key: cord-0692096-72voooxk authors: Wanlapakorn, Nasamon; Suntronwong, Nungruthai; Phowatthanasathian, Harit; Yorsaeng, Ritthideach; Thongmee, Thanunrat; Vichaiwattana, Preeyaporn; Auphimai, Chompoonut; Wongsrisang, Lakkhana; Klinfueng, Sirapa; Sudhinaraset, Natthinee; Poovorawan, Yong title: Immunogenicity of heterologous inactivated and adenoviral-vectored COVID-19 vaccine: real-world data date: 2022-04-18 journal: Vaccine DOI: 10.1016/j.vaccine.2022.04.043 sha: 612106d18e389f6235a4c703e20a564452c3f62a doc_id: 692096 cord_uid: 72voooxk Limited data are available on the responses to heterologous vaccine regimens for SARS-CoV-2, especially among countries using inactivated and adenoviral-vectored vaccines. A total of 77 participants who received heterologous inactivated COVID-19 vaccine (CoronaVac) and adenoviral-vectored vaccine (AZD1222) were enrolled in our study. There were two comparison groups vaccinated with the homologous CoronaVac (N = 79) and AZD1222 (N = 78) regimen. All sera samples were tested for anti-receptor-binding-domain IgG (anti-RBD IgG) using a chemiluminescent microparticle immunoassay (CMIA). The neutralizing activity in a subset of serum samples was tested against the original Wuhan strain and variants of concern, B.1.1.7, B.1.617.2 and B.1.351, using an enzyme-linked immunosorbent assay (ELISA)-based surrogate virus neutralization test (sVNT). The heterologous CoronaVac/AZD1222 vaccine induced higher levels of anti-RBD IgG than that of two-dose homologous CoronaVac or AZD1222 vaccines (p < 0.001). Sera samples of the CoronaVac/AZD1222 vaccine recipients elicited higher neutralizing antibody activity against the original Wuhan and all variants of concern than in the recipients of the two-dose CoronaVac. The heterologous CoronaVac followed by AZD1222 is an alternative regimen to combat with the SARS-CoV-2 variants in case of vaccine shortage with improved immunogenicity compared to the homologous CoronaVac regimen. Limited data are available on the responses to heterologous vaccine regimens for SARS-CoV-2, especially among countries using inactivated and adenoviralvectored vaccines. In the Com-COV trial, heterologous prime-booster combinations of the adenoviral-vectored (AZD1222) and BNT162b mRNA vaccines induced higher serum anti-spike IgG and neutralizing antibody titers than the homologous efficacyproven adenoviral-vectored two-dose regimen [1] . The CombiVacS trial found that the Pfizer's BNT162b2 mRNA vaccine given as a second dose in individuals prime vaccinated with AZD1222 induced a robust immune response, with an acceptable and manageable reactogenicity profile [2] . Nevertheless, the Com-COV trial found an increase in systemic reactogenicity after booster vaccination reported by participants in heterologous AZD1222-BNT162b schedules in comparison to homologous vaccine schedules [3] . Some countries have recommended the heterologous vaccine regimen, offering the mRNA vaccines as a second dose to young people who have already received the AZD1222 vaccine because of concerns about vaccine-induced thrombotic thrombocytopenia (VITT) [4] . A prospective observational cohort study in Germany showed that the heterologous vaccine regimen (AZD1222/BNT162b2) at a [10] [11] [12] week vaccine interval induced similar anti-RBD IgG responses but increased T-cell responses and neutralization capacity against SARS-CoV-2 alpha and beta variant compared with homologous AZD1222 and BNT162b2 vaccination [5] . Further research is required to comprehensively elucidate the immunological implications following different vaccine types and administration schedules. [10] . Although this self-limited adverse event is rare, individuals experiencing this side effect sought another regimen for their second dose, which was AZD1222. It is possible to mix and match vaccines in specific situations such as a vaccine shortage or for adverse reactions following vaccine administration. This study aims to assess the immunogenicity of heterologous inactivated and the adenoviral-vectored COVID-19 vaccine currently available in Thailand to provide preliminary data on their immunogenicity. We performed a cross-sectional study in which leftover sera samples from participants seeking antibody testing following vaccination at the Center of Excellence For the homologous