CRISISS: A Novel, Transcriptionally and Post-Translationally Inducible CRISPR/Cas9-Based Cellular Suicide Switch

Abstract

With the ever-increasing developing rate of gene and cellular therapy applications and growing accessibility due to products receiving regulatory approval, the need for effective and reliable safety mechanisms to prevent or eliminate potentially fatal side effects is of the utmost importance. In this study, we present the CRISPR-induced suicide switch (CRISISS) as a tool to eliminate genetically modified cells in an inducible and highly efficient manner by targeting Cas9 to highly repetitive Alu retrotransposons in the human genome, causing irreparable genomic fragmentation by the Cas9 nuclease and resulting cell death. The suicide switch components, including expression cassettes for a transcriptionally and post-translationally inducible Cas9 and an Alu-specific single-guide RNA, were integrated into the genome of target cells via Sleeping-Beauty-mediated transposition. The resulting transgenic cells did not show signs of any impact on overall fitness when uninduced, as unintended background expression, background DNA damage response and background cell killing were not observed. When induced, however, a strong expression of Cas9, a strong DNA damage response and a rapid halt of cell proliferation coupled with near complete cell death within four days post-induction were seen. With this proof-of-concept study, we present a novel and promising approach for a robust suicide switch with potential utility for gene and cell therapy in the future.

State-of-the-Art Molecular Genetics and Genomics in Germany

“…Amberger and colleagues bring up another application for CRISPR/Cas9 technology here, which focuses on reducing the cell biological side effects of gene and cell therapies to render these techniques reliably safe [ 4 ]. The authors constructed an inducible cell killing system (“suicide switch”) to reduce unwanted risks such as inflammation, neurotoxicity, and tumorigenic potential of, e.g., grafted genetically engineered cells.…”