Convalescent plasma in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial (preprint)

Background: Treatment of COVID-19 patients with plasma containing anti-SARS-CoV-2 antibodies may have a beneficial effect on clinical outcomes. We aimed to evaluate the safety and efficacy of convalescent plasma in patients admitted to hospital with COVID-19. Methods: In this randomised, controlled, open-label, platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]) several possible treatments are being compared with usual care in patients hospitalised with COVID-19 in the UK. Eligible and consenting patients were randomly allocated to receive either usual care plus high titre convalescent plasma or usual care alone. The primary outcome was 28-day mortality. Findings: Between 28 May 2020 and 15 January 2021, 5795 patients were randomly allocated to receive convalescent plasma and 5763 to usual care alone. There was no significant difference in 28-day mortality between the two groups: 1398 (24%) of 5795 patients allocated convalescent plasma and 1408 (24%) of 5763 patients allocated usual care died within 28 days (rate ratio [RR] 1.00; 95% confidence interval [CI] 0.93 to 1.07; p=0.93). The 28-day mortality rate ratio was similar in all prespecified subgroups of patients, including in those patients without detectable SARS-CoV-2 antibodies at randomisation. Allocation to convalescent plasma had no significant effect on the proportion of patients discharged from hospital within 28 days (66% vs. 67%; rate ratio 0.98; 95% CI 0.94-1.03, p=0.50). Among those not on invasive mechanical ventilation at baseline, there was no significant difference in the proportion meeting the composite endpoint of progression to invasive mechanical ventilation or death (28% vs. 29%; rate ratio 0.99; 95% CI 0.93-1.05, p=0.79). Interpretation: Among patients hospitalised with COVID-19, high-titre convalescent plasma did not improve survival or other prespecified clinical outcomes.

Virological and serological characterization of critically ill patient with COVID-19 in the UK: a special focus on variant detection (preprint)

Background. Treatment of COVID-19 patients with convalescent plasma containing neutralising antibody to SARS-CoV-2 is under investigation as a means of reducing viral loads, ameliorating disease outcomes, and reducing mortality. However, its efficacy might be reduced in those infected with the emerging B.1.1.7 SARS-CoV-2 variant. Here, we report the diverse virological characteristics of UK patients enrolled in the Immunoglobulin Domain of the REMAP-CAP randomised controlled trial. Methods. SARS-CoV-2 viral RNA was detected and quantified by real-time PCR in nasopharyngeal swabs obtained from study subjects within 48 hours of admission to intensive care unit. Antibody status was determined by spike-protein ELISA. B.1.1.7 strain was differentiated from other SARS-CoV-2 strains by two novel typing methods detecting the B.1.1.7-associated D1118H mutation with allele-specific probes and by restriction site polymorphism (SfcI). Findings. Of 1260 subjects, 90% were PCR-positive with viral loads in nasopharyngeal swabs ranging from 72 international units [IUs]/ml to 1.7x10^11 IU/ml. Median viral loads were 45-fold higher in those who were seronegative for IgG antibodies (n=314; 28%) compared to seropositives (n=804; 72%), reflecting in part the latter group's possible later disease stage on enrolment. Frequencies of B.1.1.7 infection increased from early November (<1%) to December 2020 (>60%). Anti-SARS-CoV-2 seronegative individuals infected with wild-type SARS-CoV-2 had significantly higher viral loads than seropositives (medians of 1.2x10^6 and 3.4 x10^4 IU/ml respectively; p=2x10^-9). However, viral load distributions were elevated in both seropositive and seronegative subjects infected with B.1.1.7 (13.4x10^6 and 7.6x10^6 IU/ml; p=0.18). Interpretation. High viral loads in seropositive B.1.1.7-infected subjects are consistent with increased replication capacity and/or less effective clearance by innate or adaptive immune response of B.1.1.7 strain than wild-type. As viral genotype was associated with diverse virological and immunological phenotypes, metrics of viral load, antibody status and infecting strain should be used to define subgroups for analysis of treatment efficacy.

Neutralising antibodies drive Spike mediated SARS-CoV-2 evasion (preprint)

SARS-CoV-2 Spike protein is critical for virus infection via engagement of ACE2, and amino acid variation in Spike is increasingly appreciated. Given both vaccines and therapeutics are designed around Wuhan-1 Spike, this raises the theoretical possibility of virus escape, particularly in immunocompromised individuals where prolonged viral replication occurs. Here we report chronic SARS-CoV-2 with reduced sensitivity to neutralising antibodies in an immune suppressed individual treated with convalescent plasma, generating whole genome ultradeep sequences by both short and long read technologies over 23 time points spanning 101 days. Although little change was observed in the overall viral population structure following two courses of remdesivir over the first 57 days, N501Y in Spike was transiently detected at day 55 and V157L in RdRp emerged. However, following convalescent plasma we observed large, dynamic virus population shifts, with the emergence of a dominant viral strain bearing D796H in S2 and{Delta} H69/{Delta}V70 in the S1 N-terminal domain NTD of the Spike protein. As passively transferred serum antibodies diminished, viruses with the escape genotype diminished in frequency, before returning during a final, unsuccessful course of convalescent plasma. In vitro, the Spike escape double mutant bearing{Delta} H69/{Delta}V70 and D796H conferred decreased sensitivity to convalescent plasma, whilst maintaining infectivity similar to wild type. D796H appeared to be the main contributor to decreased susceptibility, but incurred an infectivity defect. The{Delta} H69/{Delta}V70 single mutant had two-fold higher infectivity compared to wild type and appeared to compensate for the reduced infectivity of D796H. Consistent with the observed mutations being outside the RBD, monoclonal antibodies targeting the RBD were not impacted by either or both mutations, but a non RBD binding monoclonal antibody was less potent against{Delta} H69/{Delta}V70 and the double mutant. These data reveal strong selection on SARS-CoV-2 during convalescent plasma therapy associated with emergence of viral variants with reduced susceptibility to neutralising antibodies.

