key: cord-0825172-3229q816
authors: Libster, R.; Perez Marc, G.; Wappner, D.; Coviello, S.; Bianchi, A.; Braem, V.; Esteban, I.; Caballero, M. T.; Wood, C. J.; Berrueta, M.; Rondan, A.; Lescano, G.; Cruz, P.; Ritou, I.; Fernandez Vina, V.; Alvarez Paggi, D.; Esperante, S.; Ferretti, A.; Ofman, G.; Ciganda, A.; Rodriguez, R.; Lantos, J.; Valentini, R.; Itcovici, N.; Hintze, A.; Oyarvide, L.; Etchegaray, C.; Neira, A.; Name, I.; Alfonso, J.; Lopez Castelo, R.; Caruso, G.; Rapelius, S.; Alvez, F.; Etchenique, F. C.; Dimase, F.; Alvarez, D. R.; Aranda, S. S.; Sanchez Yanotti, C.; DeLuca, J.; Jarez Baglivo, S.; Laudanno, S. L.; Nowog,
title: Prevention of severe COVID-19 in the elderly by early high-titer plasma
date: 2020-11-21
journal: nan
DOI: 10.1101/2020.11.20.20234013
sha: e6075e75b46a2d30736977bf23be8d93c9022181
doc_id: 825172
cord_uid: 3229q816

Background. Therapies to interrupt progression of early COVID-19 remain elusive. Among them, convalescent plasma in hospitalized patients was unsuccessful, perhaps because antibody should be administered earlier. We advanced plasma infusions to the first 72 hours of symptoms to arrest COVID-19 progression. Methods. A randomized, double-blind, placebo-controlled trial of convalescent plasma with high IgG titers against SARS-CoV2 in elderly subjects within 72 hours of mild COVID-19 symptoms. The primary endpoint was severe respiratory disease defined as a respiratory rate [≥]30 and/or an O2 sat<93% in room air. The study was interrupted at 76% of its projected sample size, because cases in the region decreased considerably and steady enrollment of study subjects became virtually impossible. Results. 160 patients underwent randomization. In the intention-to-treat analysis (ITT), 13/80(16.2%) patients receiving plasma vs. 25/80(31.2%) receiving placebo experienced severe respiratory disease [RR(95%CI)= 0.52(0.29,0.94); p=0.026)] with an RRR=48%. A modified ITT analysis, excluding six subjects who experienced the primary endpoint before infusion, showed a larger effect size [RR(95%CI) = 0.40(0.20, 0.81), p=0.007]. High- and low-titer donor analyses, based on a median IgG titer=1:3,200, evidenced a dose-dependent response with an RRR=73.3% for recipients of high-titer plasma (p=0.016) and a number needed to treat (NNT)=4.4. All secondary endpoints exhibited trends towards protection. No solicited adverse events were observed. Conclusions. Early administration of high-titer convalescent plasma against SARS-CoV2 to mildly ill infected seniors reduced COVID-19 progression. This safe, inexpensive, outpatient intervention facilitates access to treatment from industrialized to LMIC, can decompress demands on hospitals, and may contribute to save lives.

SARS-CoV2, the etiologic agent of coronavirus disease 2019 , elicits a particularly severe illness for the elderly, who experience high hospitalization rates and account for most fatalities worldwide [1, 2] . Various comorbidities aggravate the prognosis of COVID-19 regardless of age, including hypertension, diabetes, cardiovascular disease, obesity, chronic renal failure, and chronic obstructive pulmonary disease (COPD) [1, 2] .

Treating COVID-19 early remains elusive. Few strategies provide some benefit, several failed, and others are under evaluation [3] [4] [5] [6] . One of the latter strategies relies on infusion of specific antibodies present in plasma of convalescent subjects [7] [8] [9] .

Plasma infusions are safe [10] and modulated other diseases in the past [11] [12] [13] . But antibodies in plasma must be administered early after infection to be effective [11] [12] [13] .

To date, late infusion of convalescent plasma against SARS-CoV2 to hospitalized patients with COVID-19 has not shown clear benefits [7] [8] [9] . Therefore, we articulated a team of >250 health professionals and volunteers to address a critical question around timing of plasma as a therapeutic. The main objective of our study was to advance administration of high-titer plasma to the first days after initiation of symptoms. Here, we evaluated the efficacy of convalescent plasma with high titers of SARS-CoV2 antibody administered within 72 hours of mild symptoms to elderly All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101 https://doi.org/10. /2020 subjects with COVID-19. The aim of our study was to prevent progression to severe disease.

