicating a pro-oxidative state after stimulation. This was accompanied by decreased cell viability after GSH-depletion, no alterations in QSOX extracellular activity, but higher QSOX nucleic immunostaining. We suggest that melanogenesis stimulation and redox impairment caused by GSH-depletion enhanced the oxidative stress in these cells, contributing to additional alterations of its metabolic adaptive response.

Bioengineered dual fluorescent carbon nano dots from Indian long pepper leaves for multifaceted environmental and health utilities.

In this article, we present the synthesis of Piper longum leaves-derived ethanolic carbon dots (PLECDs) using the most simplistic environmentally friendly solvothermal carbonization method. The PLECDs fluoresced pink color with maximum emission at 670nm at 397nm excitation. Additionally, the dried PLECDs dissolved in water showed green fluorescence with higher emission at 452nm at 370nm excitation. The UV spectra showed peaks in the UV region (271.25nm and 320.79nm) and a noticeable tail in the visible region, signifying the efficient synthesis of nano-sized carbon particles and the Mie scattering effect. Various functional groups (-OH, -N-H, -C-H, -C = C, -C-N, and -C-O) were identified using Fourier transform infrared spectroscopy (FTIR). Its nanocrystalline property was revealed by the sharp peaks in the X-ray diffraction (XRD). High-resolution transmission electron microscopy (HRTEM) photomicrograph displayed a roughly spherical structure with a mean size of 2.835nm. The energy dispersive X-ray (EDAX) and X-ray photoelectron spectroscopy (XPS) revealed the elemental abundance of C, O, and N. The high-performance thin-layer chromatography (HPTLC) fingerprint of PLECDs showed an altered pattern than its precursor (Piper longum leaves ethanolic extract or PLLEE). The PLECDs sensed Cu2+ selectively with a limit of detection (LOD) and limit of quantification (LOQ) of 0.063μM and 0.193μM, respectively. It showed excellent cytotoxicity toward MDA-MB-231 (human breast cancer), SiHa (human cervical carcinoma), and B16F10 (murine melanoma) cell lines with excellent in vitro bioimaging outcomes. It also has free radical scavenging activity. The PLECDs also showed outstanding bacterial biocompatibility, pH-dependent fluorescence stability, photostability, physicochemical stability, and thermal stability.

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

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. 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. 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. Therefore, the essential oil, MeOH, and different fractions of R. × damascena had promising antimelanogenic activity via repression of mushroom tyrosinase activity and ROS production.

Slow-cycling murine melanoma cells display plasticity and enhanced tumorigenicity in syngeneic transplantation assay

Slow-cycling cancer cells (SCC) contribute to the aggressiveness of many cancers, and their invasiveness and chemoresistance pose a great therapeutic challenge. However, in melanoma, their tumor-initiating abilities are not fully understood. In this study, we used the syngeneic transplantation assay to investigate the tumor-initiating properties of melanoma SCC in the physiologically relevant in vivo settings. For this we used B16-F10 murine melanoma cell line where we identified a small fraction of SCC. We found that, unlike human melanoma, the murine melanoma SCC were not marked by the high expression of the epigenetic enzyme Jarid1b. At the same time, their slow-cycling phenotype was a temporary state, similar to what was described in human melanoma. Progeny of SCC had slightly increased doxorubicin resistance and altered expression of melanogenesis genes, independent of the expression of cancer stem cell markers. Single-cell expansion of SCC revealed delayed growth and reduced clone formation when compared to non-SCC, which was further confirmed by an in vitro limiting dilution assay. Finally, syngeneic transplantation of 10 cells in vivo established that SCC were able to initiate growth in primary recipients and continue growth in the serial transplantation assay, suggesting their self-renewal nature. Together, our study highlights the high plasticity and tumorigenicity of murine melanoma SCC and suggests their role in melanoma aggressiveness.

