Antibody titer 6 months after the third dose of COVID-19 mRNA vaccination

Introduction

As of the middle of July 2022, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) had infected over 558 million individuals worldwide and caused more than 6.36 million deaths. A third dose of SARS-CoV-2 vaccine was administered from December, 2021, in Japan. A number of reports have been published regarding the humoral responses induced by and the effectiveness of the third dose of Pfizer BioNTech (BNT162b2) vaccine [1], [2], [3], [4], [5], [6], [7], [8]. However, the long-term persistence of the immune response after the third dose of vaccine has not been reported, especially in healthy volunteers. Our group recently reported the antibody titers and side effects observed after two doses of BNT162b2 mRNA vaccine [9], the decline in antibodies 6 months after the first vaccination [10] and the potent incremental increase in the IgG antibody titer after the third vaccination [11]. Furthermore, we demonstrated that the production of IgG after the third dose of BNT162b2 vaccine decreased age-dependently, and the number of side effects generally decreased with age [12]. In the current study, we examined the levels of SARS-CoV-2 antibodies among healthy volunteers at Tohoku Medical and Pharmaceutical University Hospital, 6 months after the third vaccination.

Materials and methods

Results

In total, 40 out of the 41 participants completed 14 months of follow-up since the first dose of BNT162b2 (Table 1). Serum samples were obtained at an average of 445 days (SD 4.8 days) after the first dose of BNT162b2 (Table 1, Figure 1A), which is 180 days after the third vaccination. At this time point, the mean anti-RBD IgG level was 159.4 AU/mL (SD 100.1 AU/mL), compared with the mean anti-RBD IgM baseline level (0.4 C.O.I.) (Table 1, Figure 1B). The mean anti-RBD IgG level was significantly higher than 144 days after the second vaccination.

Figure 1:

(A) Schematic presentation of the sampling procedure. (B) Box plot of the anti-SARS-CoV-2 IgG antibody levels following vaccination with the BNT162b2 vaccine. Current data at day 445 are shown in black, and data at baseline, and on days 14, 35, 180, 264 and 279 are shown in gray and are presented for comparison. Data are shown as boxplots representing the 25th percentile, 75th percentile, median, and 95% confidence interval using Prism software (version 9.0; GraphPad software Inc., La Jolla, CA, USA). Statistical significance of the data at each time point compared with that at day 445, calculated by Tukey’s multiple comparison test, is indicated by asterisks (*p<0.05, **p<0.01, ***p<0.001, ****p<0.0001).

The decline in the IgG level, 180 days after the third vaccination, was 74.1% (SD 16.1%), which was significantly lower than the decline (88.6% [SD 4.4%) even 144 days after the second vaccination (Figure 2). Interestingly, before (R²=0.3512, Spearman’s r=−0.6429, p<0.00001) and 14 days after (R²=0.1481, Spearman’s r=−0.4563, p=0.0027) the third dose of COVID-19 mRNA vaccination, we observed a clear decrease in IgG level with age (12). By contrast, in the current study, at 180 days after the third vaccination, this difference disappeared and no relationship between IgG levels and age was detected (R²=0.000579, Spearman’s r=−0.0572, p=0.8828) (Figure 3A). We further revealed that, the reduction in IgG from 14 to 180 days after the third vaccination significantly inversely correlated with age (R²=0.1514, Spearman’s r=−0.4693, p=0.0131) (Figure 3B). We further calculated percental loss of IgG and found that this reduction was also tended to be decreased with age (R²=0.07426, Spearman’s r=−0.3,056,837, p=0.0889).

Figure 2:

Comparison of the humoral responses and the duration of the IgG titer after the first and second (red) and third (black) vaccinations. Circles and squares represent the mean value for each time point and the standard deviations are also shown. p-values were calculated from the end point decline using the Mann–Whitney test and Prism software (version 9.0; GraphPad software Inc., La Jolla, CA, USA). Additionally, the non-liner regression curve, calculated using the least-squares method, and the extra sum of the squares F-test was employed to validate this difference (****p<0.0001).

Figure 3:

Correlation between age at day 445 post-vaccination and (A) IgG levels, and (B) reduction in IgG levels (C) percental loss of IgG levels (i.e., the decline in IgG from 14 days after the third vaccination), with the superimposed linear regression lines (with 95% CI) shown. Black circles represent males and gray circles represent females. Statistical methods were based on Spearman’s rank correlation coefficient and p-values are indicated. Linear regression lines, blue.

Discussion

Recently, Padoan et al. [13] reported the SARS-CoV-2 anti-spike protein RBD IgG levels and the neutralizing antibody levels 3–4 months after the third dose of vaccination in healthcare workers. Their study lacked a time point before the third dose vaccination, which meant that it was difficult to interpret the decrease in titer during this 3 to 4-month period. However, in their study, after the second dose vaccination, IgG levels were 2,719.0 (723–4,154.0) kBAU/L in COVID (˗) subjects, compared with 1,076.0 (529.3–3,409.0) kBAU/L 3–4 months after the third dose, which was approximately 40% of the level detected after the second dose. In our current study, the decline in IgG level was 88.6% after the second dose and 74.1%, a significantly lower decrease, after the third dose, which is relatively consistent with Padoan et al. findings that the decline is not so profound after the third dose (13) compared with after the second dose [10]. It was recently published that 3 months after the third vaccination with BNT162b2, the higher levels of neutralization of Delta variants are maintained [14], further confirming the durability of the third vaccination. A similar report was published by Herzberg et al. [15], that antibody levels dropped significantly within a short period of 11 weeks after the third dose, but even after this decline, levels still remained higher than after the second dose.

To date, no report has been published regarding antibody levels 6 months after the third vaccination. Recently, Xin et al. reported the neutralization titers and safety of four regimens involving 6-month follow-up after a booster dose of vaccination in two single-center phase 2 clinical trials [16]. Although they did not assess IgG antibody titers, they reported that the neutralization titers declined approximately 4-fold and 2.5-fold from days 28 to 180 after the third dose in adults aged 18–59 years and those aged 60 years and older, respectively [16]. Our results are consistent with this study, confirming that the older population maintains a similar level of IgG titer compared with the younger population. This finding may indicate the durability of the long-term humoral response in the older population.

The long-term efficacy of BNT162b2 vaccination remains to be determined. However, we found that 6 months after the third vaccination, the mean anti-RBD IgG level was significantly higher than 144 days after the second vaccination, and also we revealed that the waning of the IgG level 6 months after the third vaccination, is significantly less compared with the decline following the second vaccination. The higher rise of IgG antibody titer after the third compared to the second vaccination and the lower decline both result in a higher mean antibody titer 6 months after the third vaccination. Collectively, our results support the utility of the third booster dose in reinforcing the duration of humoral immunity.