ArticleMolecular biology reports2026
Optimising the method to develop spheroids from MDA-MB-468 human triple negative breast cancer cells.
Article in Molecular biology reports, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
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Who cites it
2 citing papers in PubMed.
- Three-dimensional spheroid models in breast cancer: tumor microenvironment complexity, cancer stem cell-driven resistance, and translational model integration.Journal of translational medicine · 2026Review
- Advantages and research progress of three-dimensional culture systems for lung cancer drug screening (Review).Oncology letters · 2026Review
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5 authors.
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Abstract
purposeThree-dimensional (3D) spheroid models are increasingly used to emulate the tumour microenvironment for preclinical drug screening. This study aimed to optimise and assess spheroid formation from MDA-MB-468 triple-negative breast cancer (TNBC) cells using hanging drop, liquid overlay, and rigid scaffold methods under normal oxygen (NOC) and low oxygen (LOC) culture conditions.
methodsSpheroids were generated and characterised using bright-field microscopy with AnaSP morphometrics (sphericity, solidity, and perimeter). Gene expression of Epithelial-Mesenchymal Transition (EMT), stemness, and hypoxia/angiogenesis markers (CD44, HIF1A, VEGFA, TWIST1, SNAI1, and NES) was quantified using qPCR. The optimised model was further evaluated using field-emission scanning electron microscopy (FE-SEM) and Hoechst fluorescence.
resultsA workflow combining hanging-drop pre-aggregation with ultra-low attachment (ULA) or agarose-coated plates under NOC produced consistent, compact spheroids. Scaffold cultures formed rapidly but showed size variability under NOC and LOC. Across methods, spheroids were less compact, and gene expression patterns deviated from expected hypoxic responses. HIF1A and VEGFA were more highly expressed under NOC, suggesting pseudo-hypoxic signalling and activation of angiogenesis-related pathways. CD44 and TWIST1 were upregulated in most spheroid types, whereas SNAI1 and NES exhibited condition- and method-specific variability.
conclusionSpheroids cultivated under normoxic conditions demonstrated enhanced structural integrity and transcriptional fidelity. Nonetheless, the study identified that the most compact and resilient spheroids were achieved through the use of hanging-drop pre-aggregation combined with ULA-plates under NOC. The enhanced structural integrity and transcriptional fidelity observed in these spheroids make them valuable models for studying cancer biology and drug responses.
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