B16F10 Murine Melanoma Cells Research Articles

Molecular Imaging of Melanoma VEGF-expressing Tumors through [99mTc]Tc-HYNIC-Fab(Bevacizumab).

Angiogenesis is a process that many tumors depend on for growth, development, and metastasis. Vascular endothelial growth factor (VEGF) is one of the major players in tumor angiogenesis in several tumor types, including melanoma. VEGF inhibition is achieved by bevacizumab, a humanized monoclonal antibody that binds with high affinity to VEGF and prevents its function. In order to successfully enable in vivo VEGF expression imaging in a murine melanoma model, we previously labeled bevacizumab with [99mTc]Tc. We observed that this was feasible, but it had prolonged blood circulation and delayed tumor uptake. The aim of this study was to develop a radiolabeled Fab bevacizumab fragment, [99mTc]Tc-HYNICFab( bevacizumab), for non-invasive in vivo VEGF expression molecular imaging. Flow cytometry was used to examine VEGF presence in the murine melanoma cell line (B16-F10). Bevacizumab was digested with papain for six hours at 37°C to produce Fab(bevacizumab), which was then conjugated to NHS-HYNIC-Tfa for radiolabeling with [99mTc]Tc. Stability and binding affinity assays were also evaluated. Biodistribution and single photon emission computed tomography/computed tomography (SPECT/CT) were performed at 1, 3, and 6 h (n = 4) after injection of [99mTc]Tc-HYNIC-Fab(Bevacizumab) in normal and B16-F10 tumor-bearing C57Bl/6J mice. Using flow cytometry, it was shown that the B16-F10 murine melanoma cell line has intracellular VEGF expression. Papain incubation resulted in the complete digestion of bevacizumab with good purity and homogeneity. The radiolabeling yield of [99mTc]Tc-HYNIC-Fab(bevacizumab) was 85.00 ± 6.06%, with a specific activity of 291.87 ± 18.84 MBq/mg (n=3), showing in vitro stability. Binding assays demonstrated significant intracellular in vitro VEGF expression. Fast blood clearance and high kidney and tumor uptake were observed in biodistribution and SPECT/CT studies. We present the development and evaluation of [99mTc]Tc-HYNIC-Fab(bevacizumab), a novel molecular VEGF expression imaging agent that may be used for precision medicine in melanoma and potentially in other VEGF-expressing tumors.

Enhanced Oncolytic Potential of Engineered Newcastle Disease Virus Lasota Strain through Modification of Its F Protein Cleavage Site.

Newcastle disease virus (NDV) is an oncolytic virus whose F protein cleavage activity is associated with viral infectivity. To explore the potential of modifying F protein cleavage activity to enhance antitumor effects, we constructed a recombinant NDV LaSota strain by replacing its F protein cleavage site with that from the mesogenic Beaudette C (BC) strain using reverse genetics techniques. The resulting virus, rLaSota-BC-RFP, demonstrated significantly enhanced infectivity and tumor cell suppression on the murine melanoma B16F10 cell, characterized by higher cytotoxicity and increased apoptosis compared to its parental strain, rLaSota-RFP. In vivo, rLaSota-BC-RFP treatment of B16F10 tumors in C57BL/6 mice resulted in significant tumor growth inhibition, improved survival rate, and induction of tumor-specific apoptosis and necrosis. Additionally, the rLaSota-BC-RFP treatment enhanced immunostimulatory effects within the tumor microenvironment (TME), characterized by increased infiltration of CD4+ and CD8+ T cells and elevated levels of antitumor immune modulator cytokines, including mouse IL-12, IFN-γ, IL-15, and TNF-α, in the rLaSota-BC-RFP-treated tumor tissues. Collectively, these findings demonstrate that the mesogenic F protein cleavage site enhances the oncolytic potential of the NDV LaSota strain, suggesting that rLaSota-BC-RFP is a promising oncolytic viral vector for gene delivery in cancer immunotherapy.

