Engineering a vascularised 3D in vitro model of cancer progression

Abstract

The hallmark of tumours is the ability of cancerous cells to promote vascular growth, to disseminate and invade to distant organs. The metastatic process is heavily influenced by the extracellular matrix (ECM) density and composition of the surrounding tumour microenvironment. These microenvironmental cues, which include hypoxia, also regulate the angiogenic processes within a tumour, facilitating the spread of cancer cells. We engineered compartmentalized biomimetic colorectal tumouroids with stromal surrounds that comprised a range of ECM densities, composition and stromal cell populations. Recapitulating tissue ECM composition and stromal cell composition enhanced cancer cell invasion. Manipulation of ECM density was associated with an altered migration pattern from glandular buds (cellular aggregates) to epithelial cell sheets. Laminin appeared to be a critical component in regulating endothelial cell morphology and vascular network formation. Interestingly, the disruption of vascular networks by cancer cells was driven by changes in expression of several anti-angiogenic genes. Cancer cells cultured in our biomimetic tumouroids exhibited intratumoural heterogeneity that was associated with increased tumour invasion into the stroma. These findings demonstrate that our 3D in vitro tumour model exhibits biomimetic attributes that may permit their use in studying microenvironment clues of tumour progression and angiogenesis.

Cancer Associated Fibroblasts Mediate Cancer Progression and Remodel the Tumouroid Stroma

“…The majority of 3D cancer models lack appropriate tensile force and stiffness associated with tumour tissue, as they commonly use soft hydrogels, which have too high a water content 42 . Our biomimetic 3D in vitro cancer model (tumouroid) has a collagen density of up to 40x higher compared to standard hydrogels and therefore mimics the in vivo stiff tumour environment more closely 22 ; an important aspect especially for CAFs 43 . Our tumouroids let us investigate invasion patterns and study the genes that may be responsible for the increase in invasion.…”

“…The gel mix was polymerised at 37˚C for 15 minutes (min), followed by plastic-compression using the 96-well RAFT TM absorbers (Lonza, Slough, UK). In order to produce 'tumouroids' 22 , the ACMs were nested into a stroma. For the stroma, collagen solution as described above was prepared, and ACMs were directly embedded into a 24-well plate (Corning® Costar® through Sigma-Aldrich, Dorset, UK) containing 1.3 mL of the non-crosslinked collagen mix.…”

Photodynamic therapy in 3D cancer models and the utilisation of nanodelivery systems

“…Incorporation of a vascular network is still in its infancy and remains a longer term goal. 120 Numerous studies have investigated the properties of nanoparticles to improve the therapeutic effects obtained from PDT treatment of various cell types in 3D models. Table 2 provides examples of nanoparticles used in PDT studies that have been carried out in 3D cancer models.…”

Multi-Fluorescence High-Resolution Episcopic Microscopy (MF-HREM) for Three-Dimensional Imaging of Adult Murine Organs

“…For the vascular channel, Figure 7D, E and F show the MF-HREM image processing pipeline, with the final vessel segmentation and skeletonisation being carried out using the APP2 algorithm from the Vaa3D neuron tracing plugin (40–42,45). The chaotic nature of the vasculature can be seen from this analysis and such vascular networks can be used in simulations of drug delivery (8) and for understand tumour vessel growth mechanism (46).…”