Click chemistry has improved the ability to synthesize biomaterial-based scaffolds for culturing cells in 3D to recapitulate the native mechanical and biochemical environment. Matrigel, a protein extract from mouse sarcoma, is a common 3D matrix yet it offers poor physical tunability and batch heterogeneity that reduce drug screening reproducibility. Materials such as HyStem, a hyaluronan (HA)/gelatin/poly(ethylene glycol) (PEG) based thiol-ene chemical hydrogel, permit cellular encapsulation but is relatively unstable to air exposure. A scaffold suitable for the growth of physiologically relevant multicellular cancer spheroids (MCS) for drug screening requires a well-defined and stable 3D matrix that mimics the tumour microenvironment.<br/>We have synthesized an aldehyde and furan functionalized HA biopolymer characterized by <sup>1</sup>H NMR. The bifunctional HA was crosslinked with PEG bis(oxyamine) and maleimide-functionalized extracellular matrix-mimetic peptides with versatile chemical and physical properties. Importantly the stiffness of the bifunctional HA scaffold was not statistically different from Matrigel and HyStem, and the concentration of immobilized adhesive peptides was tunable on the bifunctional matrix. The hydrogel supported the formation of 3D MCS for two human breast cancer epithelial cell lines. These 3D multicellular structures, along with the HA hydrogel, were stable for over four weeks. Cell growth and morphology in the presence of the anticancer drugs, 5-fluorouracil (5-FU) and Doxorubicin (DOX), were compared with 2D tissue culture plastic and 3D Matrigel controls. Our bifunctional HA scaffold using orthogonal click-chemistries may provide a more representative and efficient drug screening method compared with conventional 2D cell culture and xenograft model systems.

Orthogonal oxime and Diels-Alder hyaluronan hydrogels as tunable matrices for multicellular tumour spheroid formation

253rd National Meeting (2017)

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