key: cord-0910348-gjrgrsue
authors: Ram, Ron; Hagin, David; Kikozashvilli, Nino; Freund, Tal; Amit, Odelia; Bar-On, Yael; Beyar-Katz, Ofrat; Shefer, Gabi; Moshiashvili, Miguel Morales; Karni, Chen; Gold, Ronit; Kay, Sigi; Glait-Santar, Chen; Eshel, Rinat; Perry, Chava; Avivi, Irit; Apel, Arie; Benyamini, Noam; Shasha, David; Ben-Ami, Ronen
title: Safety and Immunogenicity of the BNT162b2 mRNA Covid-19 Vaccine in Patients after Allogeneic HCT or CD19-based CART therapy – a Single Center Prospective Cohort Study
date: 2021-06-30
journal: Transplant Cell Ther
DOI: 10.1016/j.jtct.2021.06.024
sha: 533c12fb371315cb42d34f6971c44d7efb63d631
doc_id: 910348
cord_uid: gjrgrsue

Data are scarce regarding both safety and immunogenicity of the BNT162b2 mRNA COVID-19 vaccine in patients undergoing immune cell therapy, thus we prospectively evaluated these 2 domains in patients receiving this vaccine after allogeneic HCT (n=66) or after CD19-based CART therapy (n=14). Overall, vaccine was well tolerated, with mild non-hematologic vaccine-reported adverse events in minority of the patients. 12% (after first dose) and 10% (after second dose) of the patients developed cytopenia and there were 3 cases GVHD exacerbation after each dose. A single case of impending graft rejection was summarized as possibly related. Evaluation of immunogenicity showed that 57% of patients after CART infusion and 75% patients after allogeneic HCT had evidence of humoral and/or cellular response to the vaccine. On cox regression model, longer time from infusion of cells, female sex, and higher CD19(+) cells were associated with a positive humoral response, whereas higher CD4(+)/CD8(+) ratiowas correlated with a positive cellular response, confirmed by ELISpot test. We conclude that BNT162b2 mRNA COVID-19 vaccine has impressive immunogenicity in patients after allogeneic HCT or CART. Adverse events were mostly mild and transient, but some significant hematologic events were observed, hence, patients should be closely monitored.

Coronavirus 2 (SARS-CoV-2) and has variable presentations. An increased risk for severe disease and death has been noted among patients after allogeneic hematopoietic cell transplantation (HCT), with a case fatality rate of 9%-30% 1, 2 . The BNT162b2 mRNA COVID-19 (Pfizer/BioNTech) vaccine was recently approved by both the FDA and EMA for the prevention of COVID-19, based on a phase 3 study that showed 94.6% efficacy 3 . While this vaccine is recommended by the FDA, EBMT and NMDP for immunosuppressed patients, data are scarce regarding vaccine effectiveness and safety in patients undergoing immune cell therapy. It is reasonable to assume that, like the response to other vaccines, patients after cell therapy would demonstrate a lower response rate compared to the general population 4, 5 . In addition, immunologic alterations that may result in exacerbation of graft vs. host disease (GVHD) and other immune phenomena, are also potential concerns. In this study, we aimed to evaluate the safety and immunogenicity of the BNT162b2 mRNA COVID-19 vaccine in patients who underwent either allogeneic HCT or CD19-based chimeric antigen receptor T (CART) cell therapy.

This was a prospective study performed at the BMT Long-Term Follow-up (LTFU) clinic, Tel Aviv Sourasky Medical Center. All sequential patients that underwent allogeneic HCT and CART therapy in the center are followed in the LTFU clinic. Patients were eligible for this study if they fulfilled the EBMT criteria for COVID-19 vaccination (COVID-19 vaccines, Version 2.0 December 21,2020) including: age>18 years and at least 3 months interval between cells infusion and referral to vaccination. In patients after CART infusion, if the CD19+ cells blood level was 0 after 3 months, vaccination was deferred to 6 months after CART infusion. After 6 months, patients were vaccinated irrespectively to the CD19+ cells blood level. In addition, our protocol exclusion criteria included grade 3-4 acute GVHD, treatment for acute or chronic GVHD with ≥0.5 mg/kg of prednisone (or an equivalent steroid formula), treatment with rituximab within the previous 6 months, treatment with mesenchymal cells within 1-month, hematologic relapse, treatment with maintenance therapy (excluding tyrosine-kinase inhibitors), previous infection with SARS-CoV-2 or recent exposure to a SARS-CoV-2-infected person, and known allergy to vaccine components. In case of GVHD exacerbation, patients were not eligible for the second dose of vaccine until they returned to the baseline status of GVHD. The study was approved by the hospital ethics committee (#1067-20) and was registered in ClinicalTrials.gov (NCT04724642). All patients signed informed consent prior to enrolment.

