key: cord-0279677-yb6jk5j0
authors: Al Jurdi, A.; Benedetti Gassen, R.; De Jesus Borges, T.; Tadeval Lape, I.; Morena, L.; Efe, O.; Solhjou, Z.; El Fekih, R.; Deban, C.; Bohan, B.; Pattanayak, V.; Kotton, C.; Azzi, J.; Riella, L. V.
title: Diminished antibody response against SARS-CoV-2 Omicron variant after third dose of mRNA vaccine in kidney transplant recipients
date: 2022-01-06
journal: nan
DOI: 10.1101/2022.01.03.22268649
sha: 8e6961971eb54fe4564d821c3b74f5d0ec41f12a
doc_id: 279677
cord_uid: yb6jk5j0

Abstract: Background: Available SARS-CoV-2 vaccines have reduced efficacy against the Omicron variant in immunocompetent individuals. Kidney transplant recipients (KTRs) have diminished antiviral responses to wild-type SARS-CoV-2 after vaccination, and data on antiviral responses to SARS-CoV-2 variants, including the Omicron variant, are limited. Methods: We conducted a prospective, multi-center cohort study of 51 adult KTRs who received three doses of BNT162b2 or mRNA-1273. Blood and urine samples were collected before and four weeks after the third vaccine dose. The primary outcome was anti-viral antibody responses against wild-type and variants of SARS-CoV-2. Secondary objectives included occurrence of breakthrough SARS-CoV-2 infection and non-invasive monitoring for rejection using serum creatinine, proteinuria, donor-derived cell-free DNA and donor-specific antibodies. Sera from pre-pandemic healthy controls and KTRs were used for comparison. Results: 67% of KTRs developed anti-wild-type spike antibodies after the third vaccine dose, similar to the Alpha (51%) and Beta (53%) variants, but higher than the Gamma (39%) and Delta (25%) variants. No KTRs had neutralizing responses to the Omicron variant before the third vaccine dose. After the third dose, fewer KTRs had neutralizing responses to the Omicron variant (12%) compared to wild-type (61%) and Delta (59%) variants. Three patients (6%) developed breakthrough SARS-CoV-2 infection at a median of 89 days. No KTRs developed allograft injury, de novo donor-specific antibodies or allograft rejection. Conclusion: In KTRs, a third dose of mRNA vaccines increases antibody responses against wild-type and variants of SARS-CoV-2, while neutralizing responses to the Omicron variant remain markedly reduced.

Coronavirus disease (COVID-19) caused by severe acute respiratory coronavirus 2 (SARS-CoV-2) has resulted in over 800,000 deaths in the United States 1 with the Delta (B.1.617.2) and more recently the Omicron (B.1.1.529) variants accounting for the majority of cases. 2 Kidney transplant recipients (KTRs) are at increased risk of mortality following SARS-CoV-2 infection, 3 and develop blunted antiviral responses following SARS-CoV-2 vaccination compared to nontransplant patients. [4] [5] [6] Furthermore, the Delta and Omicron variants have been shown to be less sensitive to neutralizing antibodies from sera of vaccinated immunocompetent individuals, 7, 8 and similar data from vaccinated KTRs are limited. 9 Therefore, in an era of clinically prevalent SARS-CoV-2 variants, evaluating antiviral responses to both wild-type (WT) and variants of SARS-CoV-2 in KTRs following SARS-CoV-2 mRNA vaccination is crucial. The aim of this study was to evaluate antiviral humoral responses against SARS-CoV-2 variants, including the Delta and Omicron variants following the third dose of SARS-CoV-2 mRNA vaccination in KTRs.

This is a prospective multicenter observational cohort study of the antiviral immune responses against SARS-CoV-2 variants following the third dose of SARS-CoV-2 mRNA vaccination in adult KTRs. Inclusion criteria were KTRs aged ≥18 years who were >3 months posttransplantation with stable allograft function and no rejection in the preceding six months. Full exclusion criteria are listed in supplementary materials. Subjects were enrolled consecutively.

. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint Enrolled KTRs had received two doses of mRNA-1273 or BNT162b2 SARS-CoV-2 mRNA vaccines and were planned to receive a third dose of the same vaccine as per national guidelines.

Participants had a baseline visit prior to the third vaccine dose and a follow-up visit one month following the third vaccine dose, where blood and urine samples were collected. Samples from fifteen pre-pandemic kidney transplant control patients and five pre-pandemic healthy controls were used for comparison.

The study was approved by the institutional review board at Mass General Brigham (IRB 2021P000043). All subjects signed written informed consent forms prior to enrollment in the study. The study was conducted in accordance with the Declaration of Helsinki and the Declaration of Istanbul. Data are reported in compliance with the STROBE statement reporting guidelines.

