Abstract
The 3D multi-cellular tumoroid (MCT) model is an in vivo-like, avascular tumor model that has received much attention as a refined screening platform for drug therapies. Several types of research have been efforted to improve the physiological characteristics of the tumor microenvironment (TME) of the in vivo-like MCTs. Size-controlled MCTs have received much attention for obtaining highly reproducible results in drug screening assays and achieving a homogeneous and meaningful level of biological activities. Here, we describe an effective method for fabricating the size-controlled in vivo-like MCTs using a cell-loss-free (CLF) microwell arrays. The CLF microwell arrays was fabricated by using the simple operation of laser carving of a poly (methyl methacrylate) (PMMA) master mold. We also demonstrated the biophysicochemical effect of tumor microenvironment (TME) resident fibroblasts through the expression of TGFβ, αSMA, Type I-, IV collagen, angiogenesis related markers on tumorigenesis, and confirmed the drug response of MCTs with anti-cancer agents. This technology for the fabrication of CLF microwell arrays could be used as an effective method to produce an in vitro tumor model for cancer research and drug discovery.
Highlights
Multi-cellular tumoroids (MCTs) have received much attention for the study of a refined screening platform for drug therapies [1, 2]
A549 human lung carcinoma cells, HUVECs human vascular endothelial cells, and MRC-5 human lung fibroblasts which served as cancer resident fibroblasts, were pre-mixed at a ratio of 2:1:2 and plated in the CLF concave microwell array to form multi-cellular tumoroid (MCT) (Fig 1(c), day 0)
By the influence of the CLF architecture, the aspiration step for removing residual cells was omitted, and the most cells were used for MCTs formation
Summary
Multi-cellular tumoroids (MCTs) have received much attention for the study of a refined screening platform for drug therapies [1, 2]. The physiological characteristics of the three-dimensional (3D) MCTs closely resemble avascular tumor nodules, micro-metastases, and inter-vascular regions of large solid tumors [3,4,5]. Conventional two-dimensional (2D) platforms are well established and easy to use for these applications [6]. The absence of 3D cell-cell and cell-matrix interactions can obscure experimental observations and result in misleading and contradictory results during drug screening [7]. The lack of the complex 3D extracellular matrix (ECM) network in monolayer culture can affect drug testing results [7, 8].
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