The monoclonal antibody combination REGEN-COV protects against SARS-CoV-2 mutational escape in preclinical and human studies.

Monoclonal antibodies against SARS-CoV-2 are a clinically validated therapeutic option against COVID-19. Because rapidly emerging virus mutants are becoming the next major concern in the fight against the global pandemic, it is imperative that these therapeutic treatments provide coverage against circulating variants and do not contribute to development of treatment-induced emergent resistance. To this end, we investigated the sequence diversity of the spike protein and monitored emergence of virus variants in SARS-COV-2 isolates found in COVID-19 patients treated with the two-antibody combination REGEN-COV, as well as in preclinical in vitro studies using single, dual, or triple antibody combinations, and in hamster in vivo studies using REGEN-COV or single monoclonal antibody treatments. Our study demonstrates that the combination of non-competing antibodies in REGEN-COV provides protection against all current SARS-CoV-2 variants of concern/interest and also protects against emergence of new variants and their potential seeding into the population in a clinical setting

Visualizing Dynamic E2F-Mediated Repression In Vivo

Although many E2F target genes have been identified recently, very little is known about how any single E2F site controls the expression of an E2F target gene in vivo. To test the requirement for a single E2F site in vivo and to learn how E2F-mediated repression is regulated during development and tumorigenesis, we have constructed a novel series of wild-type and mutant Rb promoter-LacZ transgenic reporter lines that allow us to visualize the activity of a crucial E2F target in vivo, the retinoblastoma tumor suppressor gene (Rb). Two mutant Rb promoter-LacZ constructs were used to evaluate the importance of a single E2F site or a nearby activator (Sp1/Ets) site that is found mutated in low-penetrance retinoblastomas. The activity of the wild-type Rb promoter is dynamic, varying spatially and temporally within the developing nervous system. While loss of the activator site silences the Rb promoter, loss of the E2F site stimulates its activity in the neocortex, retina, and trigeminal ganglion. Surprisingly, E2F-mediated repression of Rb does not act globally or in a static manner but, instead, is a highly dynamic process in vivo. Using neocortical extracts, we detected GA-binding protein α (GABPα, an Ets family member) bound to the activator site and both E2F1 and E2F4 bound to the repressor site of the Rb promoter in vitro. Additionally, we detected binding of both E2F1 and E2F4 to the Rb promoter in vivo using chromatin immunoprecipitation analysis on embryonic day 13.5 brain. Unexpectedly, we detect no evidence for Rb promoter autoregulation in neuroendocrine tumors from Rb(+/−); RbP-LacZ mice that undergo loss of heterozygosity at the Rb locus, in contrast to the situation in human retinoblastomas where high RB mRNA levels are found. In summary, this study provides the first demonstration that loss of an E2F site is critical for target gene repression in vivo and underscores the complexity of the Rb and E2F family network in vivo

Antibody cocktail to SARS-CoV-2 spike protein prevents rapid mutational escape seen with individual antibodies.

Antibodies targeting the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) present a promising approach to combat the coronavirus disease 2019 (COVID-19) pandemic; however, concerns remain that mutations can yield antibody resistance. We investigated the development of resistance against four antibodies to the spike protein that potently neutralize SARS-CoV-2, individually as well as when combined into cocktails. These antibodies remain effective against spike variants that have arisen in the human population. However, novel spike mutants rapidly appeared after in vitro passaging in the presence of individual antibodies, resulting in loss of neutralization; such escape also occurred with combinations of antibodies binding diverse but overlapping regions of the spike protein. Escape mutants were not generated after treatment with a noncompeting antibody cocktail