The primary outcome was the development of antiviral humoral responses following the third vaccine dose as assessed by 1) measurement of antibodies directed against the spike protein and receptor-binding domain (RBD) of WT and Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2) and Omicron (B.1.1.529) variants of SARS-CoV-2 by a Luminex-based multiplex assay and by enzyme-linked immunosorbent assay (ELISA); and 2) Antibodies neutralization capacity against WT, Delta and Omicron variants of SARS-CoV-2 using a surrogate virus neutralization test. 10 Secondary objectives included 1) the occurrence of any severe or grade 4 related adverse events; 2) development of breakthrough SARS-CoV-2 infection; and 3) monitoring for rejection using serum creatinine, urine protein-to-creatinine ratio, donor-derived cell-free DNA . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint and donor-specific antibodies (DSAs). Details of sample processing and assays are described in supplementary materials and methods.

Continuous variables are presented as means (± standard deviation) or as medians (with interquartile ranges or full ranges) depending on normality of distribution. Categorical variables are presented as frequencies and percentages. Differences between paired samples were assessed using a paired t-test, a Wilcoxon matched-pairs signed rank test, a repeated measures ANOVA or Friedman test as appropriate. For comparisons between continuous variables between three or more groups, if testing reached statistical significance, then pairwise testing was performed to determine significant differences between groups, using Dunn's correction to adjust for multiple comparisons. For categorical variables, the differences in proportions were calculated using a Pearson's Chi squared test or Fisher's exact test as appropriate. All tests used were two-sided and a two-sided α level of 0.05 was considered to be statistically significant. SPSS v24 (Chicago, IL) and GraphPad Prism v9.1.2 (San Diego, CA) were used for statistical analysis and creation of figures.

Fifty-one vaccinated KTRs were enrolled in the study. Baseline characteristics are shown in Table   1 . Median age was 63 years, 43% were female and forty-eight (94%) patients received the BNT162b2 vaccine. The median time between the second and third vaccine doses was 187 days . Baseline samples were collected at a median of 5 days (IQR 0-18) prior to the . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint third vaccine dose and post-vaccination samples were obtained at a median of 29 days (IQR 26-33) post-vaccination.

To evaluate antibody-mediated antiviral responses, we measured antibodies directed against the nucleocapsid (NC), spike trimer, S1 and RBD regions of the WT SARS-CoV-2 prior to and following the third vaccine dose in KTRs using a Luminex-based multiplex assay. After the third vaccine dose, there was no significant change in the MFI ratios of anti-NC antibodies (p=0.278) but there was a significant increase in the MFI ratios for antibodies directed against the spike trimer, S1 and RBD regions (p<0.001 for all, Fig. 1A ). Using the recommended assay positivity threshold, all KTRs had a negative anti-NC antibody result before and after the third vaccine dose.

The percentage of KTRs with anti-WT spike antibodies increased from 29% to 67% (p<0.001), anti-WT S1 antibodies increased from 10% to 47% (p<0.001) and anti-WT RBD antibodies increased from 12% to 45% (p<0.001) after the third vaccine dose. In comparison, all prepandemic healthy controls (HCs) and pre-pandemic kidney transplant patient controls (KCs) had negative results for antibodies against all four WT SARS-CoV-2 antigens (Fig. 1B) . After the third vaccine dose, there was a significant increase in the MFI ratios of anti-spike antibodies directed against the Alpha, Beta, Gamma and Delta variants (p<0.001 for all, Fig. 1C ), while all prepandemic HCs and KCs had undetectable antibodies against those variants (Fig. 1D ). When evaluating differences in anti-spike antibody responses to SARS-CoV-2 variants, we found that prior to the third vaccine dose a higher percentage of KTRs (29%) had anti-spike antibodies against the WT virus compared to the Alpha (12%, p=0.028), Gamma (6%, p=0.002) and Delta (2%, p<0.001) variants but not the Beta variant (22%, p=0.364, Fig. 1E ). After the third vaccine dose, . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint a higher percentage of KTRs (67%) had anti-spike antibodies against the WT virus compared to the Gamma (39%, p=0.006) and Delta (25%, p<0.001) variants but not the Alpha (51%, p=0.108) or Beta variants (53%, p=0.158, Fig. 1F ).

Anti-RBD antibody measurement against Omicron variant and its neutralization capacity We then evaluated neutralizing antibody responses using a surrogate virus neutralization test (SVNT). 10 We found a significant increase in the percentage of neutralization against the WT (p<0.001), Delta (p<0.001) and Omicron variants (p<0.001, Fig. 2C ) after the third vaccine dose.

However, the percentage of neutralization was significantly lower for the Omicron variant compared to the WT and Delta variants (p<0.001). In comparison, there was no significant neutralizing response against the WT, Delta and Omicron variants in pre-pandemic HCs and KCs (Fig. 2D) .

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In addition, we found that the percentage of KTRs who had a neutralizing response increased from 29% to 61% for WT virus (p=0.002), from 24% to 59% for the Delta variant (p<0.001), and from 0% to 12% for the Omicron variant following the third vaccine dose (p=0.012, Fig. 2E ). Prior to the third vaccine dose, fewer KTRs had a neutralizing response against the Omicron variant, is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint change in serum creatinine (p=0.212), urine protein-to-creatinine ratio (p=0.257), or donor-derived cell-free DNA levels (p=0.434, Table S1) compared to pre-vaccination. At a median follow-up of 89 days (IQR 78-100) after the third vaccine dose, no patients developed allograft rejection.