CoronaVac and heterologous CoronaVac/AZD1222 groups, participants tested positive for anti-N IgG were not excluded as it was likely to be from a post vaccination with inactivated vaccine. CoronaVac is an inactivated virus vaccine created from African green monkey kidney cells (Vero cells) that have been inoculated with SARS-CoV-2 (CZ02 strain). The virus was inactivated with β-propiolactone and finally absorbed onto aluminum hydroxide. Each vial contains 0.5 mL with 600 SU (equal to 3 µg) of inactivated SARS-CoV-2 virus as antigen [11] . Chimpanzee adenovirus Oxford 1 (ChAdOx1)vectored vaccine (AZD1222) from Oxford/AstraZeneca is a non-replicating viral vector vaccine that stimulates an immune response against the coronavirus spike protein. One dose (0.5 mL) contains no less than 2.5 × 10 8 infectious units of chimpanzee adenovirus encoding the SARS-CoV-2 spike glycoprotein (ChAdOx1-S) [12] . The CoronaVac vaccine was given as a two-dose regimen administered 21 days apart and prioritized for adults aged 18-59 years. AZD1222 was given as a two-dose regimen administered 10 weeks apart for adults ≥ 18 years and prioritized for the elderly above 60 years of age. Venous blood samples (5 ml) samples were collected at a variety of timepoints after second dose vaccination in participants who received heterologous inactivated and adenoviral-vectored vaccine. For participants who received homologous CoronaVac and AZD1222 vaccines, venous blood samples (5 ml) were collected at 21-35 days after the second dose vaccination. All sera samples were tested for SARS-CoV-2 anti-nucleocapsid (anti-N) and anti-RBD IgG by SARS-CoV-2 IgG II Quant assay (Abbott Diagnostics, Sligo, Ireland). This assay quantifies specific IgG against the RBD and nucleocapsid protein of the spike protein by using a chemiluminescent microparticle immunoassay (CMIA). The anti-RBD IgG result was expressed as arbitrary units per milliliter (AU/mL) and the positive cut-off level was ≥50 AU/mL. The AU/mL unit was converted to binding antibody units per milliliter (BAU/mL) according to the World Health Organization standard. The anti-N IgG results were used to exclude participants in the homologous AZD1222 group (data not shown). The neutralizing activity in a subset of serum samples was tested against the original Wuhan strain and variants of concern, B. RBD IgG titers were presented as geometric mean titer (GMT) and 95% confidence interval (CI). The sVNT ACE2-RBD binding percent inhibition was presented as median with interquartile range. Baseline characteristics including potential confounders such as age, sex and presence of comorbidity were adjusted using ANCOVA with Bonferroni correction. Statistical analysis was performed using SPSS v23.0 (IBM Corp, Armonk, NY). Figures were generated using Prism 8.0 (GraphPad, San Diego, CA). A P-value of <0.05 was considered statistically significant. All participants were Thai. The median age of participants in the homologous AZD1222 vaccine cohort (median 50 years; IQR 33-64) was significantly higher than that in the homologous CoronaVac (median 43 years; IQR 35-48) and the heterologous CoronaVac/AZD1222 (median 38 years; IQR 31-44) (P<0.001) because of vaccine prioritization during initial implementation (Table 1) Table 1) . Differences between all available data and data from participants with similar characteristics among all groups were not significant. Therefore, we presented all available data in the results section. Apart from age, there was a difference in the presence of comorbidities among the three groups (Table 1) . Participants in the homologous AZD1222 group had higher comorbidity rate than the homologous and the heterologous CoronaVac/AZD1222 (P=0.013). Potential confounders including age and presence of comorbidities were taken into account and adjusted in all analyses of this study. Anti-RBD IgG was detected in 100% of participants in all groups after dose 2, with the GMT (95% CI) of 142.8 (118.8-171.6), 562.6 (472.5-670.0), and 165.6 (135.9-201.8) BAU/ml among the homologous CoronaVac, heterologous CoronaVac/AZD1222 and homologous AZD1222 groups, respectively. CoronaVac followed by AZD1222 vaccine induced higher levels of anti-RBD IgG than two-dose CoronaVac (approximately 3.9-fold) and AZD1222 vaccines (approximately 3.4-fold) (P<0.001) (Fig. 