Patients were vaccinated through the national Israeli vaccination program that started in mid-December 2020. All patients had a baseline serology test to detect anti-nucleocapsid antibodies to ensure a SARS-CoV-2 negative status and a baseline quantification of peripheral blood CD19 + , CD4 + , and CD8 + cells. Prior to first dose of BNT162b2 mRNA COVID-19 vaccine, patients were reassessed for suitability to vaccination, including physical examination, assessment of GVHD status, complete blood count and liver function tests. One week after the administration of the first vaccine dose, patients were interviewed for post-vaccination adverse events, underwent physical evaluation and repeated laboratory tests, and were then scheduled for their second vaccine dose. Patients were reassessed 7-14 days after the second vaccine dose and had a blood test for SARS-CoV-2 serology and ELISpot assay. Patient demographics, disease characteristics and GVHD parameters were collected prospectively. Concomitant immunosuppressive therapy (IST) was also documented, and we defined high intensity IST (as opposed to low intensity) as either prednisone dose of ≥ 0.25 mg/kg/day or another IST medication.

The primary endpoint was the incidence of grade 3-4 adverse events and GVHD exacerbation.

Secondary endpoints included overall adverse effects and humoral and cell-mediated response to vaccine (measured by anti S IgG and ELISpot tests, respectively).

We graded adverse events according to CTCAE v5.0, acute GVHD according to the MAGIC criteria, and chronic GVHD according to the NIH 2014 grading and response criteria 6 . Causality of adverse events was defined according to the WHO-UMC categories (certain, probable, possible, and unlikely) (http://who-umc.org/Graphics/24734.pdf).

For Serology detection of antibodies and ELISpot assay see Supplement. 

Continuous variables were described as the mean, median, standard deviation and range of values, as applicable. Categorical data were described with contingency tables including frequency and percent. Antibody titers were compared between patient groups using either Pearson Chi-Square or t test, as appropriate. The association of various parameters with the serology/ ELISpot test results was performed using Bilinear Logistic Regression with a twosided P value of <.05 considered to be statistically significant. IBM SPSS Statistics, version 27 was used to perform all analyses. (Figure 4b) . Of note, there was no evidence of GVHD, and she was still receiving low dose cyclosporine. To investigate this, we performed bone marrow aspiration that showed trilineage hematopoiesis with no evidence for increased blast percentage, and hypolobulated megakaryocytes. The latter finding was not noted in a previous bone marrow evaluation.

Cytogenetic analysis showed normal karyotype, however the percentage of donor's chimerism gradually dropped from 97% to 59%. Cyclosporine was stopped and she received 2 separate doses of DLI (1x10^6/ kg and 5x10^6/kg), Figure 4b . This impending secondary rejection was considered as possibly vaccine-associated adverse event.

GVHD exacerbation -After the first vaccine dose, there were 3 cases (4.5%) of exacerbation of GVHD, all developed within the 1st week after injection (grade 2 oral GVHDresolved 18 days after intervention with steroid mouth wash (n=1); grade 2 liver and grade 1 oralresolved within a week with no intervention (n=1); and grade 2 lower gutresolved within 3 weeks after a short steroid course of prednisone 0.25 mg/kg per day (n=1). All 3 patients subsequently received the second dose. After the second dose of vaccine there were also 3 cases of exacerbation of GVHD, all occurred within 1 week after injection (grade 2 liver and arthralgia-resolved within 2 weeks with a short low dose steroids course (n=1); grade 3 fasciitis and skin rashreturned to base line after increasing prednisone dose from 0.15 mg/kg to 0.25 mg/kg for 2 weeks; grade 2 oral GVHDresolved within 2 weeks with steroid-based mouth wash (n=1)). Since all patients had a stable chronic GVHD for at least a month prior to vaccination, we considered all cases of GVHD exacerbation to be possibly related to vaccination.