Contact effects by mitochondria: biological destruction of cultivated B16 – F10 melanoma cells

In recent years therapeutic effects produced by mitochondria, transplanted to pathogenic regions in an organism, which demonstrate their high migration activity and tropicity with respect to filling of energetic vacuum, have been inspiring a renewed interest in the scientific community. In this context, this raises the question of the source of metabolically active mitochondria and their histological compatibility required for the mitochondrial transplantation. Aim. The aim of our research work is to study characteristics/ quality of mitochondria from cells of the liver and the heart in rats, which in the in vitro system have produced their impact on different biological features of the B16-F10 murine melanoma cell culture. Materials and methods. In our research work we have used cells of the B16-F10 murine melanoma cell line culture. In the framework of the study, an experiment with mitochondria harvested from the liver and the heart of a rat has been conducted. Mitochondria have been isolated using differential centrifugation with a high-speed refrigerated centrifuge. With the B16-F10 culture cells, we have designed the following variants of our experiments: 1) use of cardiac mitochondria (1 mg/mL, in terms of total protein); 2) use of cardiac mitochondria (1 mg/mL) + succinic acid (10-4%); 3) mitochondria of the liver (1 mg/mL, in terms of total protein); 4) use of mitochondria of the liver (1 mg/mL) + succinic acid (10-4%); 5) use of succinic acid (10-4%) solely; 6) the reference specimen with no use of mitochondria and the above agents. An assessment of the impact made by mitochondria on the migration of the B16-F10 cells has been performed with the scratch wound healing test. For the purpose of an analysis of the effect produced by mitochondria on the energetic metabolism of the B16-F10 cell culture we have measured main parameters of the cell respiration and glycolysis in stress tests with adding some toxic chemicals. The rate of the cellular respiration has been assessed by measuring the amount of oxygen taken in (oxygen consumption rate, OCR), and the glycolysis level has been evaluated by the extracellular acidification rate (ECAR). Results. Adding cardiac and hepatic mitochondria to the cultivated B16-F10 cells has produced a pronounced cytopathic effect, which has become more remarkable upon expiration of two days of the cell cultivation and which has consisted in cytoplasm granulation and partial detaching of the cells. Introducing mitochondria of the heart to the cultivated B16-F10 cells has induced a considerable decrease both in the background-related and the maximum level of oxygen consumption by the B16-F10 cells as against the reference samples without adding of mitochondria. Adding the cardiac mitochondria has led to a statistically significant decrease in the base level of ECAR by 14,36 mpH/min (t = 3,12, df = 10) as compared with the reference values. Introducing hepatic mitochondria has also resulted in a reduction of the average value of ECAR as against the background by 4,8 mpH/min. Conclusion. Metabolically active mitochondria are capable of reformatting energetic fluxes in tumor cells and change their cellular respiration that leads to the most effectively realized death of the cultivated B16-F10 tumor cells.

Echinochrome A Inhibits Melanogenesis in B16F10 Cells by Downregulating CREB Signaling.

Excessive increase in melanin pigment in the skin can be caused by a variety of environmental factors, including UV radiation, and can result in spots, freckles, and skin cancer. Therefore, it is important to develop functional whitening cosmetic reagents that regulate melanogenesis. In this study, we investigated the effects of echinochrome A (Ech A) on melanogenesis in the B16F10 murine melanoma cell line. We triggered B16F10 cells using α-MSH under Ech A treatment to observe melanin synthesis and analyze expression changes in melanogenesis-related enzymes (tyrosinase, tyrosinase-related protein 1 (TYRP1), and tyrosinase-related protein 2 (TYRP2)) at the mRNA and protein levels. Furthermore, we measured expression changes in the microphthalmia-associated transcription factor (MITF), CREB, and pCREB proteins. Melanin synthesis in the cells stimulated by α-MSH was significantly reduced by Ech A. The expression of the tyrosinase, TYRP1, and TYRP2 mRNA and proteins was significantly decreased by Ech A, as was that of the MITF, CREB, and pCREB proteins. These results show that Ech A suppresses melanin synthesis by regulating melanogenesis-related enzymes through the CREB signaling pathway and suggest the potential of Ech A as a functional agent to prevent pigmentation and promote skin whitening.

Antitumor Properties of Epitope-Specific Engineered Vaccine in Murine Model of Melanoma.

