This iatrogenic phenomenon, known little or not by many practitioners, responsible for significant dental and periodontal complications, both functional and aesthetic, is called “Wire Syndrome” (WS). It is therefore considered an undesirable event of bonded orthodontic retainers, which must be differentiated from an orthodontic relapse. The objective was to perform, for the first time, a systematic review of the literature in order to define the prevalence of WS and to study its associated clinical characteristics. (2) Methods: A systematic review of the literature was performed following the guidelines of Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and recommendations using an electronic search strategy on four databases complemented by a manual search. All the prospective and retrospective clinical studies, including case reports and series, written in English or French, clearly mentioning the description, detection, or management of WS were included. Three independent blinding review authors were involved in study selection, data extraction, and bias assessment using the Mixed Methods Appraisal Tool (MMAT). (3) Results: Of 1891 results, 20 articles published between 2007 and 2021 fulfilled the inclusion criteria, with a globally high risk of bias since 16 articles were case report/series. The analysis of each article allowed the highlighting of WS through 13 categories, as follows: prevalence, apparition delay, patient characteristics, arch and tooth involved, families of movements, dental and periodontal consequences, type of wire, risk factors, etiologies, treatment, and preventive approach. (4) Conclusion: This systematic review of the literature elaborated a synthesis on WS, allowing general practitioners, periodontists, and orthodontists to understand this adverse event, to facilitate the diagnostic approach, and to underline preventive measures against WS. This review was registered in the International Prospective Register of Systematic Reviews . The long-term follow-up of orthodontic bonded retainers remains a challenge for orthodontists, but also for periodontists, as well as general practitioners. Whereas fixed retainer placement is a common procedure after orthodontic treatment, complications, which can be severe, can happen. Indeed, in 2007, Katsaros et al.  were theThe synthesis of clinical experience evoked by the authors cited above allows us to define and characterize Wire Syndrome (WS) as follows: Fixed orthodontic retainers can provoke aberrant, unexpected, unwanted, or unexplained tooth movement on teeth still bonded by a fixed retainer placed after orthodontic treatment, which could induce progressively iatrogenic dental and periodontal complications, functional and/or aesthetic, ranging from minor teeth displacement to teeth expulsion from the bone with loss of vitality. In the presence of severe WS, the retainer may become detached or fractured. WS is not a classic orthodontic relapse, and the position of the teeth does not correspond to any previous situation.However, neither general practitioners nor dental specialists, such as orthodontists and periodontists, are aware of the Wire Syndrome phenomenon. Concerning general practitioners, a lack of knowledge has been detected. A survey in eastern France showed that only 18.6% of general dentists were aware of the risks of adverse tooth movement associated with unintentionally active fixed retainers . HoweverTherefore, the aim of this study was to perform the first systematic review of the literature on Wire Syndrome (WS) in order to define the prevalence, to study its associated clinical characteristics, and, specifically, to facilitate the diagnostic approach of practitioners and to underline preventive and curative measures against WS.A systematic review of the literature (SRL) was performed, following as closely as possible the guidelines of Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and recommendations (reference). The protocol was registered in the International Prospective Register of Systematic Review (PROSPERO) (CRD42021269297).–Participants (P): Patients with “Wire Syndrome” (WS), i.e., dental movements described as aberrant, unexpected, unexplained, unwanted, or excessive;–Interventions (I): Fixed orthodontic retainer bonded at the maxilla and/or mandible after orthodontic treatment;–Comparisons (C): Patients not affected by “Wire Syndrome” (only for studies including a control group);–Outcomes (O): Define the prevalence of “Wire Syndrome” and the characteristics associated with it.–Study designs (S): All prospective and retrospective clinical studies, including case reports or series, written in English or French, clearly reporting the description, detection, or management of “Wire Syndrome” or tooth movement described as aberrant, unexpected, unwanted, or unexplained in the presence of a bonded fixed retainer placed after orthodontic treatment were included, regardless of the length of the follow-up. In vitro studies, narrative reviews, author opinions, editorials, or commentaries were excluded.According to the question formulated using the “Population, Intervention, Comparison, Outcome and Study Designs (PICOs) model”: ❖Electronic searchA search strategy, tailored to each database, combining keywords, Medical Subject Headings (MeSH) terms, and Boolean operators was performed without date restriction.The electronic search was conducted on four different databases on 1 September 2021 .❖Manual search–From the bibliography of articles selected by the electronic search;–○American Journal of Orthodontics and Dentofacial Orthopedics;○European Journal of Orthodontics;○Journal of Orthodontics;○Journal of Clinical Orthodontics;○Orthodontic & Craniofacial Research;○The Angle Orthodontist;○Revue d’Orthopédie Dento-Faciale;○L’information Dentaire.From the search engine of a selection of orthodontic and dental journals:To complement the electronic searches, a manual search was conducted:https://www.zotero.org accessed on 1 September 2021). Two authors carried out the entire procedure independently. In the case of disagreement, a third author was interviewed, and a mutual discussion was conducted to reach a consensus.This research was conducted using the reference management software Zotero version 5.0.96.2 .Given the heterogeneity of the studies selected for this SRL, a tool that can evaluate the methodological quality of different types of studies within the same SRL was employed: The “Mixed Methods Appraisal Tool” (MMAT)  .❖Single-stranded, round wire bonded on the canines only; diameter: 0.036 inch.❖Unknows strands with diameter: 0.0175 inch; 0.0215 inch; 0.0195 inch;Three strands with a diameter of 0.0155, 0.0195, or 0.0195 inches (heat treatment);Five strands with a diameter of 0.0215 inched (gold-plated);Six strands with a diameter of 0.0175 inches.Round, twisted, stainless steel wire:❖Six strands with a diameter of 0.018 inches.Round, coaxial, stainless steel wire:Most cases of WS are seen in the presence of round, twisted, stainless steel wires ,18,21,23Risk Factors: Predisposing factors of WS were investigated in three studies [❖p < 0.0001) [p = 0.029) [p = 0.049) [Patient-related factors: Different parameters were found to be significant in WS patients, such as lower facial level increase ( 0.0001) , vestibu= 0.029) , and pre= 0.049) . However= 0.049)  did not ❖p = 0.03) [p < 0.01) [p = 0.270), inter-canine distance (p = 0.065), or change in incisor inclination (p = 0.151). Orthodontic treatment-related factors: Different parameters were found to be significant in WS patients, such as debonded at a young age ( = 0.03) , canine  = 0.03) , and abs < 0.01) . In cont < 0.01)  found no < 0.01)  showed n❖p ≤ 0.05) [p = 0.562) [p = 1.000) [Wire-related factors: No significant differences were found in patients with WS regarding debonded wire ( ≤ 0.05) , (p = 0.= 0.562)  and type= 1.000) . studies ,18,22:❖PEtiologies: Different etiological hypotheses were mentioned in the included studies, which can be grouped into three categories, as follows . After identifying 1891 articles, 20 articles were selected and analyzed, with a globally high risk of bias. Given the limited number of existing publications on WS and the relevant information found in the case reports and case series, these types of study designs were included. The description of WS is recent; the first publication on WS appeared in 2007 , followeRegarding the delay in apparition, WS appeared between 1 and 21 years after the placement of a retainer in the included studies. The majority of cases of WS appeared within the first five years after the placement of the retainer. This time interval should be considered with caution for several reasons. First of all, it is difficult to date the apparition of WS with precision, as retainer visits are intermittent. Moreover, it is difficult to detect early WS, and the patient often consults when complications are already severe . ConcernIn the presence of WS, the bonded retainer is intact in most cases but, in severe cases, the wire may become partially debonded or fractured due to important dental movement. In any situation, various and different clinical dental and periodontal signs can be found. The detection of one sign related to WS must immediately alert the practitioner to the possible presence of WS in order to stop the iatrogenic evolutive process and to start adapted therapy. Indeed, this syndrome is progressive and starts with minor dental and periodontal consequences until the loss of vitality and/or tooth expulsion. In addition, when treatment is not carried out ,21, clinWith regard to WS prevention, the most important preventive measure is the use of a bonded passive retainer. The use of a dental model to perfectly fit the retainer to the teeth before placement is recommended, as well as an indirect bonding protocol. Furthermore, special care should also be taken when the fixed retainer needs to be repaired. When a composite comes loose, small tooth movements may have already occurred and the wire is no longer a perfect fit. In this case, using an instrument to “push” the wire to better fit the teeth results in an active wire that could potentially be responsible for subsequent WS. Therefore, a new passive wire should be bonded rather than repaired. Another strategy for preventing WS is double retention, which combines a fixed bonded retainer with a removable thermoplastic retainer ,5,8,18. Although there are explanatory hypotheses that could justify the risk factors mentioned by the authors, not all studies agreed. Indeed, WS seems to be due to a combination of different and multifactorial etiologies. In addition, it appears that the delay in apparition varies with etiology . Early WTM; CA Digital GmbH, Mettmann, Germany) versus round, 0.0175 inch (in), six-stranded, twisted, stainless steel wire retainers  and showed no significant difference between these two types of retainer after one year of placement. To reach a consensus on the preferred type of wire to use, randomized controlled studies must be conducted on a large sample size and over long observation periods.The type of wire seems to have an influence on the occurrence of WS. In the study of Padmos et al. , the mecFinally, some additional points should be made regarding this systematic review of the literature. First, only one study designed as a randomized controlled trial was included for analysis, and the risk of bias was considered high, as the majority of included studies were designed as case series/reports. Second, there was a lack of information in the included studies. For example, few data were provided on patient characteristics prior to orthodontic treatment , orthodontic biomechanics employed to treat the patient , bonding and retainer placement protocol, history of retainer failures , etc. Furthermore, the periodontal conditions, such as type of phenotype , tractioThis first review of the literature on Wire Syndrome (WS) included 20 articles published between 2007 and 2021, with a majority of case report/series leading to a globally high risk of bias. However, the analysis of the overall article provided an understanding of this adverse event associated with fixed orthodontic retainers, emphasized the importance of an early diagnosis, and highlighted preventive measures against WS for dental professionals worldwide, including general practitioners (GP), periodontists, and orthodontists. Indeed, the WS problem must involve all the dental health professions, including the general practitioners who will be able to refer, if necessary, the patient to a specialist practitioner; the continuity of the collaboration and the “ortho–paro–gp” link will then be prolonged during the therapeutic time, thus guaranteeing optimal patient care. Further studies are needed to improve the knowledge about fixed orthodontic retainers, based on a large, well-documented sample and conducted over a very long observation period.


