key: cord-0693816-wmixrj1t authors: Benning, Louise; Morath, Christian; Bartenschlager, Marie; Kim, Heeyoung; Reineke, Marvin; Beimler, Jörg; Buylaert, Mirabel; Nusshag, Christian; Kälble, Florian; Reichel, Paula; Töllner, Maximilian; Schaier, Matthias; Klein, Katrin; Benes, Vladimir; Rausch, Tobias; Rieger, Susanne; Stich, Maximilian; Tönshoff, Burkhard; Weidner, Niklas; Schnitzler, Paul; Zeier, Martin; Süsal, Caner; Hien Tran, Thuong; Bartenschlager, Ralf; Speer, Claudius title: Neutralizing antibody response against the B.1.617.2 (delta) and the B.1.1.529 (omicron) variants after a third mRNA SARS‐CoV‐2 vaccine dose in kidney transplant recipients date: 2022-04-18 journal: Am J Transplant DOI: 10.1111/ajt.17054 sha: aba3cfd572caa2c8c04a5c3a32ac17e6b75f82db doc_id: 693816 cord_uid: wmixrj1t Seroconversion after COVID‐19 vaccination is impaired in kidney transplant recipients. Emerging variants of concern such as the B.1.617.2 (delta) and the B.1.1.529 (omicron) variants pose an increasing threat to these patients. In this observational cohort study, we measured anti‐S1 IgG, surrogate neutralizing, and anti‐receptor‐binding domain antibodies three weeks after a third mRNA vaccine dose in 49 kidney transplant recipients and compared results to 25 age‐matched healthy controls. In addition, vaccine‐induced neutralization of SARS‐CoV‐2 wild‐type, the B.1.617.2 (delta), and the B.1.1.529 (omicron) variants was assessed using a live‐virus assay. After a third vaccine dose, anti‐S1 IgG, surrogate neutralizing, and anti‐receptor‐binding domain antibodies were significantly lower in kidney transplant recipients compared to healthy controls. Only 29/49 (59%) sera of kidney transplant recipients contained neutralizing antibodies against the SARS‐CoV‐2 wild‐type or the B.1.617.2 (delta) variant and neutralization titers were significantly reduced compared to healthy controls (p < 0.001). Vaccine‐induced cross‐neutralization of the B.1.1.529 (omicron) variants was detectable in 15/35 (43%) kidney transplant recipients with seropositivity for anti‐S1 IgG, surrogate neutralizing, and/or anti‐RBD antibodies. Neutralization of the B.1.1.529 (omicron) variants was significantly reduced compared to neutralization of SARS‐CoV‐2 wild‐type or the B.1.617.2 (delta) variant for both, kidney transplant recipients and healthy controls (p < .001 for all). Due to waning humoral immunity and a rapid increase in breakthrough infections, a third vaccine dose was recommended for the general population, including immunocompromised individuals with impaired vaccination response. [9] [10] [11] First results describe an increased immune response in transplant recipients to a third vaccine dose with an induction of serologic response in 25%-49% of previous non-responders and a significant increase in antibody titers for patients who were seropositive already before the third dose. [12] [13] [14] [15] Emerging variants of concern (VoCs), such as the B.1.617. 2 (delta) and the B.1.1.529 (omicron) variants with partial immune escape are rapidly displacing other circulating strains and increasingly lead to breakthrough infections. In particular, the omicron variant escapes antibody neutralization and early real-world data indicate reduced effectiveness and reduced protection from hospitalization after two-dose vaccination or prior infection in the general South African population where omicron was first described. 16 18, 19 We recently first demonstrated impaired neutralization of the We used the SARS-CoV-2 Total Assay (Siemens) and the Elecsys anti-SARS-CoV-2 assay (Roche) to detect anti-S1 IgG and antinucleocapsid antibodies, respectively. A semi-quantitative index of ≥1 defines positivity for both assays. Surrogate neutralizing antibodies were detected using a surrogate virus neutralization test (Medac) that mimics the virus interaction with the host cell by direct protein-protein interaction using purified RBD protein from the viral spike and the ACE-2 host cell receptor. 21 An inhibition ≥30% of RBD:ACE-2 binding defines positivity for this assay. A bead-based multiplex assay for the Luminex platform (LabScreen Covid Plus, One Lambda Inc.) was used to detect IgG antibodies against four different SARS-CoV-2 target epitopes and IgG antibodies against the spike S1 of four common cold coronaviruses. 22 The mean fluorescence intensity (MFI) was analyzed using a Luminex 200 device (Luminex Corporation). All assays were performed according to the manufacturer's in- SARS-CoV-2 genomes were sequenced with the ARTIC protocol using the NEBNext ARTIC FS kit to prepare sequencing libraries. To increase sample throughput the I.DOT liquid dispenser (Dispendix) was employed, and 4 × 96 libraries were pooled for sequencing in paired-end mode (2 × 75 bp) on a NextSeq instrument. Sequencing adapters were trimmed using trim_galore and host-read contamination was assessed and filtered using kraken2, as described previously. 31, 32 Reads were aligned to the SARS-CoV-2 reference genome using bwa and alignments were sorted and indexed using samtools and quality-controlled using alfred. [33] [34] [35] Priming regions were masked with iVar, followed by variant calling with FreeBayes, normalization of variants with bcftools, and annotation of variants with the Ensembl Variant Effect Predictor (VEP). [36] [37] [38] [39] iVar was employed to compute a viral consensus sequence that was then classified by Pangolin and Nextclade to determine the viral lineage and clade, respectively. 