SARS-CoV-2 has currently precipitated the COVID-19 global health crisis. We developed a medium-throughput drug screening system and identified a small molecule library of 34 of 430 protein kinase inhibitors that were capable of inhibiting SARS-CoV-2 cytopathic effect in human epithelial cells. These drug inhibitors are in various stages of clinical trials. We detected key proteins involved in cellular signaling pathways mTOR-PI3K-AKT, ABL-BCR/MAPK, and DNA-Damage Response that are critical for SARS-CoV-2 infection. A drug-protein interaction based secondary screen confirmed compounds such as the ATR kinase inhibitor berzosertib and torin2 with anti SARS-CoV-2 activity. Berzosertib exhibited potent antiviral activity against SARS-CoV-2 in multiple cell types and blocked replication at post-entry step. Berzosertib inhibited replication of SARS-CoV-1 and MERS-CoV as well. Our study highlights key promising kinase inhibitors to constrain coronavirus replication as a host-directed therapy in the treatment of COVID-19 and beyond as well as provides an important mechanism of host-pathogen interactions.

Target 2035, an international federation of biomedical scientists from the public and private sectors, is leveraging ‘open’ principles to develop a pharmacological tool for every human protein.