Exploiting CRISPR Cas9 in Three-Dimensional Stem Cell Cultures to Model Disease

Three-dimensional (3D) cell culture methods have been widely used on a range of cell types, including stem cells to modulate precisely the cellular biophysical and biochemical microenvironment and control various cell signaling cues. As a result, morein vivo-like microenvironments are recapitulated, particularly through the formation of multicellular spheroids and organoids, which may yield more valid mechanisms of disease. Recently, genome-engineering tools such as CRISPR Cas9 have expanded the repertoire of techniques to control gene expression, which complements external signaling cues with intracellular control elements. As a result, the combination of CRISPR Cas9 and 3D cell culture methods enhance our understanding of the molecular mechanisms underpinning several disease phenotypes and may lead to developing new therapeutics that may advance more quickly and effectively into clinical candidates. In addition, using CRISPR Cas9 tools to rescue genes brings us one step closer to its use as a gene therapy tool for various degenerative diseases. Herein, we provide an overview of bridging of CRISPR Cas9 genome editing with 3D spheroid and organoid cell culture to better understand disease progression in both patient and non-patient derived cells, and we address potential remaining gaps that must be overcome to gain widespread use.