Immune response in the participants on day-60 post 1st dose.
We obtained blood samples for neutralizing activity evaluation from a total of 225, 220, 211, 210, and 208 vaccine recipients on days 7, 28, 60, 90, and 150 post 1st doses, respectively28. Demographic characteristics of the participants are shown in Table 1. As of the time of enrollment, the average age of the participants was 41.8 years (range: 21 to 72 years), and 69.8% of the participants were female serving as a physician, nurse, paramedical staff, or administrative staff. None of the participants was in the immunodeficient state or was receiving immunosuppressants or steroids.
In the present study, we focused upon the blood samples taken on day-60 post 1st dose from 211 vaccinated participants. First, we determined the neutralizing activity of each serum against cytopathic effect induced by wild-type SARS-CoV-205−2N infection and replication in the VeroE6 cell-based assay. As shown in Fig. 1, 50% neutralization titer (NT50) levels were substantially diverse among the participants: the average NT50 values was 284 (range 30 − 1,290), and five participant’s sera showed NT50 values less than cut-off level (< 20).
High and moderate responder’s sera show reduced anti-SARS-CoV-2 activity against various VOCs and VUM.
Next, we selected and designated day-60 participants, whose sera had NT50 values against SARS-CoV-205−2N greater than 500 as high responders (n = 15; shown in red), and participants, whose sera had NT50 values around average of all participants (range 220 to 380) as moderate responders (n = 30; shown in blue) (Fig. 1), and evaluated neutralizing activity of each serum belonging above mentioned two groups against nine infectious SARS-CoV-2 variants; variants of concerns (VOCs) and variants under monitoring (VUM), including Alpha, Beta, Gamma, Delta, Kappa, and E484K/D614G- amino acid subs carrying variants (E484K variants).
Average NT50 values of high responder’s sera against three Alpha variants, SARS-CoV-2QHN001, SARS-CoV-2QK002, and SARS-CoV-2QHN002 were 351, 348, and 287, respectively, and %reductions from the average NT50 values against SARS-CoV-205−2N (811) were by 56.7, 57.1, and 64.6%, respectively (Fig. 2 and Table 2). High responder’s average NT50 value against the Beta variant, SARS-CoV-2TY8–612, decreased remarkably to 92 (%reduction by 88.7%). High responder’s sera showed moderate reduction with the Gamma variant, SARS-CoV-2TY7–501 with their average NT50 value being 283 and %reduction by 65.1% (Fig. 2 and Table 2). Against the Kappa variant, SARS-CoV-2K5356, high responder’s sera showed high reduction level of neutralizing activity, with NT50 value being 147 and %reduction by 81.9%, whereas high responder’s sera showed lesser reduction against the Delta variant, SARS-CoV-2K1734 with NT50 of 297 and %reduction by 63.4% (Fig. 2 and Table 2). We also examined neutralizing activity of sera against two E484K-carrying variants, SARS-CoV-2F − 76107 and SARS-CoV-2F − 76137, which were isolated in Japan before the emergence of Alpha variants. High responder’s sera showed average NT50 value of 307 to 504 against both variants with reduction by 37.9–62.1% (Fig. 2 and Table 2). As for the group of moderate responders, similar trends were observed with the results of high responder’s group, neutralizing activity more significantly reduced against Beta, Kappa, and Gamma variants, with reduction by 83.4, 73.4, and 71.8%, respectively. Against the other strains tested, moderate responder’s sera showed reductions by 57.5 to 67.4% (Fig. 2 and Table 2).
Overall, sera in both groups showed reduced neutralizing activity by greater than 50% against all variants examined in this study compared to those against wild-type SARS-CoV-205−2N, and the greatest reduction of neutralizing activity occurred with the Beta variant.
IgG fractions obtained from high and moderate responders’ sera show different properties regarding anti-SARS-CoV-2 activity against various VOCs and VUM.
In attempt to further quantify anti-SARS-CoV-2 activity induced by BNT162b2, we obtained IgG fractions from each of high and moderate responder’s serum and carried out the VeroE6TMPRSS2 cell-based anti-SARS-CoV-2 assay determining EC50 (50% effective concentration: µg/mL) value of each IgG fraction against wild-type SARS-CoV-205−2N and nine distinct SARS-CoV-2 variants. We believe that such attempt can exclude any effects caused by non-IgG factors existing in sera, enable to evaluate actual neutralizing activity of IgG alone as induced by the BNT162b2 vaccination against SARS-CoV-2.
As shown in Fig. 3 and Table 3, average EC50 values of IgG eluted from high responder’s sera and moderate responder’s sera against SARS-CoV-205−2N were 10.8 and 23.8 µg/ml, respectively. Against three Alpha variants, SARS-CoV-2QHN001, SARS-CoV-2QK002, and SARS-CoV-2QHN002, high responder’s IgG (IgGhighs) showed EC50 values of 25.6, 24.5, and 26.3 µg/ml, respectively, and fold changes compared with EC50 value against SARS-CoV-205−2N were 2.4, 2.3, and 2.4, respectively. On the other hand, moderate responder’s IgG (IgGmoderates) generally showed more than two-times higher EC50 values compared to those of IgGhighs, as 50.1, 64.0, and 61.4 µg/ml, respectively, against the same three Alpha variants, and fold changes were by 2.1, 2.7, and 2.6, respectively.
