A Drug Screening Platform for Protein Expression Levels in Neurological Disorders

Summary Neurological and psychiatric diseases and disorders affect more than half of the population. Many of these diseases are caused by the malfunctioning of protein synthesis, where too little or too much production of a protein harms a cell and its functions within the brain. We developed a drug screening platform to identify compounds that target the primary cause of these diseases, namely protein expression amounts. This cellular assay monitors protein expression of a target disease gene along with the protein expression of a control gene using the Protein Quantitation Ratioing (PQR) technique. PQR tracks protein concentration using fluorescence. We used human cells and CRISPR-Cas9 genome editing to insert the Protein Quantitation Reporter into target genes. These cells are used in high-throughput drug screening measuring the fluorescence as the assay. Drug hits can be validated using the same PQR technique or animal models of the disease. Highlights The assay can identify drugs that directly address the molecular cause of a disease. The Protein Quantitation Ratioing (PQR) technique allows for tracking and measuring protein amounts over time in single living cells before, during, and after drug administration. Genome editing to insert the PQR into the target gene allows tracking of endogenous protein expression. Using human cell lines allow for faster production of knock-in cells. Patient mutations can be replicated using genome editing during the knock-in step. Using induced pluripotent stem cells allow for an unlimited supply of genome edited differentiated cells such as neurons with the PQR knock-in reporter.