Limited window for donation of convalescent plasma with high live-virus neutralizing antibodies for COVID-19 immunotherapy

The optimal timeframe for donating convalescent plasma to be used for COVID-19 immunotherapy is unknown. To address this important knowledge deficit, we determined in vitro live-virus neutralizing capacity and persistence of IgM and IgG antibody responses against the receptor-binding domain and S1 ectodomain of the SARS-CoV-2 spike glycoprotein in 540 convalescent plasma samples obtained from 175 COVID-19 plasma donors for up to 142 days post-symptom onset. Robust IgM, IgG, and viral neutralization responses to SARS-CoV-2 persist, in the aggregate, for at least 100 days post-symptom onset. However, a notable acceleration in decline in virus neutralization titers ≥160, a value suitable for convalescent plasma therapy, was observed starting 60 days after first symptom onset. Together, these findings better define the optimal window for donating convalescent plasma useful for immunotherapy of COVID-19 patients and reveal important predictors of an ideal plasma donor, including age and COVID-19 disease severity score. One Sentence Summary Evaluation of SARS-CoV-2 anti-spike protein IgM, IgG, and live-virus neutralizing titer profiles reveals that the optimal window for donating convalescent plasma for use in immunotherapy is within the first 60 days of symptom onset.

We discovered robust IgM, IgG, and VN responses in the majority of individuals, with 80 moderate to strong correlation regardless of assay type (Figure 1a  reported that anti-S/RBD and S/ECD IgG titers are excellent surrogates for in vitro VN and help 106 identify plasma donors for therapeutic uses 1,14 . Specifically, we have shown that anti-S/RBD or 107 anti-S/ECD antibody titers of ≥1350 are strong proxies for a VN titer ≥160, the FDA-108 recommended value for use in COVID-19 convalescent plasma therapy 1 , and transfusion of 109 anti-S/RBD IgG ≥1350 titer plasma within 72 hours (h) of hospitalization significantly improves 110 survival and health outcomes 15,16 .

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Our large and well-characterized convalescent plasma library with longitudinally donated 112 samples enabled detailed assessment of VN response persistence. We found that the 113 proportion of individuals with a VN titer ≥160 remained above 80% through the first 60 DPO but 114 declined sharply to less than 20% between DPO 61 and 120 ( Figure 2c). These results suggest 115 that the time period in which donated convalescent plasma is likely to have a high VN titer and 116 optimal therapeutic potential is within the first 60 DPO. This has important implications for 117 convalescent plasma donation and passive immunotherapy programs, some of which have 118 already transfused more than 60,000 individuals in the United States as of August 13, 2020 119 (https://www.uscovidplasma.org).

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Facile methods to identify suitable convalescent plasma donors are needed as the gold 121 standard live-virus VN assays used herein are labor intensive, cumbersome, take several days 122 to perform, and require specialized expertise and access to a high containment (Biosafety Level 123 3) laboratory and regulatory clearances. ELISAs are easier to implement than VN assays, 124 especially in resource-limited countries and environments. We previously reported that an 125 S/RBD ≥1350 titer may serve as a good marker for identifying plasma donors with VN ≥160 1 126 (Supplementary Table S2 Table S2). Thus, an S/RBD ≥1350 titer is a promising marker for 132 identifying suitable plasma donors early, but not late, after first symptom onset. In contrast, 133 S/RBD IgG ≥1350 appears to be a reliable predictor of VN ≥160, and S/RBD IgG ≥1350 survival 134 is statistically indistinguishable from that of VN ≥160 (Figure 2e), with an overall LR+ of 3.18 and 135 a negative likelihood ratio (LR-) of 0.26 (Supplementary Table S2).

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We next investigated the survival and predictive values of S/RBD IgM ≥450 as compared  Table S2). An S/RBD IgM titer ≥450 was selected 138 because the magnitude of IgM response was approximately three-fold lower than that of IgG 139 ( Figure 1f). The results showed that S/RBD IgM ≥450 had a similar survival profile to VN ≥160 140 but waned significantly faster (P <0.01; Figure 2f). While S/RBD IgM ≥450 had an overall LR+ of 141 3.72, it also had a LR-of 0.69, which would likely result in an unacceptable number of suitable 142 donors with VN ≥160 being excluded. Together, these results indicate that S/RBD IgG ≥1350, 143 but not IgM ≥450 or S/RBD or S/ECD total antibody ≥1350, serves as a good marker to identify 144 suitable plasma donors for COVID-19 immunotherapy.

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To determine the kinetics and persistence of IgM, IgG, and VN responses, we next  period. All individuals with a detectable starting titer remained, on average, between one or two 151 dilutions above or below the initial titer (Extended Data Figure 1). Of particular note, only 5 of 60 152 individuals (8.3%, 95% CI: 2.8-18.4%) with an initial VN titer of ≥ 5.3 (1:40) showed a 153 subsequent increase in titer, emphasizing the importance of recruiting and screening 154 convalescent plasma donors quickly, as VN titers are unlikely to rise from initial levels.

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We next assessed whether particular donor characteristics predicted a more robust 156 serological and neutralization response. The results show that individuals 30 years of age or younger had significantly lower VN, IgG and IgM antibody titers than those in the older age 158 groups ( Figure 3a). Individuals between 20-30 years of age also had significantly faster decline 159 in IgG (P <0.05) and IgM (P <0.05) than did those > 60 years of age (

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In conclusion, these data refine our understanding of the kinetics, magnitude, and 184 durability of human serologic responses to SARS-CoV-2 spike protein, the primary vaccine 185 candidate being studied worldwide. This integrative analysis of serological and VN profiles 186 identifies an optimal donation window of up to 60 DPO for high-titer anti-spike protein 187 convalescent plasma as immunotherapy for COVID-19 patients. Our analysis found that 188 additional characteristics of an ideal potential donor include a recovered patient >30 years old 189 with a high COVID-19 disease severity score. In the aggregate, these data permit a more  followed. An aliquot of collected plasma was tested for antibodies by ELISA and/or VN assays.

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Cohort characteristics are described in Table 1