The most common malignancy diagnosed in childhood is acute lymphoblastic leukemia (ALL). With significant advances in ALL treatment and excellent supportive care, overall survival is now up to 90%, depending on the risk group. As a result, the survivor population is growing significantly. At the same time, it is observed that survivors experience many health problems that substantially reduce their quality of life. Emerging evidence suggests that the increased susceptibility to multiple disorders related to accelerated aging in CCS may be attributed to chronic inflammation. Therefore, this study aimed to investigate whether asymptomatic survivors of ALL have a biological phenotype of accelerated cellular senescence early in life. For this purpose, a broad panel of 51 cytokines was tested. We show that survivors after ALL treatment have an inflammatory profile associated with premature cellular aging. Factors that increase the risk of immunological alterations include younger age at diagnosis, high-risk protocols, and radiation therapy.p < 0.05). Increased levels of pro-inflammatory cytokines, including the IL-1 family , IL-6 (p < 0.001), IL-17 (p < 0.001), IL-18 (p < 0.05), TNFα (p < 0.01), IFNα2 (p < 0.05), and IFNγ (p < 0.01), were found elevated in the entire study group, compared with the controls. Subjects treated previously according to the high-risk protocol had higher IL-18 levels than low- and intermediate-risk groups (p < 0.05). Elevated levels of IL-1ra, IL-6, IL-12 (p70), IL-17, LIF, M-CSF, CSF, and VEGF were found in ALL survivors treated before the age of 5, compared with subjects treated over 5 years of age (p < 0.05). Moreover, individuals who received radiotherapy presented elevated levels of both IL-18 (p < 0.05) and MIG (p < 0.05). In conclusion, we found that young asymptomatic survivors after ALL treatment demonstrated a biological profile of complex low-grade chronic inflammation.Childhood acute lymphoblastic leukemia (ALL) survivors are at higher risk of developing many late effects later in life. They experience multiple health problems that have significant public health implications, such as frailty, premature onset of lifestyle diseases, and second tumors. There is some evidence that chronic inflammation causes accelerated aging in childhood cancer survivors; however, the available data are very limited. The aim of the study was to evaluate the broad panel of cytokines among asymptomatic ALL survivors after anticancer treatment. The study included 56 subjects with a mean age of 16.11 ± 3.98 years. The commercially available Bio-Plex Pro Human Cytokine Screening 48-Plex Panel Assay and Bio-Plex TGF-β Assay were used for simultaneous determination of 48 cytokines and 3 isoforms of TGF-β. Among 51 tested cytokines, the levels of 33 were statistically significantly higher in ALL survivors than in the control group ( Acute lymphoblastic leukemia (ALL) is the most common malignancy in children. Currently, the overall long-term survival rate in children treated for ALL is approximately 80% due to advanced treatment and excellent supportive care ,2. Over One hypothesis holds that the differences in health status between childhood cancer survivors (CCS) and their equals result from premature cellular aging. Furthermore, emerging evidence suggests that the increased susceptibility to multiple disorders related to accelerated aging in CCS may be attributed to chronic inflammation . DespiteAnticancer therapy may lead to treatment-induced accelerated aging in several possible ways. They include cellular senescence, oxidative stress, telomere attrition, DNA damage, stem cell exhaustion, mitochondrial dysfunction, epigenetic alterations, etc. In general, cellular senescence is defined as the loss of the cells’ ability to proliferate properly, resulting in an irreversible arrest of cell growth ,8. This In recent years, there has been an increase in the number of publications on the role of premature aging in patients with CCS; however, only few have examined the contribution of cytokines, and they have been limited to only few single proinflammatory cytokines  . MoreoveThe aim of this study was to investigate whether asymptomatic children and young adult survivors of acute lymphoblastic leukemia have a biological phenotype of accelerated cellular senescence early in life. For this purpose, a broad panel of 51 cytokines was tested.2). The waist-to-height ratio (WHR) was calculated by dividing waist circumference by height. Blood pressure was measured using a standardized sphygmomanometer. Hypertension (HT) was defined as a mean value of systolic blood pressure (SBP) and/or diastolic blood pressure (DBP) level ≥ 95th percentile adjusted for age, sex, and height. Echocardiography was performed to assess the heart’s structures and fraction shortening and ejection fraction by a pediatric cardiology specialist. The study was conducted in accordance with the Declaration of Helsinki and was approved by the local ethics committee (Permission Number: R-I-002/328/2019).Fifty-six White survivors of acute lymphoblastic leukemia, without clinical features of any comorbid disease, were recruited for the study. They gave written informed consent prior to the enrollment. The characteristics of the survivors’ cohort are presented in The commercially available Bio-Plex Pro Human Cytokine Screening 48-Plex Panel Assay  and Bio-Plex TGF-β Assay  allowed the simultaneous determination of 48 cytokines, as well as 3 isoforms of TGF-β  in each well of a 96-well plate, respectively. The Bio-Plex assays were performed on serum specimens according to the manufacturers’ instructions. The assay principle was based on the reaction with an antibody against a specific protein, which was covalently bound to fluorescently dyed magnetic beads, each with a distinct wavelength unique to the target protein. The bead-conjugated antibodies react with the sample containing the protein of interest. After the series of washes, biotinylated antibodies optional for different epitopes of the target proteins were added to the reaction. The final complex was made by adding a streptavidin–phycoerythrin (SA-PE) conjugate. A dual-laser, flow-based microplate reader—Bio-Plex 200 Reader —detected the internal fluorescence of the individually dyed beads and the intensity of the signal on their surface. The obtained signal was expressed as median fluorescence intensity (MFI), analyzed, and presented as concentration (pg/mL) using the Bio-Plex Manager Software . The concentration of the analyte attached to the individual beads was proportional to the MFI of the phycoerythrin signal.2 test were used. For independent variables, the Mann–Whitney U test or Student’s t-test were used. We compared risk groups  using univariate analysis of variance (ANOVA), and Tukey’s test was performed for post hoc analysis. Correlations were calculated by Spearman’s test. A statistical significance was determined at 0.05.The statistical analysis was performed using GraphPad Prism version 9.3.1 . All values are expressed as a mean ± standard deviation (SD) or median (Me) and interquartile range (IQR) when appropriate. In the univariate analysis, Fisher’s exact test and χp = 0.6) and median age of the study group did not differ from the control group—16.36 (13.32–19.27) years vs. 16.12 (14.16–17.65) years, p = 0.915.The characteristics of the study group are shown in p < 0.05). A comparison of the cytokine profile in the study and reference groups is presented in Among 51 tested cytokines, the levels of 33 were statistically significantly higher in ALL survivors than in the control group (p < 0.0001), IL-6 (p < 0.001), IL-17 (p < 0.001), IL-18 (p < 0.05), TNFα (p < 0.01), IFNα2 (p < 0.05), and IFNγ (p < 0.01)—were found elevated in the entire study group [Increased levels of proinflammatory cytokines—namely, the IL-1 family , as shown in To determine the effect of treatment intensity on cytokine levels, survivors were divided into three groups according to the treatment protocols used in childhood. As a result, subjects treated previously according to the high-risk protocol had only higher IL-18 levels, compared with low- and intermediate-risk groups (p < 0.05) , IL-17, LIF, M-CSF, CSF, and VEGF were found in ALL survivors treated before the age of 5, compared with subjects treated over 5 years of age ( < 0.05) .The potential impacts of some cytostatic agents  on serum cytokines and chemokines levels were