Three patients (all after allogeneic HCT) that developed COVID-19 infection after the first vaccine dose were excluded from this analysis. All 14 patients after CART infusion were evaluated by serology and 12 (86%) by ELISpot assay, while among the patients after allogenic HCT (n=63), 57 (90%) and 37 (59%) patients, respectively, were evaluated. On multivariate analysis, a positive humoral response to the vaccine was associated with increased time from infusion of cells (p=.032), female sex (p=.028), and higher number of CD19 + cells (p=.047), while age, active GVHD and intensity of concomitant IST did not predict response, Table 2 . Focusing on the subgroup of patients after CART infusion revealed that patients with B cell reconstitution had a higher incidence of positive serology, compared to those with B cell aplasia (66% vs. 11%, p=.025). In the allogeneic HCT group, there was only 1 patient with complete B cell aplasia and this patient had a negative serology test.

Because of low number of positive ELISpot results we did not perform analysis to identify potential predictors; however, ELISpot results were correlated with the CD4 + /CD8 + ratio (Pearson correlation=.54 (95% CI .29-.72), p<.001), while the number of CD19 + , CD4 + , and CD8 + cells did not significantly correlate with the probability of positive test results.

In this study we evaluated the safety profile and response to the BNT162b2 mRNA COVID-19 vaccine. To our knowledge, this is the first study that focused on patients after cell therapy, including allogeneic HCT and CART therapy. We showed that in this population the vaccine was relatively safe and there were no grade 3-4 non-hematologic adverse events. However, ~5% of the patients developed transient grade 3-4 cytopenia. GVHD exacerbation was noted in ~5% of the patients and was easily controlled. Remarkably, humoral response was documented in 82% of the patients after allogeneic HCT. Perhaps even more surprising, humoral response was observed in 36% of patients after B-cell depleting CART therapy, and 50% of those had evidence for cellular response. After measuring overall (both humoral and cellular) response, an in vitro immune response was documented in 75% and 57% of allogeneic HCT and CART recipients, respectively. Patients after allogeneic HCT or CART therapy are often immunosuppressed for months and years due to conditioning regimens, maintenance therapy, immunosuppressive medications, persistent hypogammaglobulinemia, and GVHD; all blunting immune response and decreasing vaccine efficacy 7, 8 . While these patients are at a high risk for complications from viral infections, the efficacy of both specific anti-viral therapy as well as vaccination is inferior compared to healthy subjects. The mRNA SARS-CoV-2 vaccines were administered in the pivotal trials, and safety data has not raised any significant concerns 3, 9 . Similarly to the reported data, nonhematologic adverse events were mild in our cohort, resolved within 2 days, and we did not observe any grade 3-4 non-hematologic toxicity 9 .

In contrast, hematologic/immunologic adverse events may be of concern in the context of cell therapy. Studies performed in healthy individuals showed that in concurrent with the production of neutralizing antibodies and the stimulation of virus-specific CD4 + and CD8 + T cells, there was a robust release of immune-modulatory cytokines, such as IFN-γ, TNF, IL-1, IL-2, and IL-12 10, 11 .

These pro-inflammatory cytokines may alter T cell function and induce, or exacerbate, GVHD 12 .

In our cohort, ~5% of allogeneic HCT recipients had exacerbation of GVHD. Half of the patients required systemic steroids to control the exacerbation and all exacerbations resolved within several weeks. In line with that, we observed grade 3-4 cytopenia in ~5% of the patients, mainly thrombocytopenia. Although rare, this phenomenon was also reported in healthy individuals, and it was postulated that these individuals may have pre-formed antibodies against components of the vaccine nanoparticles or impaired platelet production as part of the post vaccination systemic inflammatory response 13, 14 . We favour the latter explanation in our cohort, as all patients spontaneously recovered counts and were closely monitored for months prior to vaccination, suggesting that the possibility of pre-formed antibodies is unlikely.

