BackgroundRecent data suggest that complications and death from coronavirus disease 2019 (Covid-19) may be related to high viral loads.MethodsIn this ongoing, double-blind, phase 1–3 trial involving nonhospitalized patients with Covid-19, we investigated two fully human, neutralizing monoclonal antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein, used in a combined cocktail (REGN-COV2) to reduce the risk of the emergence of treatment-resistant mutant virus. Patients were randomly assigned (1:1:1) to receive placebo, 2.4 g of REGN-COV2, or 8.0 g of REGN-COV2 and were prospectively characterized at baseline for endogenous immune response against SARS-CoV-2 (serum antibody–positive or serum antibody–negative). Key end points included the time-weighted average change in viral load from baseline (day 1) through day 7 and the percentage of patients with at least one Covid-19–related medically attended visit through day 29. Safety was assessed in all patients.ResultsData from 275 patients are reported. The least-squares mean difference (combined REGN-COV2 dose groups vs. placebo group) in the time-weighted average change in viral load from day 1 through day 7 was −0.56 log10 copies per milliliter (95% confidence interval [CI], −1.02 to −0.11) among patients who were serum antibody–negative at baseline and −0.41 log10 copies per milliliter (95% CI, −0.71 to −0.10) in the overall trial population. In the overall trial population, 6% of the patients in the placebo group and 3% of the patients in the combined REGN-COV2 dose groups reported at least one medically attended visit; among patients who were serum antibody–negative at baseline, the corresponding percentages were 15% and 6% (difference, −9 percentage points; 95% CI, −29 to 11). The percentages of patients with hypersensitivity reactions, infusion-related reactions, and other adverse events were similar in the combined REGN-COV2 dose groups and the placebo group.ConclusionsIn this interim analysis, the REGN-COV2 antibody cocktail reduced viral load, with a greater effect in patients whose immune response had not yet been initiated or who had a high viral load at baseline. Safety outcomes were similar in the combined REGN-COV2 dose groups and the placebo group. (Funded by Regeneron Pharmaceuticals and the Biomedical and Advanced Research and Development Authority of the Department of Health and Human Services; ClinicalTrials.gov number, NCT04425629.)
Christos Kyratsous, Vice President of Research, Infectious Diseases, and Viral Vector Technologies at Regeneron Pharmaceuticals, and Alina Baum, Associate Director, Infectious Diseases Associate at Regeneron Pharmaceuticals, discuss the development of antibody therapeutics targeting the spike protein of SARS-CoV-2.
An urgent global quest for effective therapies to prevent and treat coronavirus disease 2019 (COVID-19) is ongoing. We previously described REGN-COV2, a cocktail of two potent neutralizing antibodies (REGN10987 and REGN10933) that targets nonoverlapping epitopes on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein. In this report, we evaluate the in vivo efficacy of this antibody cocktail in both rhesus macaques, which may model mild disease, and golden hamsters, which may model more severe disease. We demonstrate that REGN-COV-2 can greatly reduce virus load in the lower and upper airways and decrease virus-induced pathological sequelae when administered prophylactically or therapeutically in rhesus macaques. Similarly, administration in hamsters limits weight loss and decreases lung titers and evidence of pneumonia in the lungs. Our results provide evidence of the therapeutic potential of this antibody cocktail.
Casirivimab and imdevimab (REGEN-COV™) markedly reduces risk of hospitalization or death in high-risk individuals with Covid-19. Here we explore the possibility that subcutaneous REGEN-COV prevents SARS-CoV-2 infection and subsequent Covid-19 in individuals at high risk of contracting SARS-CoV-2 by close exposure in a household with a documented SARS-CoV-2-infected individual. Individuals ≥12 years were enrolled within 96 hours of a household contact being diagnosed with SARS-CoV-2 and randomized 1:1 to receive 1200 mg REGEN-COV or placebo via subcutaneous injection. The primary efficacy endpoint was the proportion of participants without evidence of infection (SARS-CoV-2 RT-qPCR-negative) or prior immunity (seronegative) who subsequently developed symptomatic SARS-CoV-2 infection during a 28-day efficacy assessment period. Subcutaneous REGEN-COV significantly prevented symptomatic SARS-CoV-2 infection compared with placebo (81.4% risk reduction; 11/753 [1.5%] vs. 59/752 [7.8%], respectively; P<0.0001), with 92.6% risk reduction after the first week (2/753 [0.3%] vs. 27/752 [3.6%], respectively). REGEN-COV also prevented overall infections, either symptomatic or asymptomatic (66.4% risk reduction). Among infected participants, the median time to resolution of symptoms was 2 weeks shorter with REGEN-COV vs. placebo (1.2 vs. 3.2 weeks, respectively), and the duration of time with high viral load (>104 copies/mL) was lower (0.4 vs. 1.3 weeks, respectively). REGEN-COV was generally well tolerated. Administration of subcutaneous REGEN-COV prevented symptomatic Covid-19 and asymptomatic SARS-CoV-2 infection in uninfected household contacts of infected individuals. Among individuals who became infected, REGEN-COV reduced the duration of symptomatic disease, decreased maximal viral load, and reduced the duration of detectable virus.(ClinicalTrials.gov number, NCT04452318.).