further investigated; however, no effect was found.p = 0.037) and MIG (p = 0.012), as shown in One of the most harmful types of anticancer treatment, which is currently only targeted at carefully selected patients, is radiation therapy. In our cohort study, individuals who received radiotherapy had elevated levels of IL-18  such as Th1 , Th2 , and Th17 (IL-17A) in 87 asymptomatic ALL survivors. They showed a significantly higher concentration of IL-2, IL-10, and IL-17a, compared with controls . Our stuIn contrast, in a study by Sulicka-Grodzicka et al. of 50 young adults with CCS, higher levels of IL-6 were not confirmed. It is noteworthy that in addition to higher levels of C-reactive protein and fibrinogen, they found a shift toward memory and activated T cells, higher activation of cluster differentiation marker (CD) 38, and a decrease in naive T lymphocytes . MoreoveInterleukin 18 (IL-18) is a member of the interleukin 1 (IL-1) family expressed by a range of inflammatory cell types. In our study, a higher level of IL-18 was found in high-risk patients than in those who were treated according to intermediate- or low-risk protocols. In addition, its higher levels were found in the subset of patients who were treated with radiation therapy compared with those in whom it was not used. IL-18, together with IL-12, induces high levels of IFNγ production by T cells, and its expression correlates with disease activities of rheumatoid arthritis and Crohn’s disease. Furthermore, in animal models, IL-18-deficient mice were more susceptible to bacterial infections than normal ones and showed uncontrolled disease progression. The assembly of the inflammasome in cells results in caspase-1 activation, followed by proteolysis and release of the cytokines IL-1b and IL-18 to induce pyroptotic cell death . All theFurther stratification of patients in this study was based on age at diagnosis. The rationale for this approach was the hypothesis that children treated at a younger age are more likely to develop chronic inflammation due to the immaturity of many organs and systems. As expected, high levels of several cytokines and growth factors among ALL survivors treated before the age of 5 were found. They included IL-1 receptor antagonist (IL-1Ra), IL-6, IL-12(p70), IL-17, leukemia inhibitor factor (LIF), macrophage colony-stimulating factor (M-CSF), stem cell factor (SCF), and vascular endothelial factor (VEGF).Unlike most cytokines of the IL-1 family, IL-1Ra has antagonistic effects on IL-1 α and β, which have exquisite proinflammatory potency. It binds competitively to the same receptor as IL-1 but does not transmit a cellular signal, thus inhibiting IL-1-induced cellular changes . ContrarInterestingly, IL-12(p70) expression was also higher in a group of younger children at the time of diagnosis. Their bioactive form IL-12(p70) is produced mainly by macrophages, monocytes, neutrophils, and dendritic cells, and it primarily mediates Th1 cell differentiation and maintenance. Moreover, it indirectly exerts an anti-infective function by activating the cytolytic activity of natural killer (NK) cells and Th1 cells . On the Leukemia inhibitory factor (LIF) is a member of the IL-6 family and is expressed in almost all types of body tissues. LIF was first discovered as a cytokine that induces blast differentiation in myeloid leukemia; however, studies of LIF in other diseases, including cancer, indicate that it may potentially contribute to many other pathologies. The role of LIF appears to vary greatly depending on the tissue in which it is expressed. LIF is involved in tumor spread and metastasis, treatment resistance, and cachexia, and has also cancer biomarker potential . Since tMacrophages play a crucial role in inflammation and tissue regeneration, and their differentiation is mainly regulated by M-CSF derived from different tissues . ElevateIncreased levels of anti-inflammatory cytokines such as IL-4, IL-10, and IL-13 were also found in this study. Interleukin 4 is a major stimulant of Th2 cells and induces IgE production by B cells. Hence, it is involved in the development of allergy and is considered an important mediator of allergic inflammation . In turnMoreover, it is important to note that ALL survivors showed elevated levels of regulated on activation, normal T-cell expressed and secreted (RANTES), which is a member of the CC chemokine subfamily and plays a chemotactic role for monocytes, T cells, and eosinophils. It may have deleterious effects by recruiting immune cells that enhance inflammatory diseases such as arteriosclerosis, arthritis, nephritis, dermatitis, colitis, and many other disorders . There iAside from premature death, the most serious late effects of cancer treatment in children are second malignancies occurring later in life. The link between SASP factors and tumorigenesis has remained one of the most engaging scientific issues for years. There is ample evidence to support this linkage. It is emphasized that SASP promotes tumor cell proliferation, dissemination, angiogenesis, and invasiveness ,35,36. AINK4a and alternate reading frame protein product of the CDKN2A locus (ARF) [In addition to cellular senescence induced by SASP factors, many potential mechanisms lead to premature aging that should be noted. These include DNA damage, stem-cell depletion, mitochondrial dysfunction, oxidative stress, telomere attrition, and epigenetic changes. Much effort is currently being made in identifying a group of measurements of accelerated aging in cancer survivors. Researchers implemented the concept of biological age to estimate accelerated aging in children with cancer to better predict the aging process than chronological age . The risus (ARF) ,39,40.Finally, there are a few limitations to note. Firstly, single-center research should be interpreted in the context of the potential bias that may arise. Secondly, this pilot study was conducted on a relatively small group of individuals, which may affect the results. Finally, only cytokine levels as part of SASP were investigated, and no other parameters related to premature aging were analyzed. However, it is worth emphasizing that cellular senescence is a very complex process, and it is not possible to account for all factors and their interrelationships in a study performed on a small cohort of patients in one minor center.Strengths of the study include a homogenous group of acute lymphoblastic leukemia survivors, a relatively long follow-up time, and no ethnic diversity. To the best of our knowledge, this is the first study to address such a broad panel of inflammatory biomarkers among childhood acute lymphoblastic leukemia survivors.In conclusion, we demonstrated that ALL survivors present a biological profile of premature aging, manifested by increased secretion of multiple SASP biomarkers in the form of chronic inflammation. In addition, children treated for cancer at a younger age had higher levels of selected cytokines and growth factors than children treated at an older age, which may suggest a greater susceptibility of their healthy tissues to anticancer treatment administered in childhood. Furthermore, it is not yet clear how activation of cellular senescence mechanisms affects the development of different diseases. Additionally, in light of available studies indicating high morbidity among cancer survivors from multiple diseases, it seems reasonable to determine the exact pathogenesis of this condition. Thus, further longitudinal studies considering the contribution of a wide range of factors are needed.In conclusion, we demonstrated that ALL survivors present a biological profile of premature aging, manifested by increased secretion of multiple SASP biomarkers in the form of chronic inflammation. In addition, children treated for cancer at a younger age had higher levels of selected cytokines and growth factors than children treated at an older age, which may suggest a greater susceptibility of their healthy tissues to anticancer treatment administered in childhood. Furthermore, it is not yet clear how activation of cellular senescence mechanisms affects the development of different diseases. Additionally, in light of available studies indicating high morbidity among cancer survivors from multiple diseases, it seems reasonable to determine the exact pathogenesis of this condition. Thus, further longitudinal studies considering the contribution of a wide range of factors are needed.