Liposome preparation of alpha-arbutin: stability and toxicity assessment using mouse B16F10 melanoma cells

ABSTRACT Melanoma is the most aggressive type of skin cancer, with few therapeutic alternatives following metastasis development. In recent years, drug delivery-associated nanotechnology has shown promising targeted results with diminished adverse effects compared to conventional treatments. This study aimed to (1) examine the effects of plant-derived α-arbutin, a natural compound and (2) compare these findings with bioactively developed liposomes containing α-arbutin utilizing the B16-F10 murine melanoma cell line as a model. Liposomes were obtained through reversed-phase evaporation by applying a spray dryer to assess their stability. The following biologic assays were measured cytotoxicity/antiproliferative (MTT, Neutral Red, and dsDNA PicoGreen). In addition, the levels of melanin and purinergic enzymes were also measured. The production of reactive oxygen species (ROS) and nitric oxide (NO) was determined as a measure of oxidative state. Treatment with nano-liposome containing alpha-arbutin induced a significant 68.4% cytotoxicity, similar to the positive control, in the B16-F10 murine melanoma cell line at 72 hr. Further, arbutin and liposomes containing alpha-arbutin increased levels of ROS and nitrite formation at 72 hr at the highest concentration (100 and 300 µg/ml) of treatments. Arbutin and liposomes containing alpha-arbutin reduced melanin levels at all tested concentrations. In addition, arbutin and alpha-arbutin containing liposomes lowered nucleotides (AMP, ADP, and ATP) and nucleoside (adenosine) levels in melanoma cells. Evidence suggests that α-arbutin containing liposome can be considered as an alternative immunosuppressive agent stimulated in melanoma treatment.

Development and evaluation of berberine-loaded bigel for the treatment of hyperpigmentation on B16F10 melanoma cell line.

Aim: The aim of this study was to optimize, develop, characterize and evaluate a topical nanobigel (BG) formulation containing Berberine (BRB) that exhibits anti-melanogenic properties.Materials & methods: The Berberine-loaded bigel (BRB@BG) formulation was prepared by homogenously mixing the optimized hydrogel and oleogel. BRB@BG was characterized in vitro and cytotoxicity study was conducted to evaluate its effects on murine skin melanoma B16F10 cell lines.Results: The optimized BRB@BG exhibited uniform texture with nanometric size, desirable spreadability and extrudability, suitable for topical applications. Cytotoxicity studies revealed that BRB@BG had a lower IC50 value (4.84μg/ml) on B16F10 cell lines compared with drug alone.Conclusion: In conclusion, the developed BRB@BG formulation showed good potential as safe and effective topical treatment for hyperpigmentation.

Syringaresinol Attenuates α-Melanocyte-Stimulating Hormone-Induced Reactive Oxygen Species Generation and Melanogenesis.

Ginseng has been utilized for centuries in both the medicinal and cosmetic realms. Recent studies have actively investigated the biological activity of ginseng berry and its constituents. (+)-Syringaresinol [(+)-SYR], an active component of ginseng berry, has been demonstrated to have beneficial effects on the skin, but its potential impact on skin pigmentation has not been fully explored. Here, the antioxidant and anti-pigmentary activity of (+)-SYR were evaluated in B16F10 murine melanoma cells and in an artificial human pigmented skin model, Melanoderm™. A real-time PCR, Western blotting, immunofluorescence staining, and histochemistry staining were conducted to confirm the effects of (+)-SYR on pigmentation. (+)-SYR reduced melanogenesis and dendrite elongation in α-melanocyte-stimulating hormone (α-MSH)-primed B16F10 cells with low cytotoxicity. (+)-SYR suppressed the expression of melanogenic genes, namely tyrosinase (TYR), tyrosinase-related protein 1 (TRP-1), and tyrosinase-related protein 2 (TRP-2). Notably, (+)-SYR attenuated α-MSH-induced cytosolic and mitochondrial reactive oxygen species (ROS) generation, which was attributable at least in part to the suppression of NADPH oxidase-4 (NOX 4) expression. Finally, the brightening activities of (+)-SYR were verified using Melanoderm™, underscoring the potential of ginseng berry and (+)-SYR as functional ingredients in skin-brightening cosmetics.

The Potential of Photodynamic Therapy Using Solid Lipid Nanoparticles with Aluminum Phthalocyanine Chloride as a Nanocarrier for Modulating Immunogenic Cell Death in Murine Melanoma In Vitro.