Selectively caring for the most severe COVID-19 patients delays ICU bed shortages more than increasing hospital capacity (preprint)

1SARS-CoV-2, the virus responsible for COVID-19, has killed hundreds of thousands of Americans. Although physical distancing measures played a key role in slowing COVID-19 spread in early 2020, infection rates are now peaking at record levels across the country. Hospitals in several states are threatened with overwhelming numbers of patients, compounding the death toll of COVID-19. Implementing strategies to minimize COVID-19 hospitalizations will be key to controlling the toll of the disease, but non-physical distancing strategies receive relatively little attention. We present a novel system of differential equations designed to predict the relative effects of hospitalizing fewer COVID-19 patients vs increasing ICU bed availability on delaying ICU bed shortages. This model, which we call SEAHIRD, was calibrated to mortality data on two US states with different peak infection times from mid-March - mid-May 2020. It found that hospitalizing fewer COVID-19 patients generally delays ICU bed shortage more than a comparable increase in ICU bed availability. This trend was consistent across both states and across wide ranges of initial conditions and parameter values. We argue that being able to predict which patients will develop severe COVID-19 symptoms, and thus require hospitalization, should be a key objective of future COVID-19 research, as it will allow limited hospital resources to be allocated to individuals that need them most and prevent hospitals from being overwhelmed by COVID-19 cases.

A haemagglutination test for rapid detection of antibodies to SARS-CoV-2 (preprint)

Serological detection of antibodies to SARS-CoV-2 is essential for establishing rates of seroconversion in populations, detection of seroconversion after vaccination, and for seeking evidence for a level of antibody that may be protective against COVID-19 disease. Several high-performance commercial tests have been described, but these require centralised laboratory facilities that are comparatively expensive, and therefore not available universally. Red cell agglutination tests have a long history in blood typing, and general serology through linkage of reporter molecules to the red cell surface. They do not require special equipment, are read by eye, have short development times, low cost and can be applied as a Point of Care Test (POCT). We describe a red cell agglutination test for the detection of antibodies to the SARS-CoV-2 receptor binding domain (RBD). We show that the Haemagglutination Test (HAT) has a sensitivity of 90% and specificity of 99% for detection of antibodies after a PCR diagnosed infection. The HAT can be titrated, detects rising titres in the first five days of hospital admission, correlates well with a commercial test that detects antibodies to the RBD, and can be applied as a point of care test. The developing reagent is composed of a previously described nanobody to a conserved glycophorin A epitope on red cells, linked to the RBD from SARS-CoV-2. It can be lyophilised for ease of shipping. We have scaled up production of this reagent to one gram, which is sufficient for ten million tests, at a cost of ~0.27 UK pence per test well. Aliquots of this reagent are ready to be supplied to qualified groups anywhere in the world that need to detect antibodies to SARS-CoV-2, but do not have the facilities for high throughput commercial tests.

Convalescent plasma therapy for the treatment of patients with COVID-19: Assessment of methods available for antibody detection and their correlation with neutralising antibody levels (preprint)

Introduction. The lack of approved specific therapeutic agents to treat COVID-19 associated with SARS coronavirus 2 (SARS-CoV-2) infection has led to the rapid implementation and/or randomised controlled trials of convalescent plasma therapy (CPT) in many countries including the UK. Effective CPT is likely to require high titres of neutralising antibody levels in convalescent donations. Understanding the relationship between functional neutralising antibodies and antibody levels to specific SARS-CoV-2 proteins in scalable assays will be crucial for the success of large-scale collection and use of convalescent plasma. We assessed whether neutralising antibody titres correlated with reactivity in a range of ELISA assays targeting the spike (S) protein, the main target for human immune response. Methods. Blood samples were collected from 52 individuals with a previous laboratory confirmed SARS-CoV-2 infection at least 28 days after symptom resolution. These were assayed for SARS-CoV-2 neutralising antibodies by microneutralisation and pseudotype assays, and for antibodies by four different ELISAs. ROC analysis was used to further identify sensitivity and specificity of selected assays to identify samples containing high neutralising antibody levels suitable for clinical use of convalescent plasma. Results. All samples contained SARS-CoV-2 antibodies, whereas neutralising antibody titres of greater than 1:20 were detected in 43 samples (83% of those tested) and >1:100 in 22 samples (42%). The best correlations were observed with EUROimmun IgG ELISA S/CO reactivity (Spearman Rho correlation co-efficient 0.88; p<0.001). Based on ROC analysis, EUROimmun would detect 60% of samples with titres of >1:100 with 100% specificity using a reactivity index of 9.1 (13/22). Discussion. Robust associations between virus neutralising antibody titres and reactivity in several ELISA-based antibody tests demonstrate their possible utility for scaled-up production of convalescent plasma containing potentially therapeutic levels of anti-SARS-CoV-2 neutralising antibodies.