Impaired response to COVID-19 vaccination is of particular concern in immunosuppressed patients. To determine the best vaccination strategy for this vulnerable group we performed a single center, 1:1 randomized blinded clinical trial. Patients who failed to seroconvert upon two mRNA vaccinations (BNT162b2 or mRNA-1273) are randomized to receive either a third dose of the same mRNA or the vector vaccine ChAdOx1 nCoV-19. Primary endpoint is the difference in SARS-CoV-2 spike antibody seroconversion rate between vector and mRNA vaccinated patients four weeks after the third dose. Secondary outcomes include cellular immune responses. Seroconversion rates at week four are significantly higher in the mRNA  as compared to the vector vaccine group . SARS-CoV-2-specific T-cell responses are reduced but could be increased after a third dose of either vector or mRNA vaccine. In a multivariable logistic regression analysis, patient age and vaccine type are associated with seroconversion. No serious adverse event is attributed to COVID-19 booster vaccination. Efficacy and safety data underline the importance of a booster vaccination and support the use of a homologous mRNA booster vaccination in immunosuppressed patients.Trial registration: EudraCT No.: 2021-002693-10. Optimizing COVID-19 vaccination strategies for patients under immunosuppressive medication is of high importance. In this clinical trial including non-seroconverted immunosuppressed patients, a homologous mRNA booster vaccination resulted in higher seroconversion rate than a switch to a vector-based vaccine. Various types of vaccines have been approved by the European Medicines Agency (EMA), including vector vaccines, such as ChAdOx1 nCoV-19 (Oxford-AstraZeneca) or Ad26.COV2-S (Johnson&Johnson) and mRNA vaccines, such as BNT162b2 (Pfizer–BioNTech) or mRNA-1273 (Moderna)7. Most recently, NVX-CoV2373 (Novavax) and VLA2001  have been approved by the EMA as a protein subunit vaccine and an inactivated whole-virus COVID-19 vaccine, respectively9. Multiple elements of the innate and adaptive immune system contribute to the vaccination response10. One way to assess the humoral immune response to different vaccines is to measure anti-SARS-CoV-2 antibodies against the receptor-binding domain (RBD). Immunocompromised individuals are less likely to mount an adequate immune response after primary vaccination. A significant number of patients do not seroconvert upon vaccination15, leaving them more susceptible to COVID-19 infections and subsequent severe disease courses16. Low antibody response rates have been observed in patients with immune-mediated inflammatory diseases, haemato-oncological malignancies, those following solid-organ transplantation, and patients undergoing hemodialysis21. Several studies have reported on the efficacy and safety of an additional booster vaccination in immunosuppressed patients. These mainly consist of the administration of a third mRNA vaccine in a homologous vaccination strategy26. Evolving evidence, however, suggests that a heterologous vaccination strategy might be more efficient in nonimmunocompromised healthy volunteers28. However, data on immunogenicity and safety of homologous versus heterologous booster vaccination strategy in patients who did not seroconvert are currently limited33. We, therefore, performed a blinded randomized controlled trial to address immunogenicity and safety of the third dose in non-seroconverted immunosuppressed patients, comparing mRNA and vector vaccines.The COVID-19 pandemic poses an unprecedented challenge to public health, and several mitigation strategies exist to combat this worldwide threat. Among such strategies, COVID-19 vaccination protects against a severe disease course and leads to accelerated viral clearanceSeventy-five patients under immunosuppressive therapy who had been immunized with two doses of an mRNA vaccine were screened for eligibility. Twenty-four patients were excluded due to the presence of detectable SARS-CoV-2-specific antibodies. Fifty-one non-seroconverted patients were randomized, of whom 25 were assigned to receive a vector and 26 to receive an mRNA vaccine as the third dose; five patients withdrew consent between the screening and the baseline visit Fig. . Thus, ap = 0.006)  as compared to the mRNA group  Fig. . Post ho8% as com+ peripheral B-cells and SARS-CoV-2 spike-specific memory B-cells were analyzed by flow cytometry. Patients without detectable CD19+ peripheral B-cells did not develop anti-RBD antibodies in either group  served as prepandemic control. Characteristics of HC and prepandemic patients can be found in Supplementary Tables p = 0.0026) or the mRNA vaccine (p = 0.0396). No difference in SFCs/106 PBMCs was observed between patients who received the third vaccination with either an mRNA- or a vector-based vaccine , who were vaccinated three times with BNT162b2 and developed a humoral immune response, served as a positive control. Material from patients under immunosuppressive therapy collected before the COVID-19 outbreak  occurred during the four-week follow-up. The prevalence of systemic reactogenicity was similar in the vector and mRNA booster vaccine groups. 7/22 (32%) of vector-vaccinated patients developed arthralgia compared to 4/24 (17%) of patients with mRNA booster vaccination. Myalgia was reported in 7/22 (32%) vector-vaccinated patients compared to 9/24 (38%) mRNA-vaccinated patients. Fatigue was present in 11/22 (50%) vectors and in 10/24 (42%) mRNA-vaccinated patients. Local pain at the injection site was more frequent in mRNA  than in vector-vaccinated patients . Headache was reported in 8/24 (33%) of the mRNA-vaccinated patients compared to 9/22 (41%) of the vector-vaccinated patients.Duration of side effects was prolonged in the mRNA as compared to the vector-vaccinated group, especially for local reaction (mRNA: 1.46 versus vector 0.46 days per patient), myalgia , and headache  Fig. .Fig. 5San = 36). None of the patients developed de novo anti-HLA antibodies.No thrombocytopenia or antibodies against platelet factor 4 (PF4) were observed after additional booster vaccination. None of the patients experienced an anaphylactoid reaction. In patients with previous organ transplantation, no acute transplant rejection was observed. Anti-HLA antibodies of patients with solid-organ transplantation were analyzed 12 weeks after the third vaccination with either a vector or an mRNA-based vaccine  than in the vector-vaccinated (heterologous) group. Overall, an additional COVID-19 vaccination resulted in humoral immune response in 41% of this initially vaccination-refractory patient population. Although the SARS-CoV-2-specific T-cell response was reduced in immunosuppressed patients as compared to HC, the cellular response could be increased after mRNA- and vector-based booster vaccination.36. While antibody levels have been determined at multiple time points during clinical routine before booster vaccination, it cannot be ruled out that individual patients had developed transient antibodies prior to study inclusion. Although case series and clinical trials have previously addressed the immunogenicity of a third vaccination in immunosuppressed patients, data comparing homologous versus heterologous vaccination strategies are currently limited33. A significant advantage was observed for the homologous booster dose in our study: the primary outcome showed a 44% higher seroconversion rate for mRNA (homologous) versus vector (heterologous) vaccination. In line with these data, anti-RBD antibody levels were significantly higher in patients who received an mRNA-based vaccination. To assess the stability of the humoral immune response, patients were invited to participate in an open-label extension study. Anti-RBD antibody levels were sustained over a period of 12 weeks, indicating a stable humoral immune response in immunosuppressed patients over at least 3 months.Patients without detectable anti-RBD antibody levels were included in our study, reflecting individuals at exceptionally high risk for severe COVID-19 disease courses. Although none of the patients seroconverted after primary vaccination, the heterogeneity of the patient cohort has to be considered since it has been reported that underlying disease-specific medication can contribute to vaccination response38. In the current trial, we excluded patients under rituximab treatment to ensure that absence of peripheral B-cells was not the main factor for an insufficient humoral immune response to COVID-19 vaccination. In addition, patients were stratified by the presence or absence of peripheral B-cells. Overall, the numbers of CD19+ peripheral B-cells did not correlate with anti-RBD antibody levels. However, when we analyzed SARS-CoV-2 spike-specific memory B-cells we observed a significant correlation with anti-RBD antibody levels, suggesting that the presence of anti-SARS-CoV-2-specific memory B-cells can be interpreted as predictive of antibody production40. ELISpot assays were performed using PBMCs to determine the in vivo effect of immunosuppressants on the development of circulating spike-specific T-cells. In line with previously published data on cellular responses in patients under immunosuppressive therapy42, we observed reduced T-cell responses in our patient cohort as compared to HC, suggesting a significant contribution of impaired T-cell function to the phenomenon of non-seroconversion after primary vaccination. A significant increase in SARS-CoV-2-specific T-cells was observed for both, mRNA- and vector-based booster vaccination, which was still diminished as compared to HC. In our study, T-cell responses to spike peptide pools from Wuhan and Omicron variants showed largely comparable results. This is in line with recent data reporting that the cellular response to SARS-CoV-2 variants is preserved in most infected and vaccinated individuals47. As T-cell responses to SARS-CoV-2 spike peptides could be influenced by pre-existing cross-reactive immunity against other human corona viruses49, we analyzed responses