Vascular endothelial growth factor-C in activating vascular endothelial growth factor receptor-3 and chemokine receptor-4 in melanoma adhesion

Vascular endothelial growth factor-C (VEGF-C) binds to receptor vascular endothelial growth factor receptor-3 (VEGFR-3) expressed on lymphatic endothelial and melanoma cells. Binding of VEGF-C to VEGFR-3 enhances receptor phosphorylation that activates mitogen-activated protein kinase (MAP-K) and phosphatidylinositol-3-kinase (PI3K). These signalling pathways regulate cell migration and adhesion in response to internal or external changes. In addition, the overexpression of VEGF-C upregulates chemokine receptor CXCR-4 in tumours (melanoma). CXCR-4 is expressed on cells of the immune system (natural killer cells) and facilitates the migration of leukocytes in response to the CXCL12 ligand. The latter is expressed by lymphatic endothelial cells and by stromal cells in the tumour microenvironment (TME). The gradient established between CXCR-4 expressed on tumour cells and CXCL12 produced by stromal and lymphatic endothelial cells enhances tumour cell metastasis. 3-(4-Dimethylamino-naphthalen-1-ylmethylene)-1, 3-dihydroindol-2-one, MAZ-51, is an indolinone-based synthetic molecule that inhibits the phosphorylation of the tyrosine kinase receptor VEGFR-3. CTCE-9908, a CXCR-4 antagonist derived from human CXCL12, hinders receptor phosphorylation and the subsequent signalling pathways that would be activated. VEGF-C is stimulated by transforming growth factor-beta 1 (TGF-β1), which facilitates cell–cell and cell-matrix adhesion by regulating cadherins through the activation of focal adhesion kinase (FAK) and mediates paxillin upregulation. Increased VEGF-C protein levels stimulated by TGF-β bound to VEGFR-3 impact on intracellular pathways that promote tumour cell adhesion. In addition, increased VEGF-C protein levels lead to enhanced CXCR-4 protein expression. Therefore, effective blocking of VEGR-3 and CXCR-4 may inhibit tumour cell metastasis by hampering intracellular proteins promoting adhesion.The Association of African Universities (AAU), the National Research Foundation (NRF) of South Africa, the School of Medicine Research Committee, the Faculty of Health Sciences, small grants programme and PhD completion funding scheme at the University of Pretoria, Pretoria, South Africa.http://wileyonlinelibrary.com/journal/jcmmhj2023Physiolog

In vitro effects of papaverine on cell migration and vascular endothelial growth factor in cancer cell lines

Papaverine (PPV) is a benzylisoquinoline alkaloid isolated from Papaver somniferum that exerts antiproliferative activity. However, several questions remain regarding the biochemical pathways affected by PPV in tumourigenic cells. In this study, the influence of PPV on cell migration (light microscopy), expression of vascular endothelial growth factor (VEGF) B, VEGF R1, VEGF R2, and phosphorylated focal adhesion kinase (pFAK) were investigated using spectrophotometry in MDA-MB-231-, A549- and DU145 cell lines. The migration assay revealed that, after 48 h, PPV (100 µM) reduced cell migration to 81%, 91%, and 71% in MDA-MB-231-, A549-, and DU145 cells, respectively. VEGF B expression was reduced to 0.79-, 0.71-, and 0.73-fold after 48 h of exposure to PPV in MDA-MB-231-, A549- and DU145 cells, while PPV exposure of 48 h increased VEGF R1 expression in MDA-MB-231- and DU145 cells to 1.38 and 1.46. A fold decrease in VEGF R1 expression was observed in A549 cells to 0.90 after exposure to 150 µM. No statistically significant effects were observed on VEGF R2- and FAK expression after exposure to PPV. This study contributes to the understanding of the effects of a phytomedicinal alkaloid compound in cancer cells and may provide novel approaches to the application of non-addictive alkaloids.The Cancer Association of South Africa, Medical Research Council, the Struwig Germeshuysen Trust, School of Medicine Research Committee of the University of Pretoria and the South African National Research Foundation.https://www.mdpi.com/journal/ijerphPhysiolog

Sulphamoylated estradiol analogue induces reactive oxygen species generation to exert its antiproliferative activity in breast cancer cell lines