40,41 The statistical analysis was performed using GraphPad Prism version 9.0.0 (GraphPad Software). Data are given as median and interquartile range (IQR) or number (N) and percent (%). Continuous variables were compared using the Mann-Whitney U test for unpaired or the Wilcoxon matched-pairs rank test for paired variables. When comparing more than three paired continuous variables, we applied the Friedman's test with Dunn's post-test. Categorial data were compared using the Fisher's exact test. Spearman's rho was calculated to describe the correlation of different commercially available assays to the current gold standard to assess humoral immunity using a live virus assay. Statistical significance was assumed at a p < .05. The funding source did not affect in the trial design, conduct, or reporting of this study. respectively, in healthy controls (p < .001 for all; Figure 1A ). IgG antibodies against the full spike, the spike S1, and the spike S2 subunits as determined by a bead-based multiplex assay were also significantly lower in kidney transplant recipients with a median (IQR) MFI of 19,084 (0-22,419) for the full spike, 9482 (0-15,117) for the spike S1, and 1716 (0-6,042) for the spike S2, respectively, when compared to healthy controls ( Figure 1B ). Healthy controls also exhibited significantly higher IgG antibodies against the four tested common cold coronaviruses, but the difference of antibody response between healthy controls and kidney transplant recipients was less pronounced than for SARS-CoV-2 (p = .003 for HCoV-229E, p = .004 for HCoV-HKU1, p = .003 for HCoV-NL63, and p < .001 for HCoV-OC43; Figure S1 ). All 25/25 (100%) healthy controls were seropositive for antispike S1 IgG, surrogate neutralizing antibodies, and anti-RBD antibodies after a third vaccine dose whereas only 26/49 (53%) kidney transplant recipients showed concurrent seropositivity in all three commercially available tests ( Figure 1C ). Figure 2B ). Corresponding interindividual changes in anti-S1 IgG, surrogate neutralizing, and anti-RBD antibodies are displayed in Figure S2 . Figure 3B ). Anti-S1 IgG showed the strongest correlation to neutralization of the B.1.1.529 (omicron) variants with a Spearman's rho of 0.85 ( Figure 3C ). which they also attribute to progressive dose reduction. 52 The two patients of our cohort with Belatacept maintenance therapy showed low antibody levels or no seroconversion which has been described previously, even after a third mRNA vaccine dose. 53 The administration of a fourth vaccine dose is another attempt to optimize vaccination response in kidney transplant recipients, ideally using a vaccine formulation that is based on more recently circulating variants such as B.1.1.529 (omicron). First results indicate an improved humoral response after a fourth vaccine dose among those with a weak response after three doses but little to no improvement among those with no response after three doses. 54, 55 Although this may suggest immunogenic potential for poor responders after a third vaccine dose, additional actions seem necessary to reach vaccine-induced immunity and protection from severe disease courses. 55 Passive immunization of those patients that do not mount immune response at all with therapeutic antibodies that have shown to inhibit SARS-CoV-2 and variants of concern including the B.1.1.529 (omicron) variants is another attempt to protect these patients from severe COVID-19 disease. 56, 57 Our data show that anti-spike S1 IgG and other commercially available tests may aid in clinical decision-making for additional booster vaccination(s) as these tests show a strong correlation to live-virus neutralization which is unfortunately not yet feasible in clinical routine. A strong correlation between commercially available tests and live-virus neutralization has been described previously by us and others in different cohorts. 20, 26, 27, [58] [59] [60] A limitation of our study is the lack of data on B and T cell responses after vaccination. Recent studies provided evidence on highly reproducible whole-blood assays to detect SARS-CoV-2 spike specific T cell response, using a similar platform to assays measuring T cell specific responses against Mycobacterium tuberculosis. 49, 50, 61 A strong correlation between anti-RBD antibodies and SARS-CoV-2 specific IFN-y T cell response was shown for healthy cohorts, immunosuppressed patients with rheumatoid arthritis, and multiple sclerosis patients on various disease-modifying therapies. 49, 50, 62 Similarly, in kidney and liver transplant recipients, a strong correlation between quantitative and functional CD4+ T-cell responses and anti-S1 IgG antibodies was demonstrated after two vaccine doses. 6, 52 These results suggest that anti-spike titers may be used as a surrogate parameter to assess immunologic response after COVID-19 vaccination as T cell studies are more resource intensive and less standardized between different laboratories. 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