ABSTRACT Neutralizing antibodies are key determinants of protection from future infection, yet well-validated high-throughput assays for measuring titers of SARS-CoV-2-neutralizing antibodies are not generally available. Here we describe the development and validation of IMMUNO-COV™ v2.0 a scalable surrogate virus assay, which titrates antibodies that block infection of Vero-ACE2 cells by a luciferase-encoding vesicular stomatitis virus displaying SARS-CoV-2 spike glycoproteins (VSV-SARS2-Fluc). Antibody titers, calculated using a standard curve consisting of stepped concentrations of SARS-CoV-2 spike monoclonal antibody, correlated closely (p < 0.0001) with titers obtained from a gold-standard PRNT50% assay performed using a clinical isolate of SARS-CoV-2. IMMUNO-COV™ v2.0 was comprehensively validated using data acquired from 242 assay runs performed over seven days by five analysts, utilizing two separate virus lots, and 176 blood samples. Assay performance was acceptable for clinical use in human serum and plasma based on parameters including linearity, dynamic range, limit of blank and limit of detection, dilutional linearity and parallelism, precision, clinical agreement, matrix equivalence, clinical specificity and sensitivity, and robustness. Sufficient VSV-SARS2-Fluc virus reagent has been banked to test 5 million clinical samples. Notably, a significant drop in IMMUNO-COV™ v2.0 neutralizing antibody titers was observed over a six-month period in people recovered from SARS-CoV-2 infection. Together, our results demonstrate the feasibility and utility of IMMUNO-COV™ v2.0 for measuring SARS-CoV-2-neutralizing antibodies in vaccinated individuals and those recovering from natural infections. Such monitoring can be used to better understand what levels of neutralizing antibodies are required for protection from SARS-CoV-2, and what booster dosing schedules are needed to sustain vaccine-induced immunity.
The SARS-CoV-2 spike (S) protein variant D614G supplanted the ancestral virus worldwide in a matter of months. Here we show that D614G was more infectious than the ancestral form on human lung cells, colon cells, and cells rendered permissive by ectopic expression of various mammalian ACE2 orthologs. Nonetheless, D614G affinity for ACE2 was reduced due to a faster dissociation rate. Assessment of the S protein trimer by cryo-electron microscopy showed that D614G disrupts a critical interprotomer contact and that this dramatically shifts the S protein trimer conformation toward an ACE2-binding and fusion-competent state. Consistent with the more open conformation, neutralization potency of antibodies targeting the S protein receptor-binding domain was not attenuated. These results indicate that D614G adopts conformations that make virion membrane fusion with the target cell membrane more probable but that D614G retains susceptibility to therapies that disrupt interaction of the SARS-CoV-2 S protein with the ACE2 receptor.
There is a need for novel immunotherapies to address patient populations resistant or refractory to immune checkpoint inhibitors (CPI). VV1 is an oncolytic vesicular stomatitis virus engineered to express interferon beta (IFNβ) to enhance anti-tumor immune responses and tumor selectivity. Phase 1 studies demonstrated VV1 anti-tumor activity in certain clinical settings either alone or in combination with a CPI. The current preclinical study aimed to identify dosing and scheduling regimens that maximize the efficacy of VV1 in combination with CPIs.
Methods
Immune-competent mice bearing syngeneic tumor models MC38 (colorectal), B16F10 (melanoma) and CMT64 (lung adenocarcinoma) tumors were dosed to test combinations of VV1 delivered intravenously with anti-PD-1 and anti-CTLA4 antibodies. Treatment was started once tumors were established, and T cell responses in the tumor and peripheral lymphoid organs were analyzed using flow cytometry and ELISPOT assays.
Results
Potent anti-tumor efficacy was observed following intratumoral or intravenous administration of VV1 combined with anti-CTLA4 and anti-PD-1. A comparison of single dose versus repeat administration of anti-CTLA4 in combination with VV1 plus continuous anti-PD-1 showed that a single dose was sufficient to maximally enhance the depth and durability of tumor response. This effect was observed consistently in multiple tumor models, including anti-PD-1 sensitive (MC38) or anti-PD-1 resistant tumor models (B16F10 and CMT64). Remarkably, the triple combination boosted T cell priming against the B16F10 and CMT64 neo-antigen peptides in TILs and tumor-draining lymph nodes. ELISPOT and multimer staining showed that, in contrast to doublet therapy using virus and a single CPI, triplet combination therapy strongly boosted CTL responses against a broad array of B16F10 and CMT64 neoepitopes, detected both in the tumor and in tumor-draining lymph nodes. This was associated with increased infiltration of CD8 T cells in the tumor, but the number of regulatory T cells was not impacted, indicating that the enhancing effect of anti-CTLA4 was not a consequence of Treg depletion.
Conclusions
Intratumoral or intravenous VV1 virotherapy combined with anti-CTLA4 and anti-PD-1 checkpoint antibodies synergistically enhances tumor control in multiple syngeneic mouse tumor models. The triple combination seems to promote a tumor-vaccination effect, by inducing a polyclonal anti-tumor T cell response and boosting anti-tumor CTL responses. This triplet combination approach will soon be evaluated clinically in patients with advanced melanoma (after progression on an anti-PD1) and first line NSCLC patients. (ClinicalTrials.gov Identifier: NCT04291105).