2-Methoxyestradiol (2ME), a 17β-estradiol metabolite, exerts anticancer properties in vitro and in vivo. To address 2ME’s low bioavailability, research led to the in silico design of sulphamoylated 2ME analogues. However, the role of oxidative stress induced in the activity exerted by sulphamoylated compounds remains elusive. In the current study, the influence of 2-Ethyl-17-oxoestra-1,3,5(10)-trien-3-yl sulphamate (ESE-one) on reactive oxygen species (ROS) induction and its effect on cell proliferation, as well as morphology, were assessed in breast tumorigenic cells (MCF-7 and MDA-MB-231). Fluorescent microscopy showed that sulphamoylated estradiol analogues induced hydrogen peroxide and superoxide anion, correlating with decreased cell growth demonstrated by spectrophotometry data. ESE-one exposure resulted in antiproliferation which was repressed by tiron (superoxide inhibitor), trolox (peroxyl inhibitor) and N,N0 -dimethylthiourea (DMTU) (hydrogen peroxide inhibitor). Morphological studies demonstrated that tiron, trolox and DMTU significantly decreased the number of rounded cells and shrunken cells in MCF-7 and MDA-MB-231 cells induced by ESE-one. This in vitro study suggests that ESE-one induces growth inhibition and cell rounding by production of superoxide anion, peroxyl radical and hydrogen peroxide. Identification of these biological changes in cancer cells caused by sulphamoylated compounds hugely contributes towards improvement of anticancer strategies and the ROS-dependent cell death pathways in tumorigenic breast cells.Cancer Association of South Africa, Medical Research Council, Struwig Germeshuysen Trust, School of Medicine Research Committee of the University of Pretoria, South African National Research Foundation and Department of Physiology and the School of Medicine, Faculty of Health Sciences, University of Pretoria.http://www.mdpi.com/journal/moleculespm2020Physiolog

Modes of cell death induced by tetrahydroisoquinoline-based analogs in MDA-MB-231 breast and A549 lung cancer cell lines

BACKGROUND : A and B rings of the steroidal microtubule disruptor, 2-methoxyestradiol, and its analogs can be mimicked with a tetrahydroisoquinoline (THIQ) core. THIQs are cytotoxic agents with potential anticancer activities. The aim of this in vitro study was to investigate the modes of cell death induced by four nonsteroidal THIQ-based analogs, such as STX 2895, STX 3329, STX 3451 and STX 3450, on MDA-MB-231 metastatic breast and A549 epithelial lung carcinoma cells. MATERIALS AND METHODS : Cytotoxicity studies determined the half-maximal growth inhibitory concentration of the analogs to be at nanomolar concentrations without the induction of necrosis. Light and fluorescent microscopy determined that compounds caused microtubule depolymerization and displayed morphological hallmarks of apoptosis. RESULTS : Flow cytometric analyses confirmed apoptosis induction as well as an increased G2/M phase on cell cycle analysis. Furthermore, intrinsic pathway signaling was implicated due to increased cytochrome c release and a decrease in mitochondrial transmembrane potential. Potential involvement of autophagy was observed due to increased acidic vacuole formation and increased aggresome activation factor. CONCLUSION : Thus, it can be concluded that these four THIQ-based analogs exert anti- proliferative and antimitotic effects, induce apoptosis and involve autophagic processes. Further investigation into the efficacy of these potential anticancer drugs will be conducted in vitro and in vivo.The abstract of this paper was presented at the 24 Biennial Congress of the European Association for Cancer Research, July 9–12, 2016, Manchester, UK, and was published in the European Journal of Cancer.Grants from the Medical Research Council of South Africa, the Cancer Association of South Africa, National Research Foundation and the Struwig-Germeshuysen Cancer Research Trust of South Africa. BVLP is a Wellcome Trust Senior Investigator (grant 101010).http://www.dovepress.com/drug-design-development-and-therapy-journalam2018Physiolog

Apoptotic profiling of chronic myeloid leukaemia patients' platelets ex vivo before and after treatment with Imatinib

Chronic myeloid leukaemia (CML) is a malignancy of the haematopoietic stem cells. The first line of treatment for CML, especially in developing countries, remains the first-generation tyrosine kinase inhibitor, Imatinib. Patients with CML are frequently diagnosed with platelet abnormalities. However, the specific mechanism of platelet abnormalities in CML remains unclear and poorly understood. The aim of this study was therefore to determine the apoptotic profiles of CML patients ex vivo on platelets before and after treatment with Imatinib. Blood samples of healthy volunteers and CML patients at diagnosis and after 6 months treatment with Imatinib were collected. Platelet counts, viability and activation were determined. Results showed that CML patients' platelet counts were elevated upon diagnosis and these levels statistically significantly decreased after 6 months of treatment. Platelet activation was significantly increased after 6 months of treatment compared to levels at diagnosis (P-value < .05). Similarly, platelet apoptosis was also increased after 6 months of treatment. Abnormalities in platelet functioning found in this study may partly be due to clonal proliferation of haematopoietic cells in CML patients, specifically of megakaryocyte precursors as well as the inhibition of platelet tyrosine kinase's and the inhibition of platelet-derived growth factor.Cancer Association of South Africa; Medical Research Council of South Africa; National Research Foundation; School of Medicine Research Committee of the Faculty of Health Sciences, University of Pretoria; Struwig-Germeshuysen Research Trust.http://wileyonlinelibrary.com/journal/cbfhj2022HaematologyInternal MedicinePhysiolog

