ABSTRACT
A safe and effective vaccine against COVID-19 is urgently needed in quantities that are sufficient to immunize large populations. Here we report the preclinical development of two vaccine candidates (BNT162b1 and BNT162b2) that contain nucleoside-modified messenger RNA that encodes immunogens derived from the spike glycoprotein (S) of SARS-CoV-2, formulated in lipid nanoparticles. BNT162b1 encodes a soluble, secreted trimerized receptor-binding domain (known as the RBD-foldon). BNT162b2 encodes the full-length transmembrane S glycoprotein, locked in its prefusion conformation by the substitution of two residues with proline (S(K986P/V987P); hereafter, S(P2) (also known as P2 S)). The flexibly tethered RBDs of the RBD-foldon bind to human ACE2 with high avidity. Approximately 20% of the S(P2) trimers are in the two-RBD 'down', one-RBD 'up' state. In mice, one intramuscular dose of either candidate vaccine elicits a dose-dependent antibody response with high virus-entry inhibition titres and strong T-helper-1 CD4+ and IFNγ+CD8+ T cell responses. Prime-boost vaccination of rhesus macaques (Macaca mulatta) with the BNT162b candidates elicits SARS-CoV-2-neutralizing geometric mean titres that are 8.2-18.2× that of a panel of SARS-CoV-2-convalescent human sera. The vaccine candidates protect macaques against challenge with SARS-CoV-2; in particular, BNT162b2 protects the lower respiratory tract against the presence of viral RNA and shows no evidence of disease enhancement. Both candidates are being evaluated in phase I trials in Germany and the USA1-3, and BNT162b2 is being evaluated in an ongoing global phase II/III trial (NCT04380701 and NCT04368728).
Subject(s)
COVID-19 Vaccines/immunology , COVID-19/immunology , COVID-19/prevention & control , Disease Models, Animal , SARS-CoV-2/immunology , Aging/immunology , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Antigens, Viral/chemistry , Antigens, Viral/genetics , Antigens, Viral/immunology , BNT162 Vaccine , COVID-19/blood , COVID-19/therapy , COVID-19/virology , COVID-19 Vaccines/administration & dosage , COVID-19 Vaccines/chemistry , COVID-19 Vaccines/genetics , Cell Line , Clinical Trials as Topic , Female , Humans , Immunization, Passive , Internationality , Macaca mulatta/immunology , Macaca mulatta/virology , Male , Mice , Mice, Inbred BALB C , Models, Molecular , Protein Multimerization , RNA, Viral/analysis , Respiratory System/immunology , Respiratory System/virology , SARS-CoV-2/chemistry , SARS-CoV-2/genetics , Solubility , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/immunology , Vaccination , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/chemistry , Vaccines, Synthetic/genetics , Vaccines, Synthetic/immunology , COVID-19 Serotherapy , mRNA VaccinesABSTRACT
Increased pro-inflammatory signaling is a hallmark of metabolic dysfunction in obesity and diabetes. Although both inflammatory and energy substrate handling processes represent critical layers of metabolic control, their molecular integration sites remain largely unknown. Here, we identify the heterodimerization interface between the α and ß subunits of transcription factor GA-binding protein (GAbp) as a negative target of tumor necrosis factor alpha (TNF-α) signaling. TNF-α prevented GAbpα and ß complex formation via reactive oxygen species (ROS), leading to the non-energy-dependent transcriptional inactivation of AMP-activated kinase (AMPK) ß1, which was identified as a direct hepatic GAbp target. Impairment of AMPKß1, in turn, elevated downstream cellular cholesterol biosynthesis, and hepatocyte-specific ablation of GAbpα induced systemic hypercholesterolemia and early macro-vascular lesion formation in mice. As GAbpα and AMPKß1 levels were also found to correlate in obese human patients, the ROS-GAbp-AMPK pathway may represent a key component of a hepato-vascular axis in diabetic long-term complications.
Subject(s)
Atherosclerosis/metabolism , GA-Binding Protein Transcription Factor/metabolism , Hepatocytes/metabolism , Hypercholesterolemia/metabolism , Protein Kinases/metabolism , Signal Transduction , AMP-Activated Protein Kinase Kinases , Animals , Atherosclerosis/etiology , Atherosclerosis/pathology , Cell Line , Cells, Cultured , Cholesterol/metabolism , GA-Binding Protein Transcription Factor/chemistry , Hypercholesterolemia/complications , Male , Mice , Mice, Inbred C57BL , Protein Multimerization , Protein Subunits/chemistry , Protein Subunits/metabolism , Reactive Oxygen Species/metabolism , Tumor Necrosis Factor-alpha/metabolismABSTRACT
In mammals, glucocorticoids (GCs) and their intracellular receptor, the glucocorticoid receptor (GR), represent critical checkpoints in the endocrine control of energy homeostasis. Indeed, aberrant GC action is linked to severe metabolic stress conditions as seen in Cushing's syndrome, GC therapy and certain components of the Metabolic Syndrome, including obesity and insulin resistance. Here, we identify the hepatic induction of the mammalian conserved microRNA (miR)-379/410 genomic cluster as a key component of GC/GR-driven metabolic dysfunction. Particularly, miR-379 was up-regulated in mouse models of hyperglucocorticoidemia and obesity as well as human liver in a GC/GR-dependent manner. Hepatocyte-specific silencing of miR-379 substantially reduced circulating very-low-density lipoprotein (VLDL)-associated triglyceride (TG) levels in healthy mice and normalized aberrant lipid profiles in metabolically challenged animals, mediated through miR-379 effects on key receptors in hepatic TG re-uptake. As hepatic miR-379 levels were also correlated with GC and TG levels in human obese patients, the identification of a GC/GR-controlled miRNA cluster not only defines a novel layer of hormone-dependent metabolic control but also paves the way to alternative miRNA-based therapeutic approaches in metabolic dysfunction.