Identification and characterisation of NANOG+/ OCT-4high/SOX2+ doxorubicin-resistant stem-like cells from transformed trophoblastic cell lines

Balahmar, Reham M.; Boocock, David J.; Coveney, Clare; Ray, Sankalita; Vadakekolathu, Jayakumar; Regad, Tarik; Ali, Selman A.; Sivasubramaniam, Shiva

doi:10.18632/oncotarget.24151

Cited by 11 publications

(9 citation statements)

References 30 publications

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“…Spectral library generation, alignment and fold change analysis were performed as described previously [68]. In brief, IDA data were searched using ProteinPilot 5.0 (iodoacetamide alkylation, biological modifications emphasised in a thorough search) against the Swiss-Prot human database (June 2017).…”

Section: Mass Spectrometrymentioning

confidence: 99%

β2-Adrenergic Signalling Promotes Cell Migration by Upregulating Expression of the Metastasis-Associated Molecule LYPD3

Gruet

Cotton

Coveney

et al. 2020

Biology

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Metastasis is associated with poor prognosis in breast cancer. Although some studies suggest beta-blockers increase survival by delaying metastasis, others have been discordant. This study provides both insights into the anomalous findings and identifies potential biomarkers that may be treatment targets. Cell line models of basal-type and oestrogen receptor-positive breast cancer were profiled for basal levels of adrenoceptor gene/protein expression, and β2-adrenoceptor mediated cell behaviour including migration, invasion, adhesion, and survival in response to adrenoceptor agonist/antagonist treatment. Protein profiling and histology identified biomarkers and drug targets. Baseline levels of adrenoceptor gene expression are higher in basal-type rather than oestrogen receptor-positive cancer cells. Norepinephrine (NE) treatment increased invasive capacity in all cell lines but did not increase proliferation/survival. Protein profiling revealed the upregulation of the pro-metastatic gene Ly6/PLAUR Domain-Containing Protein 3 (LYPD3) in norepinephrine-treated MDA-MB-468 cells. Histology confirmed selective LYPD3 expression in primary and metastatic breast tumour samples. These findings demonstrate that basal-type cancer cells show a more aggressive adrenoceptor-β2-activated phenotype in the resting and stimulated state, which is attenuated by adrenoceptor-β2 inhibition. This study also highlights the first association between ADRβ2 signalling and LYPD3; its knockdown significantly reduced the basal and norepinephrine-induced activity of MCF-7 cells in vitro. The regulation of ADRβ2 signalling by LYPD3 and its metastasis promoting activities, reveal LYPD3 as a promising therapeutic target in the treatment of breast and other cancers.

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Section: Mass Spectrometrymentioning

confidence: 99%

β2-Adrenergic Signalling Promotes Cell Migration by Upregulating Expression of the Metastasis-Associated Molecule LYPD3

Gruet

Cotton

Coveney

et al. 2020

Biology

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“…Hence, combination of autophagy inhibitors (e.g., hydroxy-chloroquine) with epirubicin could prove therapeutically beneficial for (LC3B-II High ) melanoma-suffering patients. Strikingly, besides elevated autophagy, increased levels of the SOX2 (NC) stem cell marker are also associated with anthracycline(s) resistance [ 340 , 349 , 350 , 351 , 352 ]. Given that WM115 cells carry elevated contents of nuclear SOX2 transcription factor ( Figure 4 ), it may be the SOX2 upregulation that critically contributes to primary (e.g., WM115) melanoma cell resistance to low doses of epirubicin ( Figure 9 F).…”

Section: Resultsmentioning

confidence: 99%

From Proteomic Mapping to Invasion-Metastasis-Cascade Systemic Biomarkering and Targeted Drugging of Mutant BRAF-Dependent Human Cutaneous Melanomagenesis