Photodynamic therapy (PDT) uses a photosensitizer to generate reactive oxygen species (ROS) that kill target cells. In cancer treatments, PDT can potentially induce immunogenic cell death (ICD), which is characterized by a well-controlled exposure of damage-associated molecular patterns (DAMPs) that activate dendritic cells (DCs) and consequently modulate the immune response in the tumor microenvironment. However, PDT still has limitations, such as the activity of photosensitizers in aqueous media and poor bioavailability. Therefore, a new photosensitizer system, SLN-AlPc, has been developed to improve the therapeutic efficacy of PDT. In vitro experiments showed that the light-excited nanocarrier increased ROS production in murine melanoma B16-F10 cells and modulated the profile of DCs. PDT induced cell death accompanied by the exposure of DAMPs and the formation of autophagosomes. In addition, the DCs exposed to PDT-treated B16-F10 cells exhibited morphological changes, increased expression of MHCII, CD86, CD80, and production of IL-12 and IFN-γ, suggesting immune activation towards an antitumor profile. These results indicate that the SLNs-AlPc protocol has the potential to improve PDT efficacy by inducing ICD and activating DCs.

CX3CL1 release during immunogenic apoptosis is associated with enhanced anti-tumour immunity.

Immunogenic cell death (ICD) has emerged as a novel option for cancer immunotherapy. The key determinants of ICD encompass antigenicity (the presence of antigens) and adjuvanticity, which involves the release of damage-associated molecular patterns (DAMPs) and various cytokines and chemokines. CX3CL1, also known as neurotactin or fractalkine, is a chemokine involved in cellular signalling and immune cell interactions. CX3CL1 has been denoted as a "find me" signal that stimulates chemotaxis of immune cells towards dying cells, facilitating efferocytosis and antigen presentation. However, in the context of ICD, it is uncertain whether CX3CL1 is an important mediator of the effects of ICD. In this study, we investigated the intricate role of CX3CL1 in immunogenic apoptosis induced by mitoxantrone (MTX) in cancer cells. The Luminex xMAP technology was used to quantify murine cytokines, chemokines and growth factors to identify pivotal regulatory cytokines released by murine fibrosarcoma MCA205 and melanoma B16-F10 cells undergoing ICD. Moreover, a murine tumour prophylactic vaccination model was employed to analyse the effect of CX3CL1 on the activation of an adaptive immune response against MCA205 cells undergoing ICD. Furthermore, thorough analysis of the TCGA-SKCM public dataset from 98 melanoma patients revealed the role of CX3CL1 and its receptor CX3CR1 in melanoma patients. Our findings demonstrate enhanced CX3CL1 release from apoptotic MCA205 and B16-F10 cells (regardless of the cell type) but not if they are undergoing ferroptosis or accidental necrosis. Moreover, the addition of recombinant CX3CL1 to non-immunogenic doses of MTX-treated, apoptotically dying cancer cells in the murine prophylactic tumour vaccination model induced a robust immunogenic response, effectively increasing the survival of the mice. Furthermore, analysis of melanoma patient data revealed enhanced survival rates in individuals exhibiting elevated levels of CD8+ T cells expressing CX3CR1. These data collectively underscore the importance of the release of CX3CL1 in eliciting an immunogenic response against dying cancer cells and suggest that CX3CL1 may serve as a key switch in conferring immunogenicity to apoptosis.

Polyfunctionalized organoselenides: New synthetic approach from selenium-containing cyanohydrins and anti-melanoma activity

A series of seleno-containing polyfunctionalized compounds was synthesized exploring cyanohydrin chemistry, including α-hydroxy esters, α-hydroxy acids, 1,2-diols, and 1,2-diacetates, with yields ranging from 26 up to 99 %. The cytotoxicity of all synthesized compounds was then evaluated using a non-tumor cell line (BALB/3T3 murine fibroblasts), and those deemed non-cytotoxic had their anti-melanoma activity evaluated using B16-F10 murine melanoma cells. These assays identified two compounds with selective cytotoxic activity against the tested melanoma cell line, showing a potential anti-melanoma application.

Maclura cochinchinensis (Lour.) Corner Heartwood Extracts Containing Resveratrol and Oxyresveratrol Inhibit Melanogenesis in B16F10 Melanoma Cells.