to the more variable S1 part containing the RBD and more conserved membrane-proximal S2 moiety of the spike protein separately. The results obtained revealed some variability between S1 and S2 responses in different groups, these, however, were not statistically significant. Although our data suggest that immunosuppressive therapy leads to impaired cellular immune responses, a bigger patient cohort would be needed to define the exact role of immunosuppressants. In addition, future in vitro assays should be performed to address the specific effects of different immunosuppressants on T-cell reactivation. Furthermore, we did not observe a significant correlation between SARS-CoV-2-specific T-cells and anti-RBD antibody levels, indicating more diverse roles of immunosuppressants in antibody production.In patients under B-cell-depleting therapy, such as rituximab, we and others have previously reported that seroconversion was impaired but a cellular immune response was preserved, supporting the importance of peripheral B-cells for seroconversion50. Common side effects  were observed, that were within the range reported in the approval studies6. While a longer duration of adverse events in patients boosted with a homologous mRNA vaccine than with the heterologous vector vaccine was observed in our patient cohort, this still lasted on average only 1.5 days.One of the biggest concerns of an additional booster vaccination in immunosuppressed patients relates to the risk of adverse reactions, which were documented by a patient diary within our study. No serious adverse event was recorded which could be related to booster vaccination. Reactogenicity was similar upon third vaccination as reported previously in heterologous vaccination regimens 51. To address this issue, we determined anti-HLA antibodies in solid-organ transplantation patients vaccinated either with vector or mRNA vaccine. In line with the most recent assessment report from the EMA and previous research53, no newly developed anti-HLA antibodies were detected 12 weeks post-vaccination.Concerns have been raised that cell culture-derived vector vaccines can induce HLA-sensitization in transplant candidates and recipients, which may negate future transplantations or activate low-level chronic anti-graft responsesOne limitation of the current study is the lack of placebo control, which was considered unethical. Despite early termination due to recruitment limitations, we were still able to demonstrate the statistical significance and benefit of homologous as compared to heterologous vaccination in immunosuppressed patients, which implies a robust difference between the two currently utilized strategies. In the current study, a heterogeneous population of immunosuppressed patients was included. Therefore, additional studies with larger patient cohorts will be needed to understand the effect of disease entity and immunosuppression on seroconversion and whether the enhanced humoral and cellular immune response in initially non-seroconverted patients also translates into clinical protection. Our data show that a humoral immune response can be mounted even in non-seroconverted patients who did not respond to primary vaccination and support a homologous vaccination strategy in immunosuppressed patients, also considering the acceptable safety profile.In this prospective blinded randomized controlled trial, adults (age ≥ 18 years) under immunosuppressive treatment without measurable SARS-CoV-2 spike protein-specific antibodies at least 4 weeks after their second COVID-19 vaccination were included. All patients had previously received two doses of an mRNA vaccine (BNT162b2 or mRNA-1273). Major exclusion criteria were known allergies to vaccines, previous infection with SARS-CoV-2, detectable anti-spike antibodies at the time of inclusion, or prior use of B-cell-depleting agents, such as rituximab. Healthy controls (HC) without immunosuppressive therapy as well as prepandemic immunosuppressed patients were recruited from the Vienna General Hospital and served as controls for the determination of SARS-CoV-2-specific T-cell responses. The detailed trial protocol can be found in the Supplementary Information. The trial was registered in the European Clinical Trials Database (EudraCT No.: 2021-002693-10) on the July 15, 2021.14, therefore patients were block-randomized in a 1:1 ratio based on the presence or absence of peripheral B-cells using a computerized algorithm (Randomizer). Patients were assigned to receive either a third dose of an mRNA vaccine (BNT162b2 or mRNA-1273 containing 100 μg mRNA), corresponding to their primary vaccination compound) or a vector-based COVID-19 vaccine (ChAdOx1 nCoV-19).The presence of detectable B-cells is critical for seroconversion29 and is outlined in more detail in the Supplementary Information. In brief, four trial visits were performed. In addition, an optional open-label fifth visit was offered to the patients 12 weeks after vaccination. At screening, the eligibility of the patients was verified. An additional booster dose was applied at visit two (baseline), the cellular immune response was assessed at week one, the humoral immune response at week 4, and optionally at the extension visit at week 12 after the third vaccination. Safety was evaluated by a patient diary for the first week, then inquired at week 4.The study was designed as described previouslyIn this trial, laboratory assessors and patients were blinded to the type of vaccine used. This was done to ensure an objective assessment of vaccine reactogenicity by the patients. Blinding of vaccines was ensured by the Central Pharmacy of the Vienna General Hospital, where dose aliquots were prearranged in syringes without reference to the vaccine type used. The study procedures followed Good Clinical Practice guidelines and the Declaration of Helsinki. The trial protocol was approved by competent authorities and the ethics committee of the Medical University of Vienna (No.: 1583/2021). At inclusion, all patients and healthy controls provided their written informed consent. All trial visits were conducted monocentric in a tertiary hospital . The trial started on July 22, 2021, with the inclusion of the first patient. The last patient finalized the 4-week follow-up on October 8, 2021.Laboratory tests, including quantification of peripheral leukocytes, anti-SARS-CoV-2 antibody testing, determination of SARS-CoV-2-specific T-and B-cell responses, and anti-HLA antibodies are described in more detail in the Supplementary Information.The primary outcome of the study was the difference in antibody seroconversion rates between the two intervention groups (vector versus mRNA vaccine). According to the manufacturer’s specification, seroconversion was defined as an anti-RBD antibody concentration of over 0.8 BAU/ml.Secondary endpoints included overall seroconversion rate and SARS-CoV-2 antibody levels at week 4 and cellular immune response before and 1 week following vaccination. Assessment of safety included incidence and severity of adverse events over 28 days. A paper-based patient diary was used to evaluate reactogenicity. The study sample size was targeted at 150 individuals, thus based on a Chi-squared test comparing the two groups, the minimal detectable difference was 21% at a power of 80%. During recruitment, routine access to the additional vaccination dose, as offered in our trial, was facilitated for high-risk patients in Austria, which substantially slowed our inclusion rates. To allow timely study completion, the decision was taken to stop inclusion prematurely at 75 enrolled patients, which would provide power to detect an effect size of at least 32% for our patient cohort. The observed effect size was ultimately larger, which eliminates considerations about a potential type II error, i.e., failure to reject the null hypothesis although it is actually false.τ). For graphical presentation, in figures with a log-scale, not-detectable anti-RBD antibodies and cellular responses have been set to 0.1. For all analyses conducted, two-sided tests were used. Data collection was facilitated by Microsoft Excel 365. GraphPad Prism (version 9.1.0) and “R” version 4.0.3  were used for graphical presentation and statistical analysis. Following packages were utilized: “ggplot2”, “ggbeeswarm”, “corrplot”, and “sjPlot” for creating plots, “pwr” for power calculation, and “tableone” to create baseline tables.The analysis included all patients vaccinated with an additional dose. Differences in seroconversion rates were statistically compared by utilizing a Chi-squared test. Post hoc analyses, including a comparison of antibody levels, type of mRNA vaccine, antibody stability, as well cellular immunity, were performed by Kruskal–Wallis- or Mann–Whitney U test in unpaired groups or paired Wilcoxon test in paired groups. Factors associated with seroconversion rates were assessed by multivariable logistic regression analysis. Variables that might influence seroconversion were included in the model . Univariate associations between continuous variables were described via Kendall rank correlation coefficient (Further information on research design is available in the Supplementary InformationReporting Summary