Finding new effective compounds of natural origin for composing anti-tumor vaccines is one of the main goals of antitumor research. Promising anti-cancer agents are the gastropodan hemocyanins–multimeric copper-containing glycoproteins used so far for therapy of different tumors. The properties of hemocyanins isolated from the marine snail Rapana thomasiana (RtH) and the terrestrial snail Helix aspersa (HaH) upon their use as carrier-proteins in conjugated vaccines, containing ganglioside mimotope GD3P4 peptide, were studied in the developed murine melanoma model. Murine melanoma cell line B16F10 was used for solid tumor establishment in C57BL/6 mice using various schemes of therapy. Protein engineering, flow cytometry, and cytotoxicity assays were also performed. The administration of the protein-engineered vaccines RtH-GD3P4 or HaH-GD3P4 under the three different regimens of therapy in the B16F10 murine melanoma model suppressed tumor growth, decreased tumor incidence, and prolonged the survival of treated animals. The immunization of experimental mice induced an infiltration of immunocompetent cells into the tumors and generated cytotoxic tumor-specific T cells in the spleen. The treatment also generates significantly higher levels of tumor-infiltrated M1 macrophages, compared to untreated tumor-bearing control mice. This study demonstrated a promising approach for cancer therapy having potential applications for cancer vaccine research.

Heme Oxygenase-1 Has a Greater Effect on Melanoma Stem Cell Properties Than the Expression of Melanoma-Initiating Cell Markers.

Melanoma-initiating cells (MICs) contribute to the tumorigenicity and heterogeneity of melanoma. MICs are identified by surface and functional markers and have been shown to display cancer stem cell (CSC) properties. However, the existence of MICs that follow the hierarchical CSC model has been questioned by studies showing that single unselected melanoma cells are highly tumorigenic in xenotransplantation assays. Herein, we characterize cells expressing MIC markers (CD20, CD24, CD133, Sca-1, ABCB1, ABCB5, ALDHhigh) in the B16-F10 murine melanoma cell line. We use flow cytometric phenotyping, single-cell sorting followed by in vitro clonogenic assays, and syngeneic in vivo serial transplantation assays to demonstrate that the expression of MIC markers does not select CSC-like cells in this cell line. Previously, our group showed that heme-degrading enzyme heme oxygenase-1 (HO-1) can be upregulated in melanoma and increase its aggressiveness. Here, we show that HO-1 activity is important for non-adherent growth of melanoma and HO-1 overexpression enhances the vasculogenic mimicry potential, which can be considered protumorigenic activity. However, HO-1 overexpression decreases clone formation in vitro and serial tumor initiation in vivo. Thus, HO-1 plays a dual role in melanoma, improving the progression of growing tumors but reducing the risk of melanoma initiation.

Biphasic effect of the dietary phytochemical linalool on angiogenesis and metastasis.

Cytotoxic chemotherapy dominates the field of cancer treatment. Consequently, anticancer phytochemicals are largely screened on the basis of their cytotoxicity towards cancer cells which are achieved at higher doses, leading to various toxic side effects. Some phytochemicals also showed pro-carcinogenic effects at certain doses. The concept of hormesis has taught us to look into biphasic responses of phytochemicals in a more systematic way. Interestingly, the monoterpenoid alcohol, linalool, also has been reported to display both anti-oxidant and pro-oxidant properties, which prompted us to explore a probable biphasic effect on cancer cells. Cytotoxicity of various concentrations of linalool (0.1-4mM) was tested on B16F10 murine melanoma cell line, and two sub-lethal concentrations (0.4 and 0.8mM) were selected for further experiments. 0.4mM linalool inhibited angiogenesis and metastasis, while 0.8mM increased them. Similarly, B16F10 cell migration, invasion, and epithelial-mesenchymal transition markers also showed inhibition and induction with lower and higher linalool concentrations, respectively. Chorioallantoic membrane assay, scratch wound assay, and Boyden's chamber were used to analyze angiogenesis and metastasis. Expression of molecular markers such as vascular endothelial growth factor (VEGF) and its receptor phosphorylated VEGF receptor II (p-VEGFRII or p-Flk-1), Hypoxia-inducible factor-1 α (HIF-1α), E-cadherin, and vimentin were detected using Western blot, ELISA, PCR, qPCR, and immunofluorescence. Finally, ChIP assay was performed to evaluate HIF-1α association with VEGF promoter. Interestingly, measurement of intracellular reactive oxygen species at the selected concentrations of linalool using DCFDA in a flow cytometer showed that the phytochemical induced significant amount of ROS at 0.8mM. This work sheds light on bimodal dose-response relationship exhibited by dietary phytochemicals like linalool, and it should be taken into consideration to elicit a desirable therapeutic effect.