Trial design and oversight. We conducted a randomized, double-blind, placebo- 23 2020). An independent Data and Safety Monitoring Board (DSMB) supervised the study. The authors designed the trial, compiled and analyzed the data, and vouch for their completeness and accuracy and for the adherence of the trial to the protocol. All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101 https://doi.org/10. /2020 Trial patients. Subjects ≥75 years of age irrespective of presenting comorbidities or between 65-74 years of age with at least one comorbidity (hypertension or diabetes under pharmacologic treatment, obesity, chronic renal failure, cardiovascular disease, and COPD defined in Suppl. Material) were identified and assessed for eligibility.

Eligible subjects had experienced at least one of each in the following two categories of signs and symptoms for <48 hours at the time of screening for SARS-CoV2 by reverse-transcriptase-polymerase-chain-reaction (RT-PCR): (1) a temperature ≥37.5°C and/or unexplained sweating and/or chills, and (2) dry cough, dyspnea, fatigue, myalgia, anorexia, sore throat, dysgeusia, anosmia, and/or rhinorrhea.

Exclusion criteria included already presenting severe respiratory disease (our primary endpoint, see below) and/or any disease outlined in Suppl. Material.

Subjects consenting to screening were visited at home and tested in nasopharyngeal and oropharyngeal secretions by RT-PCR for SARS-CoV2 (Atila iAMPâ COVID-19, Atila BioSystems). Those with detectable SARS-CoV2 RNA were transported to study hospitals and invited to sign the informed consent. Legal guardians signed consent for participating patients with cognitive impairment since July 22 2020. Starting July 27 2020, as several geriatric institutions facing SARS-CoV2 outbreaks were transformed into low-complexity units for mildly symptomatic All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 residents, we also screened and invited residents meeting study criteria to participate in the study on site.

Randomization and treatment regimens. After obtaining written informed consent, eligible patients were randomized to receive 250 ml of convalescent plasma with an IgG titer against SARS-CoV2 spike (S) protein >1:1,000 (COVIDAR IgG, Instituto Leloir, Argentina) or placebo (normal saline 0.9%) administered over 1.5 to 2 hours in a 1:1 ratio. Patients were randomly assigned using a computer-generated randomization sequence, with balanced permuted blocks of two. The sequence was prepared centrally at the Data Center. Both treatment and placebo were concealed using opaque bags and tape to cover the infusion line. Treatment was administered <72 hours from initiation of symptoms. Subjects were monitored for 12 hours after treatment for adverse events. 479 volunteers infected with SARS-CoV2 for a minimum of 10 days, asymptomatic for ≥3 days, and with two negative RT-PCR tests [14] were identified through hospital lists and an online campaign. Candidates were visited at home and screened for SARS-CoV2 S IgG titers >1:1,000 in serum. 135 (28.2%) were invited to donate 750 ml of plasma at four hemotherapy centers in Buenos Aires (Suppl.

All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 Clinical and laboratory monitoring. Twenty-four hours after completing the infusion, a sample of venous blood (5 ml) was obtained from participants and preserved at -20°C until completion of the study to measure anti-S IgG SARS-CoV2 using COVIDAR IgG validated with SARS-CoV-2 Spike S1-RBD IgG ELISA Detection Kit (GenScript). In addition, we assayed samples with the SARS-CoV-2 Surrogate Virus Neutralization Test Kit (GenScript). Patient evolution was monitored daily by study physicians until day 15 to assess the primary endpoint at the hospitals, in participating geriatric institutions, or at home if patients were discharged. Those persistently symptomatic were followed until resolution of symptoms or a maximum of 25 days for secondary endpoints. Study physicians used predesigned questionnaires to collect clinical information. Data were recorded using paper forms scanned digitally due to a COVID-19 mitigation protocol and subsequently double-entered in an electronic database.

Trial endpoints. The primary endpoint of the trial was development of severe respiratory disease defined as a respiratory rate (RR)≥30 and/or an O 2 sat<93% when breathing room air determined between 12 hours after infusion of the investigational product (IP) and day 15 of study participation. Pre-specified secondary clinical endpoints assessed, as needed, until day 25 of study participation included (a) life threatening respiratory disease, defined as need for 100% oxygen supplementation All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 and/or non-invasive or invasive ventilation and/or admission to intensive care; (b) critical systemic illness, defined as respiratory failure (PaO2/FiO2 ≤ 200 mm Hg) and/or shock and/or multiorganic distress syndrome (defined in Suppl. Material) and (c) death associated with COVID-19. On July 22 2020, we amended the protocol to include a fourth secondary endpoint including any of the three endpoints described above, alone or in combination.

Early study termination. In late July, our region faced a dramatic increase in SARS-CoV2 infections and participating hospitals were overburdened with severe cases.