Cell fate following irradiation of MDA-MB-231 and MCF-7 breast cancer cells pre-exposed to the setrahydroisoquinoline sulfamate microtubule disruptor STX3451

The compound STX3451 is not commercially available.SUPPLEMENTARY MATERIAL : TABLE S1: Data analysis comparing flow cytometric quantification of individual cell cycle phases across 24-h and 48-h timelines. TABLE S2: Data analysis of flow cytometric quantification of the cell cycle distribution in MCF-7 cells exposed to STX3451 and radiation. TABLE S3: Statistical analysis of cell cycle distribution in MCF-7 cells exposed to STX3451 and radiation. TABLE S4: Data analysis comparing flow cytometric quantification of individual cell cycle phases across 24-h and 48-h timelines in MDA-MB-231 cells. TABLE S5: Statistical analysis of cell cycle progression in MDA-MB-231 cells exposed to STX3451 and radiation. TABLE S6: Statistical analysis of cell cycle distribution in MDA-MB-231 cells exposed to STX3451 and radiation. TABLE S7: Annexin-V analysis of MCF-7 cells 48-h. TABLE S8: Annexin-V statistical analysis of MDA-MB-231 48-h. TABLE S9: Colony formation in MCF-7 cells. TABLE S10: Colony formation in MDA-MB-231 cells. TABLE S11: The total number of Mn in MCF-7 cells that were terminated 2- and 24-h after radiation. TABLE S12: The total number of Mn in MDA-MB-231 cells terminated 2- and 24-h after radiation. TABLE S13: Number of Mn per cell in MCF-7 cells terminated 2-h after radiation. TABLE S14: Number of Mn per cell in MCF-7 cells terminated 24-h after radiation. TABLE S15: Number of Mn per cell in MDA-MB-231 cells terminated 2-h after radiation. TABLE S16: The number of Mn per cell in MDA-MB-231 cells that were terminated 24-h after radiation. TABLE S17: Superoxide detection in MCF-7 cells treated with the various modalities. TABLE S18: Superoxide detection in pre-sensitized MDA-MB-231 cells. TABLE S19: Statistical analysis of ATM expression in combination treated MCF-7 and MDA-MB-231 cells 2- and 24-h post-radiation. TABLE S20: Nontumored animal toxicity assay; VIDEO S1: not applicable.Atetrahydroisoquinoline (THIQ) core is able tomimic theAand B rings of 2-methoxyestradiol (2ME2), an endogenous estrogen metabolite that demonstrates promising anticancer properties primarily by disrupting microtubule dynamic instability parameters, but has very poor pharmaceutical properties that can be improved by sulfamoylation. The non-steroidal THIQ-based microtubule disruptor 2-(3-bromo-4,5-dimethoxybenzyl)-7-methoxy-6-sulfamoyloxy-1,2,3,4-tetrahydroisoquinoline (STX3451), with enhanced pharmacokinetic and pharmacodynamic profiles, was explored for the first time in radiation biology. We investigated whether 24 h pre-treatment with STX3451 could pre-sensitize MCF-7 and MDA-MB-231 breast cancer cells to radiation. This regimen showed a clear increase in cytotoxicity compared to the individual modalities, results that were contiguous in spectrophotometric analysis, flow cytometric quantification of apoptosis induction, clonogenic studies and microscopy techniques. Drug pre-treatment increased radiation-induced DNA damage, with statistically more double-strand (ds) DNA breaks demonstrated. The latter could be due to the induction of a radiation-sensitive metaphase block or the increased levels of reactive oxygen species, both evident after compound exposure. STX3451 pre-exposure may also delay DNA repair mechanisms, as the DNA damage response element ataxia telangiectasia mutated (ATM) was depressed. These in vitro findings may translate into in vivo models, with the ultimate aim of reducing both radiation and drug doses for maximal clinical effect with minimal adverse effects.The Research Committee of the University of Pretoria, the Struwig-Germeshuysen Trust, the Cancer Association of South Africa (CANSA), the National Research Foundation (NRF) and the Research Development Programme of the University of Pretoria (RDP-UP).https://www.mdpi.com/journal/moleculesam2023Physiolog

The effects of kynurenine metabolites on cell proliferation and morphology in melanoma and neuroblastoma cell lines

Research Development Programme (RDP) (UP)UCD Granthttps://drive.google.com/file/d/1juIb6_CBLaV_p1W3lCCoiy6DKY4CoWJB/view?usp=sharinghttps://drive.google.com/drive/folders/15c8nNl2iUi3wTrFeIUNHHMx9ctqb2gcc?usp=sharinghttps://drive.google.com/drive/folders/1fwqlMm1hRp4TyUwXiAXdvF6yDWyPHgcb?usp=sharin

