Restoration of Neutralization Activity Against Omicron BA.2 and BA.5 in Older Adults and Individuals With Risk Factors Following the Fourth Dose of Severe Acute Respiratory Syndrome Coronavirus 2 BNT162b2 Vaccine

To the Editor—We read with interest the article by Mwimanzi et al about humoral responses after second and third doses of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) messenger RNA vaccination in older adults [1]. Here, we determined the effects of consecutive 4 BNT162b2 doses on neutralizing activity in sera from participants aged ≥60 years and those with risk factors against SARS-CoV-2 Wuhan and 2 Omicron sublineages, and analyzed longitudinal changes of neutralization activity pre– and post–second through fourth BNT162b2 doses in a prospective study over 490 days.

In the present prospective clinical study, 32 of 225 healthcare workers in Kumamoto General Hospital, Japan (225 were initially recruited in the primary clinical study [2]), who were either ≥60 years of age and/or had preexisting diseases/risk factors (see demographic characteristics in Supplementary Table 1A) were enrolled. These 32 participants received a fourth BNT162b2 dose as they had risk of developing severe coronavirus disease 2019. The SARS-CoV-2 neutralizing activity (50% neutralization titer [NT₅₀]) of their sera against the Wuhan strain of SARS-CoV-2 was determined over 490 days using sera collected consecutively on (i) 1 week post–second dose, (ii) 2 weeks post–third dose, (iii) 1 week pre–fourth dose, and (iv) 2 weeks post–fourth dose (Supplementary Methods), representing a continuation of our previous studies [2–4]. We also evaluated the profile of S1-binding immunoglobulin G (IgG) levels following pre–/post–fourth dose (on days 300/490). In addition, we determined NT₅₀ of the same sera using VeroE6^TMPRSS cells against infectious Omicron BA.2 and BA.5 variant sublineages, whose emergence has been associated with the present explosive increases globally [5].

Against SARS-CoV-2 Wuhan, moderate neutralizing activity was seen on day 28, 1 week after second-dose samples (Figure 1A, mean NT₅₀ = 307), whereas there was a remarkable rise in neutralizing activity of the same participants’ sera of 2 weeks after third-dose administration (day 300), achieving a mean NT₅₀ of 2238 (Figure 1A). On day 470 (1 week pre–fourth dose), the mean NT₅₀ decreased to 541, 24% of the peak value on day 300 (2 weeks post–third dose) (Figure 1A). However, by day 490, 2 weeks post–fourth dose, neutralization activity was restored to 2096 (Figure 1A). The mean S1-binding IgG levels in pre–/post–fourth dose sera also showed an increase, from 247 on day 470 (1 week pre–fourth dose) to 1152 on day 490 (2 weeks post–fourth dose) (Supplementary Table 1B).

Figure 1.

Neutralizing activity of post–second, third, and fourth dose sera against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Wuhan strain, Omicron BA.2, and Omicron BA.5. Temporal changes of neutralizing activity of participants’ sera over 490 days post–first dose are shown. BNT162b2 doses were administered on days 0, 21, 287, and 477. A, The 50% neutralization titers (NT₅₀) of participants’ sera against infection by SARS-CoV-2^05–2N strain were determined on days 7, 28, 60, 90, 150, 280, 300, 360, 470, and 490 post–first dose using VeroE6^TMPRSS2 cell-based neutralization assay. Solid circles denote NT₅₀ titers of each participant's serum and filled bars denote average NT₅₀ titers of 32 participants’ sera at each time point. Geometric mean NT₅₀ titers and ranges of NT₅₀ at each time point are shown at the bottom. B, Temporal changes of neutralizing activity of participants’ sera at day 28, 300, 470, and 490 post–first dose against Omicron BA.2 and BA.5 variants are shown. Solid circles denote NT₅₀ titers of each participant's serum and filled bars denote average NT₅₀ titers of 32 participants’ sera at each time point. The NT₅₀ values were found to be normally distributed following conversion to the log₁₀ values and the differences between day 300 and day 490 values were calculated using paired t test.

On the other hand, mean NT₅₀ values against BA.2 and BA.5 on day 28, 1 week post–second dose were only 27 and 24, respectively, close to the cutoff value (<20) (Figure 1B). By contrast, mean NT₅₀ of sera on day 300 (2 weeks post–third dose) against BA.2 and BA.5 increased to 223 and 191, respectively (Figure 1B). By day 470, 1 week pre–fourth dose, the mean NT₅₀ had decreased to 28% and 25% of those observed on day 300 sera against BA.2 and BA.5, respectively (Figure 1B, mean NT₅₀ = 62 and 47, respectively); however, the mean NT₅₀ against BA.2 and BA.5 was restored to 292 and 205, respectively, on day 490, to the extent seen after the third dose (Figure 1B).

Compared to the significantly boosted response elicited by the third dose, the magnitudes of neutralizing activity against SARS-CoV-2 Wuhan following the fourth dose were only comparable to and not greater than those following the third dose (Figure 1). The magnitudes of neutralizing activity against 2 Omicron variants following the fourth dose were also comparable to and not greater than those following the third dose. The present data, that a fourth vaccine dose restores protection but does not further enhance the humoral response, may be related to “original antigenic sin” [6], wherein high-affinity memory B cells inhibit the recruitment of naive B cells against subsequent antigenic stimuli, in particular, against new stimuli. Thus, it is likely that despite the fourth dose, breakthrough infections continue to occur. It remains to be determined whether the upcoming Omicron-specific booster vaccines strengthen or attenuate the Omicron-specific protection already acquired through 4 regular doses in older individuals and those with other risks.

Supplementary Data

Supplementary materials are available at The Journal of Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.

Notes

Author contributions. M. A. and H. M. had access to all data in this study and take responsibility for the integrity of the data and the accuracy of the data analysis. Concept and design: M. A. and H. M. Acquisition, analysis, or interpretation of data: M. A. and S. O. Obtained funding: H. M. Administrative and material support: Y. I., N. H.-K., and S. S. Supervision: S. M. and H. M. Original draft writing: M. A. and H. M. Approved the final version of the manuscript: All authors.

Acknowledgments. The authors greatly appreciate Yukari Uemura, PhD (Department of Data Sciences, Center for Clinical Sciences, National Center for Global Health and Medicine [NCGM]) for the statistical analysis, and Kenji Maeda, MD, PhD, Yuki Takamatsu, MD, Shinichiro Hattori, PhD, Mariko Kato, and Asuka Fujiwara (all from NCGM Research Institute), and Kiyoto Tsuchiya, PhD (NCGM hospital), for their dedicated experimental support. The authors also thank Mari Kinoshita (Japan Community Health Care Organization Kumamoto General Hospital) for handling blood samples, and Kenji Sadamasu, DVM (Tokyo Metropolitan Institute of Public Health) and Yukihito Ishizaka, MD, DMS (NCGM Research Institute) for providing SARS-CoV-2²⁰³⁷ (Omicron/BA.2) and SARS-CoV-2^TKYS14631 (Omicron/BA.5), respectively.

Financial support. This research was supported by the Japan Agency for Medical Research and Development (grant number 20fk0108502 to H. M.); the Intramural Research Program of the National Center for Global Health and Medicine (grant number 19A3001 to H. M.); and the Intramural Research Program of the Center for Cancer Research, National Cancer Institute, National Institutes of Health (to H. M.).

Data availability

The data sets generated during this study are available from the corresponding author upon request.