The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the COVID-19 pandemic, has given rise to many new variants with increased transmissibility and the ability to evade vaccine protection. The 78-kDa glucose-regulated protein (GRP78) is a major endoplasmic reticulum (ER) chaperone that has been recently implicated as an essential host factor for SARS-CoV-2 entry and infection. In this study, we investigated the efficacy of YUM70, a small molecule inhibitor of GRP78, to block SARS-CoV-2 viral entry and infection in vitro and in vivo. Using human lung epithelial cells and pseudoviral particles carrying spike proteins from different SARS-CoV-2 variants, we found that YUM70 was equally effective at blocking viral entry mediated by original and variant spike proteins. Furthermore, YUM70 reduced SARS-CoV-2 infection without impacting cell viability in vitro and suppressed viral protein production following SARS-CoV-2 infection. Additionally, YUM70 rescued the cell viability of multi-cellular human lung and liver 3D organoids transfected with a SARS-CoV-2 replicon. Importantly, YUM70 treatment ameliorated lung damage in transgenic mice infected with SARS-CoV-2, which correlated with reduced weight loss and longer survival. Thus, GRP78 inhibition may be a promising approach to augment existing therapies to block SARS-CoV-2, its variants, and other viruses that utilize GRP78 for entry and infection.