COVID-19 is associated with an increased mortality in hemodialysis patients. Therefore, achieving a long-lasting effective immune response to SARS-CoV-2 vaccines is essential. This study describes the humoral immune response in hemodialysis patients following three doses of mRNA vaccines against SARS-CoV-2, and explores the factors associated with a sustained immune response.We analyzed the monthly serological evolution of SARS-CoV-2 anti-S(RBD) antibodies for 1 year in 178 chronic hemodialysis patients who received three doses of SARS-CoV-2 mRNA vaccines. The primary outcome was sustained effective humoral response defined as anti-S(RBD) levels > 1,000 AU/ml after 4 months from the third dose. Multivariate logistic regression analyses were used to identify features associated with a sustained humoral immune response.After the initial two SARS-CoV-2 mRNA vaccine doses, 77.8% of patients showed an immediate effective humoral response, decreasing to 52.5% after 4 months. Antibody levels were significantly higher in COVID-exposed patients and HBV vaccine responders. After the third dose, 97% of patients showed an effective humoral response, and remained in 91.7% after 4 months. The mean monthly rate of antibody titer decline decreased from 33 ± 14.5 to 25 ± 16.7%. Multivariate regression analysis showed that previous exposure to COVID-19 and response to HBV vaccines were associated with an effective sustained humoral immune response.Immunization with SARS-CoV-2 mRNA vaccines elicits an effective immediate humoral immune response in hemodialysis patients, with a progressive waning in antibody levels. A third booster dose enhances the immune response with significantly higher antibody levels and more sustained humoral immune response. COVID-naïve patients and patients without previous response to HBV vaccines are likely to benefit from receiving more booster doses to maintain an effective immune response. Patients on hemodialysis (HD) have an increased incidence of infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) compared to the general population, and when infected their mortality risk is higher –3. For tThere are no specific recommendations concerning the most adequate vaccine regimen for HD patients. Around 90% of HD patients without previous infection with SARS-CoV-2 develop an immediate humoral and cellular immune response after the administration of two doses of mRNA vaccines , 11. DesThere are no longitudinal studies concerning SARS-CoV-2 vaccine-induced immune responses for more than 4 weeks after receiving the third mRNA vaccine dose in patients on HD, which are needed to optimize clinical care and plan preventive strategies in this population.In this study, we describe the serological monthly changes against SARS-CoV-2 in HD patients after 1 year of follow-up from an initial immunization with two doses of mRNA vaccines. A third booster dose was administered after 5 months from the second dose, with continuous monthly serological assessment for 4 months following the third vaccine dose. We aimed to describe the factors associated with a sustained humoral response and identify the group of patients who are more likely to benefit from receiving a third booster dose.We designed an observational, longitudinal study to evaluate the immune response induced by the different mRNA vaccines against SARS-CoV-2 in patients attending two HD units: an in-center hospital dialysis unit and its affiliated satellite dialysis unit. The study comprised all HD patients who received vaccination against SARS-CoV-2 with a two-dose mRNA vaccine between December 28th 2020 and June 30th 2021, either BNT162b2 (BionTech/Pfizer) or mRNA-1273 (Moderna), given according to the manufacturer’s recommendations. The type of vaccine was assigned according to the available vaccines at the local vaccination point or at our center.Patients were classified as having had a prior COVID-19 infection if there was clear medical documentation with a positive SARS-CoV-2 PCR swab or in presence of nucleocapsid-IgG specific antibodies in serological analyses before the administration of the first dose of the vaccine. Patients who had received the vaccination schedule before starting HD, patients with an unknown serological status for SARS-CoV-2 before vaccination and patients who either received an adenoviral vector-based vaccine or did not complete the full SARS-CoV-2 mRNA vaccine schedule were excluded from the study.The study was conducted according to the guidelines of the Declaration of Helsinki and received the approval of the Ethical Committee for Clinical Investigation of the Hospital Universitario Fundación Alcorcón. All patients signed an informed consent form and approved the use of their anonymized clinical information for medical research purposes.Patients received two initial doses of either mRNA vaccine (Pfizer or Moderna) separated by 4 weeks, followed by a third booster dose after 5 months. Serum samples for SARS-CoV-2 anti-spike 1 receptor binding domain [anti-S(RBD)] IgG antibody titers were obtained 2 months prior to the administration of the first dose of the vaccine as well as immediately before vaccination, immediately after the administration of the first and the second dose of vaccine, and thence in a monthly basis up to 4 months after the third booster dose, which would correspond to 10 months after the administration of the first dose . This seWe examined associations between anti-S(RBD) IgG antibodies and potential predictors such as demographic and clinical data, type of dialysis and type of the vaccine, previous exposure to COVID-19, history of effective humoral response to hepatitis B virus (HBV) vaccines, and treatment with immunosuppressants. Medical histories of study participants were extracted from their medical records.Antibody response was determined using the chemiluminescent microparticle immunoassay of IgG antibodies to SARS-CoV-2 in human serum  which provides quantitative measures of IgG antibodies specific for SARS-CoV-2 S1-RBD. Results are given as arbitrary units per milliliter (AU/ml), and were interpreted as positive according to the manufacturer’s instructions, with a cutoff index value of > 50 AU/ml. The upper limit of quantification was 40,000 AU/ml. The humoral response was considered as effective when the value of SARS-CoV-2 anti-S(RBD) antibodies was > 1,000 AU/ml, which strongly correlates with a sustained neutralizing antibody response of 1/80 measured by plaque reduction neutralization titer , 19. TheDialysis vintage was defined as the time between the first HD session and the administration of the first dose of mRNA vaccine. Response to HBV vaccine was defined as an anti-HBs antibody titer > 10 IU/ml after four doses of Engerix-B or Fendrix. Immediate humoral response to SARS-CoV-2 vaccines was defined as an anti-S(RBD) antibody titer > 50 AU/ml 4 weeks after the second dose of the vaccine, and the same cutoff was used in every monthly serological assay. Effective humoral response was defined as an anti-S(RBD) antibody titer > 1,000 AU/ml, and sustained humoral response was defined as a persistent antibody titer > 1,000 AU/ml for more than 4 months.t-test, if normally distributed, or by the Mann-Whitney test if abnormally distributed. A Chi-square test was used for categorical variables. Multivariable logistic stepwise regression was used to determine the independent factors associated with an effective humoral response  4 months after the first two mRNA doses and 4 months after the third dose. The covariates of interest were selected based on the associations with serological response in univariate analyses, and any variables that were at the significance level p less than 0.10 in univariate analyses were included in these models. P-value of < 0.05 was considered statistically significant. Statistical analysis was performed with IBM SPSS Statistics for Windows, Version 22.0 .Demographic and clinical information are described for all patients included in the study. Data were presented as a number (percentage) for categorical variables, as mean ± standard deviation for continuous variables that were normally distributed, and as median  for continuous variables that were non-parametric. Continuous variables were first tested for normal distribution using Shapiro-Wilk, and then compared by One hundred and ninety-six patients on maintenance HD were screened for this study, 18 patients did not meet the inclusion criteria and were therefore excluded. Ultimately, 178 patients were eligible and were included in the study. p < 0.001). After receiving the third mRNA vaccine dose, the proportion of patients who seroconverted increased to 98.3%, and this rate remained stable for the following 4 months. Median antibody levels rose up to 26,037 AU/ml  and decreased to 9,180 AU/ml  after 4 months from the booster dose (p < 0.001).The evolution of IgG anti-S(RBD) antibody levels in the whole cohort throughout the study period is represented in p < 0.001].The proportion of patients with IgG anti-S(RBD) antibody levels over 1,000 AU/ml changed in a similar manner, from 77.8% of patients after receiving the first two doses, to 52.5% of patients 4 months after the second dose, rising up to 97% after receiving the third booster dose, and remaining in 91.7% of patients after 4 months from the third dose . Overalln = 33) and those who were COVID-19 naïve at the time of vaccination (n = 145) (p = 0.04), and lower proportion of patients with online haemodiafiltration  in the group of patients with prior COVID-19 infection. Pre-vaccination levels of anti-S(RBD) IgG antibodies were elevated in 78.8% of COVID-exposed patients, with a median antibody titer of 1,123 AU/ml, of which 51.5% had antibody levels higher than 1,000 AU/ml before vaccination. On the other hand, in the COVID-naïve group only two patients (1.4%) had positive anti-S(RBD) antibody levels that were slightly elevated (56 and 63 AU/ml) with negative IgG nucleocapsid antibodies and no history of COVID-19 infection, and were thus considered COVID-naïve.We stratified the study population according to previous SARS-CoV-2 exposure into patients with prior COVID-19 infection before vaccination (n = 145) . There wp < 0.001), and remained so at every monthly assay. Serological changes in patients with previous COVID-19 exposure and in COVID-naïve patients is represented in p < 0.001).The proportion of patients who achieved seroconversion was similar in both groups . Howeverp = 0.002), and these differences remained for the following 4 months after the initial two-dose vaccine. Nonetheless, after the third SARS-CoV-2 vaccine dose, the proportion of patients who had positive anti-S(RBD) antibody levels was matched in both groups  had received a full HBV vaccination regimen, of which 84 patients (68.9%) had an effective humoral response. h groups , but theh groups .p < 0.001)  antibodies of 33 ± 