In Vitro Anticancer Activity of Nanoformulated Mono‐ and Di‐nuclear Pt Compounds

Nanoformulations of mononuclear Pt complexes cis-PtCl2 (PPh3 )2 (1), [Pt(PPh3 )2 (L-Cys)] ⋅ H2 O (3, L-Cys=L-cysteinate), trans-PtCl2 (PPh2 PhNMe2 )2 (4; PPh2 PhNMe2 =4-(dimethylamine)triphenylphosphine), trans-PtI2 (PPh2 PhNMe2 )2 (5) and dinuclear Pt cluster Pt2 (μ-S)2 (PPh3 )4 (2) have comparable cytotoxicity to cisplatin against murine melanoma cell line B16F10. Masking of these discrete molecular entities within the hydrophobic core of Pluronic® F-127 significantly boosted their solubility and stability, ensuring efficient cellular uptake, giving in vitro IC50 values in the range of 0.87-11.23 μM. These results highlight the potential therapeutic value of Pt complexes featuring stable Pt-P bonds in nanocomposite formulations with biocompatible amphiphilic polymers.

Evaluation of the oncolytic property of recombinant Newcastle disease virus strain R2B in 4T1 and B16-F10 cells in-vitro

Recombinant Newcastle disease virus vectors have gained a lot of interest for its oncolytic virus therapy and cancer immune therapeutic properties due to its selective replication to high titers in cancer cells. The aim of this study was to find out the oncolytic effects of mesogenic recombinant NDV strain R2B-GFP on murine mammary tumor cell line 4T1 and murine melanoma cell line B16-F10. The anti-tumor effects of R2B-GFP virus were studied via expression of virus transgene GFP in cancer cells, evaluating its cytotoxicity and cell migration efficacies by MTT and wound healing assays respectively. In addition, the underlying apoptotic mechanism of R2B-GFP virus was estimated by TUNEL assay, colorimetric estimation of Caspase-3, 8 and 9 and the estimation of Bax to Bcl-2 ratio. The results showed a significant decrease in viability of both 4T1 and B16-F10 cells infected with R2B-GFP virus at 0.1 and 1 MOI. R2B-GFP virus could significantly induce apoptosis in the 4T1 and B16-F10 cells as compared to the uninfected control. Further, a flow cytometry analysis on apoptotic cells percentage and mitochondria membrane permeability test was also studied in R2B-GFP virus treated 4T1 and B16-F10 cell lines. The R2B-GFP virus caused an increase in loss of mitochondrial membrane permeability in both 4T1 and B16-F10 cells indicating the involvement of mitochondrial regulated cell death. Thus, the recombinant virus R2B-GFP virus proved to be a valid candidate for oncolytic viral therapy in 4T1 and B16-F10 cells.

Expression of MITF Gene in Melanogenesis with Psoralea corylifolia, Zingiber officinale, Terminalia chebula, Punica granatum and Eclipta alba Based on Poly-Herbal Formulation

Pigmentation is an important physical characteristic of human beings due to the presence of a biopolymer molecule called melanin. Melanin pigment in the human skin and several other tissues were due to the activation and inhibition of different genes at the downstream signaling pathway like Tyrosinase-related protein 1 (TYRP1) and Tyrosinase-related protein 2 (TYRP2) and Microphthalmia-associated transcription factor (MITF) genes. In this study, we tried to deliver the molecular approach of TYRP1, TYRP2, and MITF by activating kinases in the process of Melanogenesis by treating with Unani formulations on the B16F10 murine melanoma cell line. RNA isolation of B16F10 cells was performed followed by cDNA conversion and expression studies at mRNA levels by RT-PCR method. To explore further western blot was performed at the protein level for the significant results which we obtained with UNIM regimes in the expression studies. The bioactive ingredient of UNIM-004 and UNIM-005 of Methanolic extracts up-regulates the expression and protein levels of the MITF gene which leads to the synthesis of melanin pigment in human beings. Therefore from this study, it indicates that the application and administration of UNIM-004 and UNIM-005 of methanolic extract will result in activating melanin in B16F10 cells by increasing mRNA expression levels in the MITF gene when compared with TYRP1and TYRP2 genes followed by Western blot experiment and found to be producing melanocytes in the progression of Melanogenesis in a dose-dependent manner at 10 μg and 100 μg concerning for to GAPDH control gene at mRNA expression level in Vitiligo patients.