This was compounded by an extraordinary demand for COVID-19 ambulances, which served as means of transportation for mildly symptomatic candidates from their homes to our hospitals. In addition, large groups of physician investigators infected or exposed to SARS-CoV2 were quarantined for weeks, compromising manpower. Hence, overburdening hospitals with elders presenting mild symptoms became challenging.

As several geriatric institutions facing outbreaks were transformed into lowcomplexity units for mildly symptomatic residents, we seek study candidates in them since July 27 2020. Eventually, COVID-19 cases in the region decreased considerably and, by late September and early October, we were enrolling one subject per week.

This enrollment pace projected at best ~5 months to complete the study in the All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 unlikely case of the virus remaining in circulation at a constant rate. Consequently, after discussions with the DSMB and having enrolled 76% of the target population, the principal investigator (FPP) and lead sub-investigators (RL, GPM and DW) decided it would be logistically impossible, and ethically questionable given the urgency of the problem, to continue the study, and stopped to examine the results.

Given the relative complexity of implementing this intervention, the minimally clinically important difference was set at a 40% relative reduction for an expected outcome rate of 50% in the control group reduced to 30% in the intervention group. A total sample size of 210 subjects (105 per trial arm) was estimated to have 80% power at a significance level (alpha) of 0.05 using a two-sided z-test with continuity correction.

We performed an intention-to-treat (ITT) analysis. A descriptive analysis was implemented by presentation of continuous variables as means and standard deviations or medians and interquartile ranges, as appropriate, and categorical variables as proportions.

Under the primary analysis strategy, we used the Kaplan-Meier product limit distribution to compare treatment groups for the time taken to reach the primary endpoint. An estimate of the relative risk and 95% confidence interval was also reported. A modified ITT (mITT) analysis was defined by the post-randomization All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 exclusion of subjects that lost eligibility between randomization and treatment administration. The protocol pre-specified a subgroup analysis by two populations defined in the inclusion criteria, subjects ≥75 years of age irrespective of presenting comorbidities or between 65-74 years of age with at least one comorbidity and a second analysis of protection correlates in donors' plasma and recipients' sera.

Study Population. 165 subjects presented symptoms that met eligibility criteria and tested positive for SARS-CoV2 RNA (Suppl. Material). Four of them lost eligibility before enrollment and one did not consent to participate in the trial. Therefore, 160 subjects with SARS-CoV2 infection underwent randomization; 80 were assigned to receive convalescent plasma and 80 to receive placebo. Five (3.1%) patients (3 allocated to plasma and 2 to placebo) received IP after meeting the study's primary endpoint. One additional patient randomized to receive plasma developed hypoxemia before the infusion and was not treated with IP.

160 subjects were included in the ITT analysis (Suppl. Material). One subject voluntarily abandoned the trial on day 11 of follow up. 38/160 (23.7%) subjects continued to experience COVID-19 symptoms and were followed until recovery or death for 16 to 25 days.

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The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 The mean age of subjects was 77.1 (S.D. 8.6) years; 100 (62.5%) were females (Table 1) . 72 (45%) were 65-74 years and 88 (55%) ≥75 years old. Baseline characteristics showed no clinically significant imbalances between treatment and control groups. Most patients had prespecified comorbidities at enrollment ( Table 1) Table 2 ). In the time to event analysis, patients in the plasma group also had a longer time to development of severe COVID-19 than patients receiving placebo [median (Q1-Q3) = 15 vs. 15 (8.8-15) ; p=0.028 in Four subjects in the intervention group and two subjects in the control group lost eligibility by reaching the primary endpoint before receiving the IP. A clinically relevant, modified ITT population was defined by excluding these subjects. The results of the mITT analysis showed a larger intervention effect size, with 9/76 All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101/2020.11.20.20234013 doi: medRxiv preprint (11.8%) patients receiving plasma vs. 23/78 (29.5%) receiving placebo experiencing severe respiratory disease [RR (95%CI) = 0.40 (0.20, 0.81), p=0.007]. In the mITT population, patients in the plasma group also had a longer time to development of severe COVID-19 than patients receiving placebo (p=0.006; Suppl. Material).

Results for secondary endpoints are presented in Table 2 .

Four (5%) plasma recipients and 10 (12.5%) placebo recipients experienced lifethreatening respiratory disease, while 5 (6.2%) and 6 (7.5%) had critical systemic illness, respectively. 2 plasma vs. 4 placebo recipients died. The combined secondary endpoint (life-threatening respiratory and/or critical systemic disease and/or death) was met by 7 (8.8%) subjects treated with plasma vs. 12 (15%) treated with placebo.