The single case of impending late graft rejection is worrisome. Although vaccination-associated rejection has been extensively reported in recipients of solid organ transplants, it has been rarely reported after HCT 15 . Possible mechanisms that facilitate rejection after vaccination are indirect activation by cytokines or NK-cells, and alternating endothelial progenitor cells that impair the bone marrow microenvironment and subsequently accelerate the rejection process 16, 17 . In the single case in our cohort, the patient received repeated doses of DLI and is being monitored weekly for both toxicity and marrow recovery.

In the current study we used 2 methods to test the immunogenicity of BNT162b2. While humoral immunity evaluation by serology testing is simple and relatively inexpensive, evaluation of cellular immunity by ELISpot is more complicated. Furthermore, because of low B cell number which may prevent antibody production, serology testing is of limited use in our cohort of patients. Indeed, 57% of the patients after CART infusion in our cohort had a complete B cell aplasia prior to vaccination. In the allogeneic HCT group, B cell aplasia is less common and may result from either delayed immune reconstitution or concomitant administration of rituximab for the treatment of GVHD. In our cohort, by using both methods, we identified immune response to vaccine in 57% of patients after CART infusion and 75% of the patients after allogeneic HCT. This is an important finding that suggests that B cell aplasia should not by itself preclude patients from starting vaccination program. Of note, the observed response rate was significantly higher than that reported for other vaccines in the same population, and similar to the response of shown patients with chronic inflammatory diseases or immune dysfunction to bnt162b2 4, 18, 19 . The high response rate may partially reflect an inherent selection bias, as our inclusion criteria were based on the EBMT recommendations. While these results support the EBMT recommendations, we postulate that the vaccine response rate in an unselected transplanted population may be inferior. In line with the published data, we found that both longer period from infusion of cells (a composite outcome to encompass immune reconstitution) as well as absolute B cell counts were associated with better humoral response and, in addition, CD4 + /CD8 + ratio was associated with cellular response 18, 20 . Interestingly, intensive IST and active GVHD did not predict response to vaccine, thus patients may be referred to vaccination even if under intensive IST therapy. CD4/CD8 (median, range) 0.48 (0.14-2.9)

IST-immunosuppressive therapy # all patients with stable mixed chimerism and myeloproliferative neoplasms * Of 38 patients given IST 

COVID-19 in transplant recipients: The Spanish experience

Risk factors and outcome of COVID-19 in patients with hematological malignancies

Safety and Efficacy of the BNT162b2 mRNA Covid-19

A randomized trial of one versus two doses of influenza vaccine after allogeneic transplantation

Comparable humoral response after two doses of adjuvanted influenza A/H1N1pdm2009 vaccine or natural infection in allogeneic stem cell transplant recipients

EBMT-NIH-CIBMTR Task Force position statement on standardized terminology & guidance for graft-versus-host disease assessment

Vaccination of the Stem Cell Transplant Recipient and the Hematologic Malignancy Patient

Randomized study of early versus late immunization with pneumococcal conjugate vaccine after allogeneic stem cell transplantation

Safety and Immunogenicity of SARS-CoV-2 mRNA-1273 Vaccine in Older Adults

COVID-19 vaccine BNT162b1 elicits human antibody and T(H)1 T cell responses

Targeting the IL17 pathway for the prevention of graft-versus-host disease

Thrombocytopenia following Pfizer and Moderna SARS-CoV-2 vaccination

Immune thrombocytopenic purpura (ITP) associated with vaccinations: a review of reported cases

Does vaccination in solid-organ transplant recipients result in adverse immunologic sequelae? A systematic review and meta-analysis

Association of an impaired bone marrow microenvironment with secondary poor graft function after allogeneic hematopoietic stem cell transplantation

Interferon-γ promotes vascular remodeling in human microvascular endothelial cells by upregulating endothelin (ET)-1 and transforming growth factor (TGF) β2

Evaluation of immune responses to seasonal influenza vaccination in healthy volunteers and in patients after stem cell transplantation

Immunogenicity and safety of anti-SARS-CoV-2 mRNA vaccines in patients with chronic inflammatory conditions and immunosuppressive therapy in a monocentric cohort

Granulocyte-macrophage colony-stimulating factor as immunomodulating factor together with influenza vaccination in stem cell transplant patients

All authors declare no conflicts of interests to disclose.