14.5%. This decrease was significantly steeper in patients who were COVID-naïve (36.5 ± 11% per month) compared to patients with previous exposure to COVID-19 (17.5 ± 6%) (< 0.001) .p < 0.001). Although the rate of antibody level decline remained significantly lower in patients with previous exposure to COVID-19 , COVID-naïve patients also showed a more sustained humoral response after the third dose compared to the initial vaccination, with a monthly decrease in the rate of antibody level decline from 36.5 ± 11 to 27.8 ± 16% (p < 0.001). Also, patients who were responders to HBV vaccines showed a more sustained response after the third dose, with a decrease in the rate of antibody decline from 33.5 ± 12.3 per month to 24.7 ± 16% per month (p < 0.001) (The mean monthly rate of antibody titer decline decreased to 25 ± 16.7% after the third dose (< 0.001) .We analyzed factors that were associated with an effective humoral response after the initial two-dose mRNA vaccination. Patients with previous exposure to COVID-19 , previous humoral response to HBV vaccines , patients who were < 70 years old  and patients who received mRNA-1273  were associated with an effective sustained humoral response after 4 months from vaccination in the univariate analysis. Patients with history of neoplasms were less likely to maintain an effective humoral response . In the multivariable linear regression model, both previous exposure to COVID-19 infection and previous humoral response to HBV vaccines maintained statistical significance. Details are presented in In the univariate analysis to determine the factors associated with a sustained humoral response after 4 months from receiving the third dose of mRNA vaccine, we found an association with response to HBV vaccines , and a tendency for a less likely effective seroconversion in patients receiving immunosuppressants . After adjusting for age, dialysis vintage, immunosuppression, and type of vaccine administered, the humoral response to HBV vaccines remained as an independent factor associated with an effective sustained humoral response after 1 year from the initial vaccination. All patients with prior exposure to COVID-19 developed an effective humoral response, and therefore could not be included as a factor in the multivariate analysis .Throughout the year following SARS-CoV-2 vaccination, there were 23 vaccinated patients (12.9%) who were diagnosed with SARS-CoV-2 infection, of which 3 cases (1.7%) were after receiving the first two doses, and the remaining cases (11.2%) were diagnosed after receiving the third dose of vaccination. Twenty-one patients (91.3% of the infected patients) remained asymptomatic. There were only two cases (1.1%) of severe COVID-19 with acute respiratory failure requiring hospital admission and high flow oxygen therapy among vaccinated patients included in this study, both of which had an inadequate humoral response to the vaccine. The first patient was an 85-year-old male patient who became infected 3 months after receiving the first two mRNA vaccine doses, having reached a peak SARS-CoV-2 anti-S(RBD) antibody titer of 1,007 AU/ml in the first month after vaccination, and decreasing to 510 AU/ml in the second month, just before infection. The second patient was a 72-year-old female patient under immunosuppression with tocilizumab due to severe rheumatoid arthritis. She never responded to the initial two doses of mRNA vaccine nor to the booster dose (peak SARS-CoV-2 anti-S-RBD antibodies 1 AU/ml) and was infected 3 months after receiving the third dose. Neither patient had a history of COVID-19 infection before vaccination, and both patients died.In this serological study we examined the monthly changes in SARS-CoV-2 anti-S(RBD) IgG levels for 1 year after vaccination. We observed that patients who were COVID-naïve and those who had not previously responded to HBV vaccines benefited the most from a third mRNA vaccine dose against SARS-CoV-2, since it provided them with an effective humoral response , both immediately after vaccination and also sustained in time for at least 4 months after receiving the booster dose, whereas the first two doses had been unable to elicit such a sustained response.Several studies have determined the factors that are associated with an immediate humoral response few weeks after vaccination, including serum albumin levels, lymphocytes, dialysis vintage, intravenous iron dose, immunosuppressant treatment, previous infection with COVID-19, and a positive response to HBV vaccines –22. In op = 0.009). In this way, 4 months after receiving the booster dose, there were no significant differences in the protective humoral response between COVID-naïve patients and those who were previously infected with COVID-19 .Naïve patients for COVID-19 infection are the ones who benefit the most from vaccination and especially from the administration of a third dose . In our During the study period, the accumulated incidence in our area was around 20,500 cases per 100,000 population , predomip < 0.001). Therefore, this group of patients not only benefits from the first two vaccine doses but also from the booster effect of a third dose, although to a lesser extent than in the COVID-naïve population.HD patients who have had a COVID-19 infection have less protection against reinfection than the general population . HoweverPrevious studies have compared the humoral response after the second and third doses. One study analyzed the humoral response after the third dose of the vaccine, showing a slight increase in the proportion of patients who achieve seroconversion, but with a very intense humoral response, reaching a 36-fold increase in median antibody titers after the booster dose , and anoIt has been previously described that there is an association between humoral responses after HBV and SARS-CoV-2 vaccines in HD , 32, 33.It should be noted that the most used vaccine in our patients was mRNA-1273, 77.5% of the patients for the first two doses and 91.7% for the third dose (with no significant differences between the different groups of patients). It has been described that the type of vaccine acts as an independent factor for response to vaccination and that mRNA-1273 is the one that obtains the best results, perhaps due to its higher dose of mRNA .We believe that in the future, the rate of antibodies will continue to decline and a significant percentage of patients will probably lose protection in the mid-term. Therefore, monitoring the levels of antibodies in the HD population should be a priority in order to propose new immunizations when necessary, as other authors have suggested .Consequently, we could revaccinate when SARS-CoV-2 anti-S(RBS) IgG levels drop below a certain threshold, in a similar way to how revaccination of HBV is recommended when anti-HBs antibodies drop below 10 mIU/mL. We suggest that monitoring of anti-S(RBD) IgG levels should be particularly frequent in patients who did not respond to HBV vaccines, in COVID-naïve patients and in those with lower antibody levels.Our study has a number of limitations. First, the rate of neutralizing antibodies was not determined, which according to other authors can decrease progressively after the first two doses and can rise very significantly after the third dose. However, other studies have found a significant correlation between anti-S(RBD) antibodies and the neutralizing antibody response, and in our study we found that the changes in anti-S(RBD) IgG antibody levels resemble those described in neutralizing antibodies , 19. AnoIn conclusion, as a consequence of receiving a third dose of SARS-CoV-2 mRNA vaccine, HD patients achieve a better and more sustained humoral immune response over time, which probably translates into lower morbidity and mortality rates in SARS CoV-2 infection. Patients who are most likely to benefit from the booster dose are COVID-naïve patients and patients who were non-responders to HBV vaccines.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by the Medical Research Ethical Committee (CEIm), Hospital Universitario Fundación Alcorcón. The patients/participants provided their written informed consent to participate in this study.EG-V and AS: conceptualization, writing—original draft preparation, and writing—review and editing. EG-V: methodology and project administration. EG-V, AS, and ML-P: validation. AS: formal analysis. JA-S: investigation. EG-V and EG: resources. EG-V and ML-P: data curation. JA-S, AC, CC-C, and GF-J: visualization. All authors have read and agreed to the published version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Lung transplant recipients (LuTX) represent a vulnerable population for severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2). Even though many vaccines are already developed, more clinical data need to support effective immunological response in immunocompromised patients.Stable LuTX recipients with no medical history of coronavirus disease (COVID‐19) were enrolled. Currently available messenger RNA (mRNA)  and non‐mRNA  vaccines were given according to availability, boosters were all mRNA‐based. SARS‐CoV‐2 Spike1 immunoglobulin G (IgG) antibody titer was evaluated before and 2 weeks after second and third dose. Difference between mRNA versus non‐mRNA vaccines was assessed.p = .002). 6/23 (26%) patients received two doses of mRNA vaccine developed COVID‐19 after the second injection in an average of 178 days, half of them recovered, half of them died in intensive care unit (ICU). 