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Therefore, alternative matrices where chemical composition and materials properties are controlled to steer maturation of HepG2 spheroids remain desired. Herein, a modular approach is taken based on a fully synthetic and minimalistic supramolecular matrix based on squaramide synthons outfitted with a cell\u2010adhesive peptide, RGD for 3D HepG2 spheroid culture. Co\u2010assemblies of RGD\u2010functionalized squaramide\u2010based and native monomers resulted in soft and self\u2010recovering supramolecular hydrogels with a tunable RGD concentration. HepG2 spheroids are self\u2010assembled and grown (\u2248150\u00a0\u00b5m) within the supramolecular hydrogels with high cell viability and differentiation over 21 days of culture. Importantly, significantly higher mRNA and protein expression levels of phase I and II metabolic enzymes, drug transporters, and liver markers are found for the squaramide hydrogels in comparison to Matrigel. Overall, the fully synthetic squaramide hydrogels are proven to be synthetically accessible and effective for HepG2 differentiation showcasing the potential of this supramolecular matrix to rival and replace naturally\u2010derived materials classically used in high\u2010throughput toxicity screening.<\/jats:p>","DOI":"10.1002\/adhm.202001903","type":"journal-article","created":{"date-parts":[[2021,4,30]],"date-time":"2021-04-30T10:43:20Z","timestamp":1619779400000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":34,"title":["Squaramide\u2010Based Supramolecular Materials Drive HepG2 Spheroid Differentiation"],"prefix":"10.1002","volume":"10","author":[{"given":"Tingxian","family":"Liu","sequence":"first","affiliation":[{"name":"Department of Supramolecular and Biomaterials Chemistry Leiden Institute of Chemistry Leiden University  P.O. Box 9502 Leiden 2300 RA Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Linda","family":"van den Berk","sequence":"additional","affiliation":[{"name":"Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research Leiden University  P.O. Box 9502 Leiden 2300 RA Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Joeri A. J.","family":"Wondergem","sequence":"additional","affiliation":[{"name":"Department of Physics Huygens\u2010Kamerlingh Onnes Laboratory Leiden University  Leiden 2300 RA Netherlands"},{"name":"Fraunhofer Institute for Silicate Research ISC  Neunerplatz 2 W\u00fcrzburg 97082 Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ciqing","family":"Tong","sequence":"additional","affiliation":[{"name":"Department of Supramolecular and Biomaterials Chemistry Leiden Institute of Chemistry Leiden University  P.O. Box 9502 Leiden 2300 RA Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Markus C.","family":"Kwakernaak","sequence":"additional","affiliation":[{"name":"Department of Supramolecular and Biomaterials Chemistry Leiden Institute of Chemistry Leiden University  P.O. Box 9502 Leiden 2300 RA Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bas ter","family":"Braak","sequence":"additional","affiliation":[{"name":"Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research Leiden University  P.O. 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