This study is one of the first to characterize antiviral humoral responses against SARS-CoV-2 variants, including the Omicron variant, after a third dose of SARS-CoV-2 mRNA vaccines in KTRs. Using a Luminex-based multiplex assay, we found that two thirds of KTRs had anti-WT spike antibodies after the third vaccine dose, similar to previous reports. 11 -16 Despite an increase in the proportion of KTRs with anti-spike antibodies against WT SARS-CoV-2 and variants after the third vaccine dose, fewer KTRs had anti-spike antibodies against the Gamma and Delta variants compared to the WT, Alpha and Beta variants. Using an ELISA we developed in our laboratory, we found an increase in anti-RBD antibody levels against the WT, Delta and Omicron variants after the third vaccine dose, although anti-RBD antibody levels were significantly lower for the Omicron variant compared to WT virus. To assess neutralization capacity of antibody responses, we used a SVNT, which has been shown to be highly correlated with live virus neutralization assays. 10 We found an increase in the percentage of neutralization against the WT, Delta and Omicron variants after the third vaccine dose as has been reported in immunocompetent individuals. 17 is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint Omicron variant after the third vaccine dose. The Omicron variant's ability to escape neutralization compared to WT SARS-CoV-2 in our study is consistent with data from vaccinated immunocompetent individuals 8, 17, 20, 21 and is likely due to its highly mutated RBD region, which has 15 mutations compared to WT RBD. [22] [23] [24] It is worth noting that a limitation of the SVNT is that it is not able to measure neutralizing antibodies directed against non-RBD regions of the spike protein as it only measures RBD-ACE2 interactions.

We were thus able to provide a detailed characterization of antibody responses to SARS-CoV-2 variants in KTRs, including the Omicron variant, and to evaluate the alloimmune safety of a third vaccine dose and found no evidence of allograft injury, de novo DSA development and no episodes of allograft rejection. This is consistent with what has been reported after two [25] [26] [27] and three vaccine doses 11,12,14 in solid organ transplant recipients. Our study has limitations, including its small sample size, observational design, lack of a control group of KTRs who did not receive a third vaccine dose, and lack of assessment of the cellular response to vaccination. Further studies evaluating the cellular response to the Omicron variant and the implications of the reduced neutralization ability with regards to risk and severity of infections in KTRs are needed.

In summary, we found that a third dose of SARS-CoV-2 mRNA vaccination in KTRs was associated with an increased antiviral antibody response against WT and variants of SARS-CoV-2, and while the neutralizing responses to the Omicron variant increased in some, overall they remained markedly diminished. Strategies designed to improve antiviral immune responses to the Omicron and future variants, such as with additional homologous 28 or heterologous 29 vaccine doses or use of higher vaccine doses, are also needed in this vulnerable high-risk group, in particular with the surge of Omicron as the predominant variant worldwide.

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The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. ; https://doi.org/10.1101/2022.01.03.22268649 doi: medRxiv preprint

We would like to thank the staff of the transplant clinic for their assistance in conducting this study.

The authors have declared that no conflict of interest exists.

The study was funded by CareDx, Inc. (Brisbane, CA) grant number 2021A008053 to LVR and JRA. The study was also supported in part by the Harold and Ellen Danser Endowed/Distinguished Chair in Transplantation at Massachusetts General Hospital (Boston, MA, USA). This was an investigator-initiated research project where the design and conduct of the study was determined by the investigator without influence from the funders.

Data to support the findings in the study are available from the corresponding author upon reasonable request.

. CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. History of allograft rejection, n (%) 11 (22) . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.

The copyright holder for this preprint this version posted January 6, 2022. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review)

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Assessment of 4 Doses of SARS-CoV-2 Messenger RNA-Based Vaccine in Recipients of a Solid Organ Transplant

Serum creatinine (mg/dL), median (IQR) 2 nd vaccine dose, median (IQR)

Days between 2 nd and 3 rd vaccine dose, median (IQR)

ALC at time of 3 rd vaccine dose, cells/mm 3 , median (IQR)

ACE: Angiotensin converting enzyme. ALC: Absolute lymphocyte count. ARB: Angiotensin receptor blocker. CMV: Cytomegalovirus. EBV: Epstein-Barr virus. ESKD: end-stage kidney disease. GFR: glomerular filtration rate. HLA: human leukocyte antigen. KDPI: kidney donor profile index. mTOR: mammalian target of rapamycin. PRA: panel reactive antibodies. RRT: renal kidney transplant recipients (KTRs). (A) Anti

529) variants of SARS-CoV-2 before and after the third vaccine dose in KTRs (n=51), and (B) in pre-pandemic healthy controls (HCs, n=5) and prepandemic kidney transplant control patients (KCs, n=15) measured by enzyme-linked immunosorbent assay. (C) Percentage of neutralization to WT, Delta and Omicron variants of SARS-CoV-2 before and after the third vaccine

HCs (n=5) and KCs (n=15) measured by surrogate virus neutralization test. Horizontal lines indicate positivity threshold for assay. (E) Proportion of KTRs with a positive neutralization response before and after the third vaccine dose in KTRs

Dunn's correction for multiple comparisons. (E) Statistic by Chi-square test