G2/M arrest and mitotic slippage induced by fenbendazole in canine melanoma cells

Euphorbia tirucalli Lineu (Euphorbiaceae) is a tropical and subtropical ornamental and toxic plant. E. tirucalli produces a latex that is commonly used to treat neoplasms. This study aimed to evaluate the effects of diluted E. tirucalli latex (DETL) on human (SK-MEL-28) and canine (CBMY) melanoma cells. SK-MEL-28 (3 × 103 cells/well) and CBMY (6 × 103 cells/well) were cultivated in 96-well plates. The cells were treated with 50 μl/well of dilutions (1/2, 1/4, 1/8, 1/16, 1/32, 1/64, 1/128, 1/256, and 1/512) of a standard solution containing 1 mg/mL of the E. tirucalli latex (ETL) in DMEM. Control group cells received 50 μl/well of DMEM. After 24, 48, and 72 h of treatment, cell viability was assessed by the MTT assay. There was a significant decrease in viability at 48 and 72 hours after treatment for human melanoma cells and at 24, 48, and 72 hours for canine cells, mainly in higher dilutions of ETL. Human melanoma cells presented a typical U shape curve, characteristic of hormesis. To our knowledge, this is the first study showing inhibitory effects of DETL on canine melanoma cells. Therefore, DETL is a potentially new antineoplastic drug.

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Investigation of the effects of mTOR inhibitors rapamycin and everolimus in combination with carboplatin on canine malignant melanoma cells

BMC Veterinary Research ◽

10.1186/s12917-021-03089-0 ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Sarah Bernard ◽

Andrew C. Poon ◽

Peyton M. Tam ◽

Anthony J. Mutsaers

Keyword(s):

Malignant Melanoma ◽

Cell Viability ◽

Cell Lines ◽

Tumour Progression ◽

Mtor Inhibitors ◽

Melanoma Cells ◽

Combination Drug ◽

Mechanistic Investigation ◽

Canine Melanoma

Abstract Background Malignant melanoma in dogs is considered to be largely resistant to conventional chemotherapy, although responses to carboplatin have been documented. Invasion and early metastasis are common features of certain melanoma subtypes that contribute to tumour progression despite aggressive local and systemic therapy. Upregulation of the PI3K/AKT/mTOR pathway has been observed in canine malignant melanoma and may represent a potential target for therapy. Rapamycin (sirolimus) and everolimus are commercially available small molecule inhibitors that target mTOR and therefore may have anticancer activity in canine melanoma. It was hypothesized that there is synergism between rapamycin or everolimus and platinum chemotherapy, and that combination drug treatment would inhibit target/downstream proteins involved in cell viability/proliferation and increase cell death in canine melanoma cells. It was further hypothesized that rapamycin or everolimus would impact metabolism by reducing glycolysis in these cells. Four canine melanoma cell lines were treated in vitro with rapamycin and everolimus as sole treatment or combined with carboplatin. Cell viability, apoptosis, target modulation, and glycolytic metabolism were evaluated by crystal violet colourimetric assay, Annexin V/PI flow cytometry, western blotting, and Seahorse bioanalyzer, respectively. Results When combined with carboplatin chemotherapy, rapamycin or everolimus treatment was overall synergistic in reducing cell viability. Carboplatin-induced apoptosis was noted at 72 h after treatment compared to the vehicle control. Levels of phosphorylated mTOR were reduced by rapamycin and everolimus in all four cell lines, but activation of the downstream protein p70S6K was not consistently reduced by treatment in two of the cell lines. Both mTOR inhibitors decreased the extracellular acidification rate of canine melanoma cells, indicating reduced cancer cell glycolytic activity. Conclusions Inhibition of mTOR by rapalogs, such as rapamycin and everolimus combined with carboplatin chemotherapy may have activity in canine melanoma. Future mechanistic investigation is warranted, including in vivo assessment of this combination therapy.

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Involvement of GLUT1 and GLUT3 in the growth of canine melanoma cells

PLoS ONE ◽

10.1371/journal.pone.0243859 ◽

2021 ◽

Vol 16 (2) ◽

pp. e0243859

Author(s):

Yoko Suwabe ◽

Rei Nakano ◽

Shinichi Namba ◽

Naoya Yachiku ◽

Manami Kuji ◽

...