Secondary endpoint results for the mITT analysis are presented in Suppl. Material.

placebo recipients differed significantly, with higher concentrations in subjects allocated to the intervention (log median (Q1-Q3) = 5.7 (4.9-6.3) vs. 3.9 (3.9-4.7); p<0.0001; Fig.2 ). Comparison between severe and mild cases detected no correlate of protection in plasma recipients' sera 24 hours after infusion (Suppl. Material).

Conversely, a dose-dependent effect was evident for SARS-CoV2 S IgG titers in plasma bags ( Table 3) . Analysis of high and low titer donors classified based on a All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101 https://doi.org/10. /2020 median titer of 1:3,200, showed a RRR=73.3% with a NNT=4.4 for recipients of high titer products (p=0.016; Table 3 ). SARS-CoV2 S IgG results were replicated using a different SARS-CoV-2 Spike S1-RBD IgG commercial assay providing a potential alternative tool for donor selection (r=0.7; p<0.001 in Suppl. Material).

In this manuscript, we report the first specific treatment against early COVID-19 of potential global access and low relative cost. Early administration of plasma with high titers of antibody against SARS-CoV2 to infected seniors reduced progression to severe COVID-19 by 48%. Selecting plasma with IgG titers above a median of 1:3,200 increased protection to 73%. Our findings stress the need to return to the old paradigm of treating acute viral infections early and define IgG targets that facilitate donor selection.

In our study, timing and disease severity at the time of treatment differed from previous reports. A randomized trial of convalescent plasma vs. standard treatment in hospitalized patients with severe or life-threatening COVID-19 in China did not impact clinical improvement [7] ; a second randomized trial in Indian hospitals found no differences in progression to life-threatening disease and mortality between plasma and placebo recipients [9] ; and a third study did not show significant differences in All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 probability of death for severely ill patients using propensity score-matched controls [8] .

Early high-titer plasma administration is a therapeutic breakthrough, as alternatives against COVID-19 are scarce. The monoclonal antibody LY-CoV555 has been shown to decrease viral load in recently diagnosed mild or moderately-ill outpatients [15] , remdesivir shortens the time to recovery in hospitalized patients with COVID-19 [16] , and dexamethasone decreases mortality in severely ill subjects [17] .

Convalescent plasma can bridge the transition to accessible vaccines as a potentially inexpensive (per patient cost in our region=u$186.25; Suppl. Material), outpatient, effective intervention with reasonably rapid access for many LMIC.

Six subjects reached the primary endpoint before receiving IP. To provide plasma of all blood types to 15 institutions, we instrumented two unblinded infusion teams that drove from a central hemotherapy station harboring the product to all hospitals after randomization. The study region was >100 miles 2 and security challenges precluded access to several hospitals after 8 pm. Exclusion of these patients in the mITT analysis increased the efficacy of the intervention to 60%. Since plasma infusions are safe and relatively easy to administer, and acknowledging that situations may vary in different regions, implementation may work best at designated outpatient clinics with donations fostered at a local level reinforcing a sense of community kinship. Therefore, we speculate that efficacy may approach the mITT in All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101 https://doi.org/10. /2020 many settings or ascend >70% if donors with IgG titers ≥1:3,200 are selected. The strong correlation between assays suggests that other S/RBD IgG immunoassays may provide reasonable alternative screening tools for implementation (Suppl. Material).

Enhancing early symptom awareness in seniors will be vital, now that there is a timelimited effective intervention available. Plasma against COVID-19 is conceptually like health insurance. It should be in-hand when it intuitively seems unnecessary.

Our study demonstrates a dose-dependent IgG effect in plasma infusions.

Plasma with IgG titers ≥1:3,200 reduced the risk of severe COVID-19 by 73%. Since IgG concentrations, and not volume, are critical for therapeutic success, numerous "super-donors" with IgG titers ≥1:12,800 could help >20 subjects by providing 750 ml. Maintenance of high IgG titers for months stress the value of fostering their loyalty for recurrent donations [18] . Interestingly, 71% of donors ≥1:3,200 among our volunteers had been hospitalized (not shown).

In summary, administration of select convalescent plasma against SARS-CoV2 to infected seniors within 72 hours of mild symptoms reduced COVID-19 progression to severe illness in a RCT. This simple, safe and inexpensive intervention can be rapidly available, decompress demands on the health care system, and may save lives. Early plasma infusions can provide a bridge to the future, until vaccines become widely available.

All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101 https://doi.org/10. /2020 preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101/2020.11.20.20234013 doi: medRxiv preprint

All rights reserved. No reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10.1101/2020.11.20.20234013 doi: medRxiv preprint

Log SARS-CoV2 IgG titer in recipients' sera placebo plasma p<0.001

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The copyright holder for this this version posted November 21, 2020. ; https://doi.org/10. 1101 

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