The involvement of a chemokine receptor antagonist CTCE-9908 and kynurenine metabolites in cancer development

Cancer is the second leading cause of mortality worldwide. Skin cancer is the most common cancer in South Africa with nearly 20,000 reported cases every year and 700 deaths. If diagnosed early, the 5‐year survival rate is about 90%, however, when diagnosed late, the 5‐year survival rate decreases to about 20%. Melanoma is a type of skin cancer with an estimated 5‐year survival rate of approximately 90%. Neuroblastoma is a paediatric cancer with a low survival rate. Sixty percent of patients with metastatic disease do not survive 5 years after diagnosis. Despite recent advances in targeted therapies, there is a crucial need to identify reliable prognostic biomarkers which will be able to contribute to the development of more precision‐based chemotherapeutic strategies to prevent tumour migration and metastasis. The compound, CTCE‐9908 inhibits the binding of CXC chemokine ligand 12 (CXCL12) to the CXC chemokine receptor 4 (CXCR4) receptor leading to reduced metastasis. Kynurenine metabolites are derived tryptophan, which is an essential amino acid. Kynurenine metabolites inhibit T‐cell proliferation resulting in cell growth arrest. For this reason, chemokines receptors represent potential targets for the treatment of cancer growth and metastasis. In this review paper, the role of the CXCL12/CXCR4 signalling pathway in the development of cancer is highlighted together with the current available treatments involving the CTCE‐9908 compound in combination with microtubule inhibitors like paclitaxel and docetaxel.The National Research Foundation, Struwig‐ Germeshuysen Research Trust, the Medical Research Council of South Africa, the Cancer Association of South Africa, the School of Medicine Research Committee of the Faculty of Health Sciences , the Research Development Programme and the University Capacity Development Programme at the University of Pretoria.http://onlinelibrary.wiley.com/journal/10.1002/(ISSN)1099-0844am2023AnatomyPhysiolog

Comparison of structures and cytotoxicity of mupirocin and batumin against melanoma and several other cancer cell lines

AIM : To determine the computer-predicted anticancer activity of mupirocin and to compare its activities with those determined for another polyene antibiotic, batumin. MATERIALS AND METHODS : Molecular docking, cytotoxicity assays, cell microscopy and cell cycle progression were studied in cancer and nontumorigenic cell lines. RESULTS AND CONCLUSION : Cytotoxicity of mupirocin against several cancerous cell lines was detected with the highest one (IC50 = 5.4 μg/ml) against melanoma cell line. The profile of cytotoxicity of mupirocin was similar to that reported for batumin. Nevertheless, the morphology of cells treated with these antibiotics and alterations in cell cycle progression suggested possible dissimilarity in their mechanisms of action. Selective cytotoxicity of mupirocin against melanoma cells potentiates further studies to discover nontoxic drugs for melanoma prevention.AM Joubert acquired grants from the National Research Foundation (NRF; 105992, 90523 and 85818), Cancer Association of South Africa (A0V741 and A0W228), the Struwig Germeshuysen Trust (A0N074), the School of Medicine Research Committee of the University of Pretoria (AOH561) and Medical Research Council (A0W110). MH Visagie was funded from the NRF (99706), the School of Medicine Research Committee of the University of Pretoria (AOH561) and Struwig Germeshysen Trust. N Lall was funded from NRF/IKS160514165042 Grant No: 105169. ON Reva was funded from NRF grant 105996.https://www.future-science.com/journal/fmchj2019BiochemistryGeneticsMicrobiology and Plant PathologyPhysiologyPlant Production and Soil Scienc

Promising anticancer activity of batumin : a natural polyene antibiotic produced by Pseudomonas batumici

AIM : To determine the computer-predicted anticancer activity of antibiotic batumin. MATERIALS AND METHODS : Cytotoxicity assays, cell morphology microscopy and cell cycle progression were studied in cancer and nontumorigenic cell lines. An in vivo experiment on Lewis lung carcinoma (3LL)-transplanted mice was conducted to evaluate potential antimetastatic. RESULTS AND CONCLUSION : Cytotoxicity against melanoma and lung carcinoma cells (IC50 ≈ 5 μg/ml) was detected. Hypercondensed chromatin and apoptotic body formation in batumin-treated cells suggested the induction of apoptosis supported also by an observed increase in the quantity of cells occupying the sub-G1 cell cycle phase. Twofold reduction in the number and volume of lung metastases in Lewis lung carcinoma (3LL)-bearing batumin-treated mice was demonstrated. Highly specific cytotoxicity of batumin against cancer cell lines potentiates further studies.https://www.future-science.com/loi/fmc2019-09-01hj2018BiochemistryGeneticsMicrobiology and Plant PathologyPhysiologyPlant Production and Soil Scienc