This study aimed to isolate and purify resveratrol and oxyresveratrol from the heartwoods of Maclura cochinchinensis, and to evaluate their inhibitory effects on melanogenesis in B16F10 murine melanoma cells. A methanol maceration process yielded a crude extract comprising 24.86% of the initial mass, which was subsequently analyzed through HPTLC, HPLC, and LC-MS/MS. These analyses revealed the presence of resveratrol and oxyresveratrol at concentrations of 4.32 mg/g and 33.6 mg/g in the extract, respectively. Initial purification employing food-grade silica gel column chromatography separated the extract into two fractions: FA, exhibiting potent inhibition of both tyrosinase activity and melanogenesis, and FM, showing no such inhibitory activity. Further purification processes led to the isolation of fractions Y11 and Gn12 with enhanced concentrations of resveratrol (94.9 and 110.21 mg/g, respectively) and fractions Gn15 and Gn16 with elevated levels of oxyresveratrol (321.93 and 274.59 mg/g, respectively), all of which significantly reduced melanin synthesis. These outcomes affirm the substantial presence of resveratrol and oxyresveratrol in the heartwood of M. cochinchinensis, indicating their promising role as natural agents for skin lightening.

DEHP (di(2-ethylhexyl)phthalate) stimulates skin pigmentation by perturbing cytoskeletal homeostasis.

Phthalates are extensively employed plasticizers crucial for conferring flexibility and plasticity to polyvinyl chloride. Phthalates, including DEHP (di(2-ethylhexyl)phthalate), present in diverse products, have been identified in fine dust and are capable of infiltrating the body, potentially posing health hazards. Importantly, melanocytes, existing at the basal layer of the epidermis, are susceptible to toxic substances. In our study, we employed the 3D human pigmented epidermis model, MelanoDerm™, along with the B16F10 murine melanoma cell line, to examine the influence of DEHP exposure on melanocytes. The exposure to low concentrations of DEHP (~ 5μM), resulted in the extension of melanocyte dendrites, indicating the stimulation of melanocytes. Analysis of gene expression and protein profiles unveiled the up-regulation of MITF, Arpc2, and TRP1 genes subsequent to DEHP exposure, indicating alterations in cytoskeletal and melanosome-related genetic and protein components in melanocytes. Notably, increased pigmentation was observed in MelanoDerm™ following DEHP exposure. DEHP-stimulated reactive oxygen species generation appeared to be involved in these events since the antioxidant, ascorbic acid attenuated ROS generation and MITF upregulation. Collectively, our study demonstrated that DEHP exposure can induce cytoskeletal disturbance and skin pigmentation through oxidative stress.

Bacillus Calmette-Guérin immunotherapy induces an efficient antitumor response to control murine melanoma depending on MyD88 signaling.

Bacillus Calmette-Guérin (BCG) is the first line treatment for bladder cancer and it is also proposed for melanoma immunotherapy. BCG modulates the tumor microenvironment (TME) inducing an antitumor effective response, but the immune mechanisms involved still poorly understood. The immune profile of B16-F10 murine melanoma cells was assessed by infecting these cells with BCG or stimulating them with agonists for different innate immune pathways such as TLRs, inflammasome, cGAS-STING and type I IFN. B16-F10 did not respond to any of those stimuli, except for type I IFN agonists, contrasting with bone marrow-derived macrophages (BMDMs) that showed high production of proinflammatory cytokines. Additionally, we confirmed that BCG is able to infect B16-F10, which in turn can activate macrophages and spleen cells from mice in co-culture experiments. Furthermore, we established a subcutaneous B16-F10 melanoma model for intratumoral BCG treatment and compared wild type mice to TLR2-/-, TLR3-/-, TLR4-/-, TLR7-/-, TLR3/7/9-/-, caspase 1-/-, caspase 11-/-, IL-1R-/-, cGAS-/-, STING-/-, IFNAR-/-, MyD88-/-deficient animals. These results in vivo demonstrate that MyD88 signaling is important for BCG immunotherapy to control melanoma in mice. Also, BCG fails to induce cytokine production in the co-culture experiments using B16-F10 and BMDMs or spleen cells derived from MyD88-/- compared to wild-type (WT) animals. Immunotherapy with BCG was not able to induce the recruitment of inflammatory cells in the TME from MyD88-/- mice, impairing tumor control and IFN-γ production by T cells. In conclusion, MyD88 impacts on both innate and adaptive responses to BCG leading to an efficient antitumor response against melanoma.