Giannopoulou

Velentzas

Anagnostopoulos

et al. 2021

Cancers

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Melanoma is classified among the most notoriously aggressive human cancers. Despite the recent progress, due to its propensity for metastasis and resistance to therapy, novel biomarkers and oncogenic molecular drivers need to be promptly identified for metastatic melanoma. Hence, by employing nano liquid chromatography-tandem mass spectrometry deep proteomics technology, advanced bioinformatics algorithms, immunofluorescence, western blotting, wound healing protocols, molecular modeling programs, and MTT assays, we comparatively examined the respective proteomic contents of WM115 primary (n = 3955 proteins) and WM266-4 metastatic (n = 6681 proteins) melanoma cells. It proved that WM115 and WM266-4 cells have engaged hybrid epithelial-to-mesenchymal transition/mesenchymal-to-epithelial transition states, with TGF-β controlling their motility in vitro. They are characterized by different signatures of SOX-dependent neural crest-like stemness and distinct architectures of the cytoskeleton network. Multiple signaling pathways have already been activated from the primary melanoma stage, whereas HIF1α, the major hypoxia-inducible factor, can be exclusively observed in metastatic melanoma cells. Invasion-metastasis cascade-specific sub-routines of activated Caspase-3-triggered apoptosis and LC3B-II-dependent constitutive autophagy were also unveiled. Importantly, WM115 and WM266-4 cells exhibited diverse drug response profiles, with epirubicin holding considerable promise as a beneficial drug for metastatic melanoma clinical management. It is the proteome navigation that enables systemic biomarkering and targeted drugging to open new therapeutic windows for advanced disease.

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“…Spheroids were produced as reported previously (Balahmar et al 2018 ). In brief, spheroid cells were grown under normal conditions for 72 h. After 72 h, cells were trypsinized and were seeded (5 × 10 6 cells/per flask) in an ultra-low attachment flask for 3–5 days.…”

Section: Methodsmentioning

confidence: 99%

“…Cell invasion assay was carried out using the BD Falcon ™ BioCoat tumour invasion systems (Corning, Arizona, USA) with fluoroBlock ™ 96 well insert plate according to manufacturer’s guidelines. This was also compared with migration of cells through uncoated BD Falcon™ FluoroBlok ™ 96 well insert plates (see Balahmar et al 2018 ) for details). In this study, spheroid cells were used that have a total diameter above 8.0 μm.…”

Section: Methodsmentioning

confidence: 99%

Doxorubicin resistant choriocarcinoma cell line derived spheroidal cells exhibit stem cell markers but reduced invasion

2022

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Cell cycle-specific cancer chemotherapy is based on the ability of a drug to halt, minimise or destroy rapidly dividing cells. However, their efficacy is limited by the emergence of a self-renewing cell pool called “cancer stem cells” (CSC). Choriocarcinoma is a tumour of trophoblastic tissue. We, in this study, analysed whether spheroids generated from doxorubicin-treated and non-treated choriocarcinoma cell lines exhibit markers of stem cells. Two choriocarcinoma cell lines, namely JEG-3 and BeWo, were used in this study. Spheroids were generated from doxorubicin-treated cells and the non-treated cells under non-adherent condition, followed by analysis of stem-cell markers’ expression, namely NANOG, OCT4 and SOX2. Immunofluorescence analysis suggested a general increase in the markers’ concentration in spheroids relative to the parental cells. RT-qPCR and immunoblots showed an increase in the stem-cell marker expression in spheroids generated from doxorubicin-treated when compared to non-treated cells. In spheroids, Sox2 was significantly upregulated in doxorubicin-treated spheroids, whereas Nanog and Oct4 were generally downregulated when compared to non-treated spheroids. Both 2D and 3D invasion assays showed that the spheroids treated with doxorubicin exhibited reduced invasion. Our data suggest that choriocarcinoma cell lines may have the potential to produce spheroidal cells, yet the drug-treatment affected the invasion potential of spheroids.

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Identification and characterisation of NANOG+/ OCT-4high/SOX2+ doxorubicin-resistant stem-like cells from transformed trophoblastic cell lines

Cited by 11 publications

References 30 publications

β2-Adrenergic Signalling Promotes Cell Migration by Upregulating Expression of the Metastasis-Associated Molecule LYPD3

β2-Adrenergic Signalling Promotes Cell Migration by Upregulating Expression of the Metastasis-Associated Molecule LYPD3

From Proteomic Mapping to Invasion-Metastasis-Cascade Systemic Biomarkering and Targeted Drugging of Mutant BRAF-Dependent Human Cutaneous Melanomagenesis

Doxorubicin resistant choriocarcinoma cell line derived spheroidal cells exhibit stem cell markers but reduced invasion

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