Generation of uniform-sized multicellular tumor spheroids using hydrogel microwells for advanced drug screening

Abstract

Even though in vitro co-culture tumor spheroid model plays an important role in screening drug candidates, its wide applications are currently limited due to the lack of reliable and high throughput methods for generating well-defined and 3D complex co-culture structures. Herein, we report the development of a hydrogel microwell array to generate uniform-sized multicellular tumor spheroids. Our developed multicellular tumor spheroids are structurally well-defined, robust and can be easily transferred into the widely used 2D culture substrates while maintaining our designed multicellular 3D-sphere structures. Moreover, to develop effective anti-cancer therapeutics we integrated our recently developed gold-graphene hybrid nanomaterial (Au@GO)-based photothermal cancer therapy into a series of multicellular tumor spheroid co-culture system. The multicellular tumor spheroids were harvested onto a two-dimensional (2D) substrate, under preservation of their three-dimensional (3D) structure, to evaluate the photothermal therapy effectiveness of graphene oxide (GO)-wrapped gold nanoparticles (Au@GO). From the model of co-culture spheroids of HeLa/Ovarian cancer and HeLa/human umbilical vein endothelial cell (HUVEC), we observed that Au@GO nanoparticles displayed selectivity towards the fast-dividing HeLa cells, which could not be observed to this extent in 2D cultures. Overall, our developed uniform-sized 3D multicellular tumor spheroid could be a powerful tool for anticancer drug screening applications.

Characterization of the Anti-Cancer Activity of the Probiotic Bacterium Lactobacillus fermentum Using 2D vs. 3D Culture in Colorectal Cancer Cells

“…As a result, researchers have identified solid tumor growth in 3D, resulting in heterozygous exposure to oxygen as well as physical and chemical stresses [36,37,38,39,40]. Overall, a uniform-sized 3D tumor spheroid platform could be a powerful tool for anti-cancer drug discovery and the development process of such drugs using materials from probiotics such as CFS and bacteria itself [36,41,42].…”

Injectable therapeutic organoids using sacrificial hydrogels

“…Sacrificial materials are widely used in micromachining of microelectromechanical systems (MEMS) to release patterned metals or semiconductors from a substrate (Figure 1a). To cellular structures, some hydrogels (such as agarose or poly(ethylene)glycol as previously demonstrated) can be non-adhesive and thereby promote cells to interact with one another and contract into microtissues and organoids 1,35 . We hypothesized that a dynamic change in the cross-linking state of alginate, which can be achieved by adding calcium or a chelator and has been demonstrated for other purposes 31,36 .…”

Automation, Monitoring, and Standardization of Cell Product Manufacturing

“…Cells spheroids have shown improved osteogenic, adipogenic and chondrogenic potential over conventional culture systems ( Yoon et al, 2012 ; Yamaguchi et al, 2014 ; Cesarz and Tamama, 2016 ; Miyamoto et al, 2017 ; Moritani et al, 2018 ; Tsai et al, 2019 ), improved vascularization in ischemic tissue ( Bhang et al, 2012 ) and constitute the first choice in cancer models and evaluation of anti-cancer drugs ( Chatzinikolaidou, 2016 ; Zanoni et al, 2016 ; Rodrigues et al, 2018 ). Various scaffold-free [e.g., seeding cells in a porous microwell agarose microchip ( Colle et al, 2020 ), seeding cells in 3D printed well inserts ( Boyer et al, 2018 ) or the hanging drop method ( Kapur et al, 2012 )] and scaffold-based [e.g., natural or synthetic hydrogels are used as substrates for spheroids growth ( Murphy et al, 2014 ; Chang et al, 2018 ; Lee et al, 2018 ; Kim et al, 2019 )] have been described in the literature. Regarding scalability, of significant importance are recent studies that describe scaffold-free cell spheroids production using the hanging drop method performed by a robotic device ( Gutzweiler et al, 2017 ) and a robotic automated droplet microfluidic platform ( Langer and Joensson, 2020 ).…”