Improvement of protocol for detachment assays in B16F10 cells

B16F10 (B16) murine melanoma cells have the characteristic of strong adhesion to culture surfaces. The use of trypsin solutions in high doses to release this type of cell is not suitable. When B16F10 cells are released with high concentrations of trypsin they do not form a precipitate after centrifugation and consequently the cells will be lost when discarding the supernatant. To solve this problem, we established a solution with phosphate buffered saline (PBS) and ethylenediaminetetraacetic acid (EDTA) at low trypsin concentrations. Finally, we verified the efficiency of this solution on the quantity of cells recovered after centrifugation.

FETPY: a Diiron(I) Thio-Carbyne Complex with Prominent Anticancer Activity In Vitro and In Vivo.

FETPY, an organo-diiron(I) complex, showed strong cytotoxicity across a panel of human and mouse cancer cell lines, combined with an outstanding selectivity compared to nonmalignant cells. Enhanced iron uptake in aggressive, low-differentiated cell lines, caused membrane lipid peroxidation, which resulted in ferroptosis in human ovarian cancer cells. FETPY induced significant morphological changes in murine B16-F1 and B16-F10 melanoma cells, leading to senescence and/or trans-differentiation into Schwann-like cells, thus significantly reducing their tumorigenic potential. Additionally, FETPY substantially suppressed tumor growth in low- and high-grade syngeneic melanoma models when administered in a therapeutic regimen. FETPY is featured by satisfactory water solubility (millimolar range), an amphiphilic character (Log Pow = -0.17), and excellent stability in a biological medium (DMEM). These important requisites for drug development are rarely met in iron complexes investigated so far as possible anticancer agents. Overall, FETPY holds promise as a safe and potent targeted antitumor agent.

Glyphosate and aminomethylphosphonic acid metabolite (AMPA) modulate the phenotype of murine melanoma B16-F1 cells

Pesticides are contaminants run-offs from agricultural areas with a global concern due to their toxicity for non-target organisms. The Brazilian Health Surveillance Agency reported about 63% of the food contain pesticide residues. Glyphosate is a herbicide used worldwide but its toxicity is not a consensus among specialists around the world. AMPA (aminomethylphosphonic acid) is a glyphosate metabolite that can be more toxic than the parental molecule. Melanoma murine B16-F1 cells were exposed to glyphosate and AMPA to investigate the cell profile and possible induction to a more malignant phenotype. Glyphosate modulated the multi-drug resistance mechanisms by ABCB5 gene expression, decreasing cell attachment, increasing cell migration and inducing extracellular vesicles production, and the cells exposed to AMPA revealed potential damages to DNA. The present study observed that AMPA exhibits high cytotoxicity, which suggests a potential impact on non-tumor cells, which are, in general, more susceptible to chemical exposure. Conversely, glyphosate favored a more metastatic and chemoresistant behavior in cancer cells, highlighting the importance of additional research in this area.

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.

Rhodamine B Amide Derivatives with Anticancer Activity and Fluorescence Properties for Potential Theranostic Applications

AbstractRhodamine B (RHB) based derivatives are especially known for their dye characteristics. Nevertheless, very limited of such revealed promising in‐vivo and in‐vitro therapeutic potency; besides demonstrating great interest due to their excellent photophysical and structural proficiency. Therefore, novel RHB derivatives with significant biological activity are requisite. Hence, current work presents the contemporary design aspect, synthesis, characterization, and anticancer activity of five new RHB amides attained by reacting RHB specifically with simple small amines being side chains of existing anticancer drugs. Therefore, this strategy resulted in a dual effect i. e. fluorescence property and considerable anticancer activity among achieved RHB amides. Obtained fluorophores exhibited emission wavelength at ∼560 nm, a fluorescence quantum yield Φf=10 to 40 % in aqueous solutions, and showed operational stability in pH 4–10, conditions. Further, a study for in‐vitro cell‐based experiments was carried out, wherein AGS (Human gastric cancer) and B16F10 (Murine melanoma) cells were used to screen the compounds. Hydroxy phenyl piperazine substituted RHB amide showed greater growth inhibition than other compounds with an IC50 value of 1.18 μM and 9.07 μM in AGS and B16F10 cells. Fluorescence imaging was performed for the compounds in AGS cells with DAPI as the counterstain using confocal laser microscopy.