1A) . The sVNT was based on antibody-mediated blockage of ACE2-RBD-protein interaction. The percent inhibition represented the ability of sera from vaccinated individuals to block the interaction between the ACE2 receptor protein and the SARS-CoV-2 RBD, which reflects the neutralizing capacity. Our results showed that the heterologous CoronaVac/AZD1222 vaccine recipients had higher neutralizing capacity measured by sVNT against the original Wuhan (wild type) (median = 91.9%) than the homologous CoronaVac (median = 60.2%) and AZD1222 vaccine recipients (median = 74.3%) (Fig. 1B) . Our results showed that sera samples of the heterologous CoronaVac/AZD1222 vaccine recipients had higher neutralizing capacity measured by sVNT against the wild-type and all variants of concern than the recipients of the two-dose CoronaVac (P<0.001) ( Fig. 2A-D) . In addition, the homologous AZD1222 vaccine recipients also had higher neutralizing activities against the wild-type and all variants of concern than in the recipients of the CoronaVac (P<0.001). Comparison of neutralizing activities between the heterologous CoronaVac/AZD1222 and the homologous AZD1222 vaccine recipients showed similar neutralizing capacity against the SARS-CoV-2 wild-type, B.1.1.7 and B.1.617.2 but lower neutralizing capacity against B.1.351 in the homologous AZD1222 group than the heterologous group (P=0.016) after adjustment of age, sex and presence of comorbidities as potential confounders. (Fig. 2C ). Our study showed that there was a reduction in neutralizing capacity of sera As of August 2021, the Department of Disease Control, Ministry of Public Health, Thailand, has implemented policies designating the first vaccine dose to be the CoronaVac vaccine and the second dose to be the AZD1222 vaccine, with more than 300,000 individuals getting vaccinated under this policy. The reasons are the shortage of AZD1222 and the decreased effectiveness of the two-dose CoronaVac vaccine against the SARS-CoV-2 delta variant that are circulating in Thailand. Our study enrolled recipients who had received heterologous inactivated vaccine (CoronaVac) and adenoviral-vectored COVID-19 vaccine (AZD1222) and sought antibody testing following vaccination, and they were compared with homologous vaccine recipients. Our immunogenicity data has shown that anti-RBD IgG was detected in 100% of heterologous CoronaVac/AZD1222 recipients after the second dose, with a higher GMT than those elicited by the two-dose CoronaVac and AZD1222 vaccines after adjusting for age, sex and presence of comorbidities. In addition, the sera samples of the CoronaVac/AZD1222 vaccine recipients had higher neutralizing capacity against the original Wuhan, B.1.1.7, B.1.617.2 and the B.1.351 strain than did the recipients of two-dose CoronaVac. Although the extent of the efficacy of the heterologous regimen has not been studied, the comparatively high level of immunogenicity compared to the homologous regimen supports its use as an alternative schedule. Researchers are investigating to determine the immune correlates of protection to use as surrogate endpoints for vaccine efficacy. In a recent study, binding and neutralizing antibodies were correlated with a reduced risk of symptomatic SARS-CoV-2 infection, for example, anti-RBD IgG level of 506 BAU/mL correlated to the vaccine efficacy of 80% against symptomatic infection with Alpha (B.1.1.7) variant [13] . Because of its ability to elicit a high anti-RBD IgG and neutralizing capacity following two-dose vaccination, a heterologous regimen with CoronaVac/AZD1222 may provide better protection than the homologous CoronaVac regimen. Regarding the reactogenicity of the heterologous regimen, we did not record the adverse events following homologous and heterologous vaccination in this cohort. Nevertheless, our previous prospective cohort study showed that the heterologous CoronaVac/AZD1222 regimen resulted in a higher percentage of local and systemic adverse events compared to the homologous group after the second dose vaccination which is in agreement with the initial reactogenicity data in the Com-COV trial [3, 8] . [14] [15] . A recent study has shown that two doses of AZD1222 had an efficacy of 10.4% against the B.1.351 [16] . Our study also showed a reduction in neutralizing capacity of sera against B. 