High responder’s average EC50 value against the Beta variant, SARS-CoV-2TY8–612, was as high as 77.3 µg/ml, the greatest value among variants we tested, and fold change was by 7.2. IgGhighs showed moderate reduction of antiviral activity against Gamma variant, SARS-CoV-2TY7–501, their average EC50 and fold change were 34.2 µg/ml and 3.2-fold, respectively (Fig. 3 and Table 3). Against Delta and Kappa variants, SARS-CoV-2K1734 and SARS-CoV-2K5356, IgGhighs showed same EC50 value, 25.7 µg/ml, and fold changes were similar with those against Alpha variants (Fig. 3 and Table 3). Also, antiviral activity of IgGhighs against two E484K variants, SARS-CoV-2F − 76107 and SARS-CoV-2F − 76137, were similar level with Alpha variants, average EC50 values were 25.3 and 30.1 µg/ml, respectively, and fold changes were 2.3 and 2.8, respectively (Fig. 3 and Table 3).
On the other hand, average EC50 value of IgGmoderates against SARS-CoV-205−2N was 23.8 µg/ml, 2.2-fold higher than that of IgGhighs. Also, average EC50 values of IgGmoderates against various VOCs and VUM we tested were higher than those of IgGhighs with range of 1.9 ~ 3.2-folds (49.0 ~ 144.5 µg/ml; Fig. 3 and Table 3).
Evaluation of anti-SARS-CoV-2 activity of IgG fractions obtained at different time points (day-28, -60, -90, and − 150 sera) reveals relatively long-lasting antiviral activity of IgG from both groups against Delta-variant.
Next, we also eluted IgG fractions from sera of 25 participants at day-28 and day-90 (n = 13 for IgGhighs and n = 12 for IgGmoderates), then determined EC50 values of each IgG fraction against SARS-CoV-205−2N, various VOCs, and VUM.
As shown in Fig. 4 and Table 4, both IgG groups maintained good anti-SARS-CoV-2 activity against SARS-CoV-205−2N, average EC50 values of IgGhighs and IgGmoderates from day-28 sera were 4.3 and 6.9 µg/ml, respectively, and 13.7 and 36.2 µg/ml, respectively, for the IgG eluted from day-90 sera.
Against Alpha variants (QHN001 and QK002), IgG from day-28 sera of both groups relatively maintain their anti-SARS-CoV-2 activity, average EC50 values of IgGhighs and IgGmoderates were lower than 50 µg/ml (21.4 ~ 30.7 µg/ml for QHN001 and 14.4 ~ 26.0 µg/ml for QK002), and EC50 values were gradually increased time-dependently, IgGhighs from day-90 sera showed EC50 values of 44.9 and 27.2 µg/ml against QHN001 and QK002, respectively, and day-90 IgGmoderates showed EC50 values of 104.9 and 104.3 µg/ml, respectively (Fig. 4A and Table 4A).
IgG from day-28 sera of both groups showed reduced anti-SARS-CoV-2 activity against Beta variant, average EC50 values of IgGhighs and IgGmoderates were 60.8 and 90.8 µg/ml, respectively, and values continued to increase up to 92.6 and 189.3 µg/ml, respectively, for IgG from day-90 sera (Fig. 4B and Table 4A).
Against Gamma variant, IgG from day-28 sera of both groups relatively maintained anti-SARS-CoV-2 activity with EC50 values lower than 50 µg/ml (30.3 µg/ml for IgGhighs and 48.2 µg/ml for IgGmoderates), although values elevated up to 57.4 and 124.6 µg/ml for day-90 IgGhighs and IgGmoderates, respectively (Fig. 4B and Table 4A).
As for Delta variant, both IgG groups from day-28 sera showed good anti-SARS-CoV-2 activity, EC50 were 10.3 µg/ml for IgGhighs and 31.8 µg/ml for IgGmoderates, and maintained relatively favorable EC50 values for IgGs from day-90 sera, EC50 were 22.0 µg/ml for IgGhighs and 65.9 µg/ml for IgGmoderates (Fig. 5 and Table 4B). This favorable antiviral activity of IgGhighs against Delta variant was maintained at least 150 days post 1st dose (Fig. 5 and Table 4B).
Against Kappa variant, IgG from day-28 sera relatively maintained anti-SARS-CoV-2 activity with EC50 values of 20.1 µg/ml for IgGhighs and 51.0 µg/ml for IgGmoderates, and EC50 values elevated up to 62.1 and 124.1 µg/ml for day-90 IgGhighs and IgGmoderates, respectively (Fig. 4C and Table 4A). As for E484K variant, similar results were observed with those of Delta variant (Fig. 4C and Table 4A).