Efficient siRNA delivery to murine melanoma cells via a novel genipin-based nano-polymer.

Small-interfering RNAs (siRNAs) are therapeutic nucleic acids, often delivered via cationic polymers, liposomes, or extracellular vesicles, each method with its limitations. Genipin, a natural crosslinker for primary amines, was explored for siRNA delivery scaffolds. Spermine/genipin-based GxS5 polymers were synthesized, showing slightly positive ζ potential at neutral pH and intrinsic fluorescence. We then tuned their polymerization adding glycine to the reaction batch, from 1 to 10 molar ratio with genipin, therefore conferring them a "zwitterionic" character. GxS5 efficiently internalized into B16F10 murine melanoma cells, and exhibited strong siRNA-complexing ability and they were able to elicit up to 60% of gene knock-down without any toxicity. This highlights GxS5's potential as a safe, replicable, and tunable platform for therapeutic nucleic acid delivery, suggesting broader applications. This innovative approach not only sheds light on the intricate genipin reaction mechanism but also underscores the importance of fine-tuning nanoparticle properties for effective siRNA delivery. GxS5's success in mitigating cytotoxicity while maintaining delivery efficacy signifies a promising step towards safer and more efficient nucleic acid therapeutics.

Infection of tumor cells with Salmonella typhimurium mimics immunogenic cell death and elicits tumor-specific immune responses.

Some properties of Salmonella-infected cells overlap with immunogenic cell death. In this study, we demonstrated that intracellular infection of melanoma with Salmonella typhimurium induced high immunogenicity in melanoma cells, leading to antitumor effects with melanoma-antigen-specific T-cell responses. Murine B16F10 melanoma cells were infected with tdTomato-expressing attenuated S. typhimurium (VNP20009; VNP-tdT), triggering massive cell vacuolization. VNP-tdT-infected B16F10 cells were phagocytosed efficiently, which induced the activation of antigen-presenting cells with CD86 expression in vitro. Subcutaneous coimplantation of uninfected and VNP-tdT-infected B16F10 cells into C57BL/6 mice significantly suppressed tumor growth compared with the implantation of uninfected B16F10 cells alone. Inoculation of mice with VNP-tdT-infected B16F10 cells elicited the proliferation of melanoma-antigen (gp100)-specific T cells, and it protected the mice from the second tumor challenge of uninfected B16F10 cells. These results suggest that Salmonella-infected tumor cells acquire effective adjuvanticity, leading to ideal antitumor immune responses.

BDE-209 exposure in murine melanoma (B16–F1) cells modulates tumor malignancy and progression in vivo

Melanoma is a type of skin cancer considered aggressive due to its high metastatic ability and rapid progression to other tissues and organs. BDE-209 (2,2′,3,3′,4,4′,5,5′,6,6′-decabromodiphenyl ether) is an additive used as a flame retardant and classified as a persistent organic pollutant that has a high bioaccumulation capacity due to its lipophilic nature. This substance has already been detected in rivers, air, soil, plants and even in different human biological samples, such as plasma, umbilical cord blood and breast milk, revealing a great concern to human populations. Thus, in the current study we investigated whether prior exposure of murine melanoma B16–F1 cells to BDE-209 modulates in vivo progression and malignancy of melanoma. B16–F1 cells were cultured and exposed in vitro to BDE-209 (0.01, 0.1 e 1 nM) for 15 days and then inoculated, via caudal vein, in C57BL/6 mice for experimental metastasis analysis after 20 days. Inoculation of BDE-209-exposed cells resulted in 82% increase of metastasis colonized area in the lungs of mice, downregulation of tumor suppressors genes, such as Timp3 and Reck, decrease of lipid peroxidation and increase of systemic and local inflammatory response. These findings are related to melanoma progression. Additionally, the histopathological analysis revealed greater number of focal points of metastases in the lungs and invasiveness of metastases to the mice brain (89%). The results showed that exposure to BDE-209 may alter the phenotype of B16–F1 cells, worsening their metastatic profile. Current data showed that BDE-209 may interfere with the prognosis of melanoma by modulating cells with less invasiveness capacity to a more aggressive profile.