1.1.7, B.1.167.2 additional benefit against variants of concern in the heterologous CoronaVac/AZD1222 regimen compared to the two-dose CoronaVac regimen. The high immunogenicity of the heterologous CoronaVac/AZD1222 regimen in this study is congruent with conclusions of another heterologous CoronaVac/ adenovirus type-5-vectored Convidecia study [17] . Heterologous administration with adenovirus type-5-vectored vaccine in CoronaVac-primed individuals resulted in improved immunogenicity in both humoral (neutralizing antibody and anti-RBD IgG) and cellular responses (T helper-1-biased response) with an increased in the occurrence of solicited injection site reactions than those receiving the homologous CoronaVac. Heterologous vaccination regimens have been previously examined with experimental vaccines for HIV [18] , malaria [19] , and Ebola [20] , a precedent for this regimen; however, the mechanism for increased immunogenicity from mixing CoronaVac/AZD1222 has yet to be elucidated. There are a few noteworthy limitations to the current study. Because our study participants were recruited in a real-world setting before the Ministry of Public Health launched the vaccine recommendation, the schedule of heterologous vaccination did not follow the recently released guideline stating that two doses should be given 28 days apart. In this study, approximately 80.5% of individuals received the first and second dose of the heterologous regimen at an interval between 14 and 35 days. Secondly, the timing of blood collection after the second dose in the heterologous group was not the same as in the homologous group. There might be residual confounding due to differences in dosing and blood collection intervals. Third, age and presence of comorbidities demographic disparity was present between different vaccination regimen groups. The inherent nature of Thailand's vaccination policy, which prioritizes vaccination with the AZD1222 vaccine in elderly people, consequently led to a higher average age for the homologous AZD1222 regimen cohort. A previous AZD1222 randomized trial showed that anti-spike IgG responses decreased with increasing age after the first dose but not the second dose vaccination [21] . The increased average age in the homologous AZD1222 regimen cohort may lead to a lower immune response as also demonstrated in a study of immunogenicity of an RNA vaccine [22] . The age-related lower immune response is likely due to the "immunosenescence" phenomenon as a result of increases in terminally differentiated memory cell populations, lymph node fibrosis, and altered cytokine production among the elderly [23] . Nevertheless, all of the potential confounders including age, sex and presence of comorbidities were adjusted. This study had a small sample size. Additional studies on a larger group of individuals are required to determine the immunogenicity of the heterologous vaccine in different settings. A surrogate viral neutralization test used in this study is a robust serological test to detect neutralizing capacity to SARS-CoV-2 with 99.93% specificity and 95-100% sensitivity [24] but it could not be directly compared with the live virus neutralization test in other studies. Lastly, cell-mediated immunity was not explored. In conclusion, a heterologous CoronaVac/AZD1222 regimen demonstrated a robust anti-RBD IgG and neutralizing antibody response. The heterologous regimen with CoronaVac followed by AZD1222 would provide greater flexibility for countries experiencing supply difficulties and individuals with adverse events following the first dose CoronaVac vaccination. All data generated during this study are contained within this manuscript and its ☐ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. ☒ The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Yong Poovorawan reports financial support was provided by The National Research Council of Thailand (NRCT). Yong Poovorawan reports financial support was provided by Health Systems Research Institute. Nungruthai Suntronwong reports financial support was provided by the second Century Fund (C2F) fellowship of Chulalongkorn University. Yong Poovorawan reports financial support was provided by The Center of Excellence in Clinical Virology of Chulalongkorn University. 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