Monosubstituted Acetophenone Thiosemicarbazones as Potent Inhibitors of Tyrosinase: Synthesis, Inhibitory Studies, and Molecular Docking.

A set of 12 monosubstituted acetophenone thiosemicarbazone derivatives (TSCs) were synthesized and their inhibitory properties toward tyrosinase activity were tested. Moreover, their ability to inhibit melanogenesis in the B16F10 murine melanoma cell line was studied. In order to investigate the nature of interactions between the enzyme and the inhibitors, molecular docking to the active site was performed. TSCs 5, 6, 8, and 9 revealed a half maximal inhibitory concentration (IC50) below 1 µM. Compound 6 turned out to be the most potent tyrosinase inhibitor. All investigated compounds showed reversible inhibition of competitive or mixed type. The para-substituted TSCs had higher affinity for the enzyme as compared to their ortho- and meta-analogues. All investigated compounds inhibited melanin production in B16F10 cells at the micromolar level. Molecular docking showed that the sulfur atom of the thiourea moiety penetrates the active site and interacts with copper ions. The above outcomes might be helpful in the design of new tyrosinase inhibitors in the food and cosmetic industries.

Synthesis of Novel Cinnamides and a Bis Cinnamate Bearing 1,2,3-Triazole Functionalities with Antiproliferative and Antimetastatic Activities on Melanoma Cells

The present investigation describes the synthesis of novel cinnamides and a bis cinnamate bearing 1,2,3-triazole functionalities and investigation of their antiproliferative and antimetastatic effects on melanoma cells. The necessity for the development of new chemotherapeutic agents for melanoma treatment motivated this work. Sixteen derivatives were obtained with yields ranging from 23-81% and fully characterized by spectroscopic (1H and 13C nuclear magnetic resonance, infrared) and spectrometric high resolution mass spectrometry (HRMS) techniques. The derivatives were in vitro evaluated against B16-F10 murine melanoma cell line. The most effective compound (a bis cinnamate) (6b) reduced the melanoma cell viability, generated cell cycle arrest, and influenced the metastatic behavior of melanoma cells by decreasing migration, invasion, and colony formation. Based on these findings, it is believed that compound 6b may represent an interesting scaffold to be explored toward the development of new antimelanoma agents.

Green does not always mean go: A sulfated galactan from Codium isthmocladum green seaweed reduces melanoma metastasis through direct regulation of malignancy features

Melanoma is the most lethal form of skin cancer, with a worldwide increase in incidence. Despite the increased overall survival of metastatic melanoma patients given recent advances in targeted and immunotherapy, it still has a poor prognosis and available treatment options carry diverse severe side effects. Polysaccharides from seaweed have been shown to exert antitumor activities. Here we show in vitro and in vivo antitumor activities of a sulfated homogalactan (named 3G4S) from Codium isthmocladum seaweed in the B16-F10 murine melanoma cell line. 3G4S did not induce cytotoxicity or proliferation changes; however, it was able to reduce solid tumor growth and metastasis, while not inducing side effects in mice. B16-F10 cells traits related to the metastatic cascade were also impaired by 3G4S, reducing cell invasion, colony-forming capacity and membrane glycoconjugates. Therefore, 3G4S shows promising antitumor activities without the commonly associated drawbacks of cancer treatments and can be further explored.

Planar and tomographic imaging dosimetry comparing to develop a new radiopharmaceutical for melanoma therapy

This work aims to compare SPECT (Single Photon Emission Tomography Computed) and planar modalities as the most efficient methodology to perform dosimetry by molecular imaging. Twenty-one male C57BL6 mice induced with murine melanoma cell line B16-F10, administered 131I-Ixolaris were used for melanoma therapy. The procedures applied to them followed the standards described for the use of experimental animals, duly approved by the National Council for Animal Experimentation Control (CONCEA from Federal University of Rio de Janeiro. The accumulated activities were obtained in order to estimate the absorbed doses in each organ. Mass and metabolic differences between mice and humans were considered and used to extrapolate data acquired at different scales. From the dose factors provided by the MIRDOSE software, the absorbed doses in the target organs irradiated by the source organs were calculated and, finally, the effective dose was estimated. From the Student's t-hypothesis test performed in the accumulated activity, absorbed dose and S-factor quantities, there is no statistically significant difference between performing the image dosimetry from SPECT and planar acquired images.