Keyword(s):

Cell Growth ◽

Glucose Uptake ◽

Cancer Cells ◽

Glucose Transporter ◽

Glucose Consumption ◽

Melanoma Cells ◽

Dependent Manner ◽

Canine Melanoma ◽

Fully Functioning ◽

Dose Dependent

The rate of glucose uptake dramatically increases in cancer cells even in the presence of oxygen and fully functioning mitochondria. Cancer cells produce ATP by glycolysis rather than oxidative phosphorylation under aerobic conditions, a process termed as the “Warburg effect.” In the present study, we treated canine melanoma cells with the glucose analog 2-deoxy-D-glucose (2-DG) and investigated its effect on cell growth. 2-DG attenuated cell growth in a time- and dose-dependent manner. Cell growth was also inhibited following treatment with the glucose transporter (GLUT) inhibitor WZB-117. The treatment of 2-DG and WZB-117 attenuated the glucose consumption, lactate secretion and glucose uptake of the cells. The mRNA expression of the subtypes of GLUT was examined and GLUT1 and GLUT3 were found to be expressed in melanoma cells. The growth, glucose consumption and lactate secretion of melanoma cells transfected with siRNAs of specific for GLUT1 and GLUT3 was suppressed. These findings suggest that glucose uptake via GLUT1 and GLUT3 plays a crucial role for the growth of canine melanoma cells.

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DNA methylation contributes toward silencing of antioncogenic microRNA-203 in human and canine melanoma cells

Melanoma Research ◽

10.1097/cmr.0000000000000183 ◽

2015 ◽

Vol 25 (5) ◽

pp. 390-398 ◽

Cited By ~ 17

Author(s):

Shunsuke Noguchi ◽

Takashi Mori ◽

Takayuki Nakagawa ◽

Kazuhito Itamoto ◽

Tomoya Haraguchi ◽

...

Keyword(s):

Dna Methylation ◽

Melanoma Cells ◽

Canine Melanoma

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Abstract B36: The effects of dasatinib in KIT L579P and NRAS Q61K mutant canine melanoma cells

10.1158/1538-7445.mel2014-b36 ◽

2015 ◽

Author(s):

Lu-Ping Lu ◽

Chen-Si Lin ◽

Albert Taiching Liao

Keyword(s):

Melanoma Cells ◽

Canine Melanoma

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Sequential detection of mRNA for the chemokine IL-8 and macrophages (F4/80) in human melanoma

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010014141x ◽

1995 ◽

Vol 53 ◽

pp. 1008-1009

Author(s):

C.D. Bucana ◽

R. Sanchez ◽

R. Singh ◽

I.J. Fidler

Keyword(s):

Tumor Cells ◽

Nude Mice ◽

Constitutive Expression ◽

Metastatic Potential ◽

Melanoma Cells ◽

Human Melanoma ◽

Angiogenic Factor ◽

Infiltrating Cells ◽

Immunoperoxidase Assay ◽

Multifunctional Cytokine

The purpose of this study was to demonstrate by ISH the presence of IL-8 mRNA, and by immunohistochemistry (IHC) the presence of the chemokine IL-8 and the distribution of infiltrating macrophages in subcutaneous melanomas in the same tumor. IL-8 is a multifunctional cytokine produced by melanoma cells, activated macrophages and monocytes and it has been shown to be a growth and angiogenic factor for tumor cells. More recently it was shown that constitutive expression of IL-8 correlated directly with metastatic potential of human melanoma cells in nude mice. IL-8 content of a solid tumor as determined by Western blot analysis does not take into account the contribution of macrophages. Previous studies showed that murine tumors contain many infiltrating cells interspersed among tumor cells whereas human tumors growing in nude mice exhibit macrophages at the periphery or between tumor islands. In this study we demonstrate the expression of IL-8 and the distribution of macrophages by immunoperoxidase assay and IL-8 mRNA by ISH.

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