3/6 (50%) patients with two doses mRNA and recovered from COVID‐19 had significantly higher level of antibody (average 20847.3 U/ml) than without infection. After the booster vaccine, 1/24 (4%) developed infection.Forty‐one patients  received two doses of SARS‐CoV‐2 vaccines: 23 of mRNA, 18 of non‐mRNA, and 24/41 (58%) received a third dose. Median 92 months passed since transplantation, and serum level of tacrolimus was median 5.5 ng/ml. Positive serology was found in 37% of all patients after the second dose, 86% had mRNA vaccine. After the third dose, 29% became positive who had no antibody before. Significantly higher level of antibody was found after the second mRNA than non‐mRNA vaccines (2.2 vs. 1568.8 U/ml, respectively, Immunosuppression therapy may induce a weaker SARS‐CoV‐2 response in LuTX recipients; therefore, third dose is a priority in transplanted patients. The highest antibody level was measured recovering from COVID after two doses. Our data confirm that booster mRNA vaccine could increase antibody levels, even if immunization was started with non‐mRNA vaccine. Serological findings following SARS CoV 2 vaccines in lung transplant recipients As LuTX patients receive high doses of immunosuppressant medications, they are at very highest risk of developing severe coronavirus disease 2019 (COVID‐19) and increased mortality.Transplant recipients were excluded from recent SARS‐CoV‐2 vaccine trials, so there are insufficient data about efficacy, durability, or safety in this patient subpopulation. Our study aims to assess specific immunological response to different types of vaccinations and determine the proportion of seroconverted patients and outcomes in Hungarian LuTX patients.22.1Before 2015 our patients underwent lung transplantation in Vienna. The first lung transplantation procedure was performed in Budapest, Hungary on December 12,  2015. The Department of Pulmonology, Semmelweis University is the only center for LuTx posttransplant care in Hungary. Our transplant activity is 20 cases per year. Our center uses the immunosuppression protocol of Medical University of Vienna. The induction therapy was antithymocyte globulin (ATG) of patients who underwent lung transplantation before 2008. From 2009 monoclonal antibody targeting CD52  30 mg intravenously was the induction therapy followed by lower maintenance immunosuppression.2.2LuTX patients in the study period between January and December 2021 included 148 individuals. Patients were recruited at least 1 year after the surgery, if they were in stable functional condition, and if no rejection nor infection was present. Those who had immunoglobulin deficiency were excluded. In Hungary, according to actual regulations individuals who were infected with SARS‐CoV‐2 within the last 3–6 months were exempt from vaccinations, including 30 LuTX patients.Out of the potentially eligible 118 patients, 41 volunteered for baseline SARS‐CoV‐2 IgG and IgM measurements at study entry to help determine baseline SARS‐CoV‐2 serostatus for the analysis of vaccine efficacy and were approved by local ethical committee (TUKEB‐IV/861‐1/2021/EKU).At the time of the study BNT162b2, mRNA‐1273, ChAdOx1, BBIBP‐CorV, and Gam‐COVID‐Vac vaccines were approved in Hungary. As there was no recommendation which vaccine should LuTX patients receive, all were vaccinated by the offered and available vaccines at centrally assigned vaccination points. SARS‐CoV‐2 Spike1 IgG antibody (Roche®) titer was measured 2 weeks after the second dose of the respective vaccine and if available, after the third dose at their regular control (2–6 weeks postvaccination). Positive serology was considered >0.8 U/ml serum level of anti‐SARS‐CoV‐2‐Spike1.2.3t test after testing for normality using Kolmogorov–Smirnov test, while χ2 test was applied for analyzing categorical data. A p value <.05 was defined as statistically significant.Statistical analysis was performed using Graph Pad software . Data are expressed as mean ± standard deviation (SD) or median. Differences between groups for parametric data were evaluated with Student's 3N = 20). The most common non‐mRNA vaccine was ChAdOx1 (N = 16). Twenty‐four of 41 (58%) patients got third dose after the second one on average 177 ± 42 days later, all third doses were mRNA‐based. No serious adverse events other than general lethargy and local skin reaction were recorded neither after the second nor the third vaccination.Forty‐one LuTX recipients were enrolled into the analysis. Baseline characteristics are summarized in Table n = 18) recipients received ATG as induction therapy and 23 alemtuzumab. Each patient received tacrolimus and prednisolon. The median dosage of prednisolone was 5 mg/day. The median tacrolimus serum level was 5.5 ng/ml (IQR: 2.5–11.2 ng/ml) before the first vaccination.Eighteen (Eleven out of 41 patients (26%) were treated with tacrolimus+everolimus+prednisolone. The total serum level of tacrolimus and everolimus was median 6.6 ng/ml (IQR: 3.6–11.2 ng/ml). Each patient except seven cases received mycophenolate‐mofetil, median 1500 g/day (IQR: 500–2000 g/day). Under the vaccination period, the immunosuppressive treatment did not change.N = 26; 64%), 3/41 patients (7%) had low‐positive antibody level (<10 U/ml) and six patients (15%) developed >1000 U/ml antibody titer 2 weeks after second vaccine. Thirteen out of 23 (57%) mRNA‐vaccinated patients became seropositive after two shots. Eighteen out of 41 patients received two doses of non‐mRNA vaccine; positive serology was found only in two cases (11%). A significant difference was found between the response of mRNA versus non‐mRNA vaccines , and the highest immune responses  were found in patients vaccinated with two doses of BNT162b2.Immune response for SARS‐CoV‐2‐Spike1 was not measurable in most cases after the second dose  still did not develop any immune response neither after the second nor the third vaccination. However, seven patients (29%) had positive antibody after the third dose who had none before and in these patients the average antibody titer was 2435 U/ml. Five of them received ChAdOx1, two of them BNT162b2 vaccines.p = .05). Only one of them was asymptomatic and recovered at home, while the other five required hospitalization. Two patients had moderate disease with 10%–20% involvement of the lungs, after a short time of hospitalization they recovered with no functional loss and high antibody titer  was measured thereafter. Three out of six patients had severe illness and needed intensive care, where they died soon, and after their second vaccination 183, 186, and 216 days have passed, respectively. Figure Six patients developed SARS‐CoV‐2 infection after the second vaccination in an average of 178 days, all of them received BNT162b2. Three patients had no detectable antibody, while the other three had 140, 160, and 1346 U/ml antibody levels respectively after two doses of vaccination. Significantly higher antibody levels were detected after recovering from infection  than after two doses of vaccines  while 57% of the patients became seropositive after two doses of mRNA vaccines. Among all patients vaccinated three times, positive antibody response increased from 17% to 46%, with a corresponding increase in neutralizing capacity after the third dose. The highest individual antibody level (>25000 U/ml) was measured after two doses of mRNA vaccines, and then recovering from SARS‐CoV‐2 infection. Infection could be life‐threatening in transplant recipients. In our patient population mortality was high (50%) among infected recipients, even though two or three doses of vaccines. No serious side effects of vaccination were observed. In the most recent recommendation, third dose is theoretically suggested, but that was not yet available in many countries.N = 48); however, 85% of the patients had high antibody response after COVID‐19.Several studies exists who studied the effectiveness of the booster vaccines in heart‐ transplanted patients,The low antibody response rate is alarming but not unexpected in LuTX recipients. Previous experiences with Influenza vaccine show a lower rate of immune response in solid organ recipients, however, vaccination demonstrated reduced influenza‐related lower respiratory tract disease and hospitalization despite low antibody response.Other concerns regarding the SARS‐CoV‐2 vaccines include the lack of long‐term safety data, potential reduction in efficacy in immunocompromised patients, unknown durability of the immune response, and potential for vaccine‐associated allograft rejection. Despite the low proportion of seroconversion observed in our patients, SARS‐CoV‐2 vaccines provide potential benefit and decreases the risk stabling transplant recipients.We know our study limitations with the low number of the cases, and vaccine efficacy cannot be defined obviously by the presence or absence of antibodies, which may be deceptive. We should also note that in lines with positive antibody response there is still no universal cutoff value. However, we observed low antibody response in patients vaccinated with non‐mRNA vaccines, none of them get COVID‐19. Additionally to neutralizing antibodies T helper1 CD4 + and CD8 + T cell response might contribute to immunity against SARS‐CoV‐2. Further data are needed to evaluate B‐ and T‐cellular responses after SARS‐CoV‐2 vaccination and disease.The emerging studies on SARS‐CoV‐2 vaccines indicate that they are safe, and our experience with different SARS‐CoV‐2 vaccines proved to be safe in lung transplant recipients. Currently, the third booster dose of SARS‐CoV‐2 vaccine is the most appropriate way to increase the immune response among lung transplant recipients. The durability of this immune response should be evaluated in the need of additional boosters in the future. However, we still have insufficient amounts of data using non‐mRNA‐based vaccines among transplant recipients, there is promising result about switching to a different type of vaccine  in nonresponders thereby offering a new strategy to increase immune response. Vaccination is the most promising way to tackle the pandemic, and third or more doses of vaccine are becoming a chance to avoid serious COVID‐19 complications.All authors have read and approved the final manuscript. All authors performed the research. Enikő Bárczi, Anikó Bohács, and Veronika Müller designed the research study, analyzed the data, and wrote the paper.The authors declare no conflicts of interest.


Overall, comprehensive analysis of a small cohort showed that co-administration of SARS-CoV-2 vaccine and IO is not associated with increased irAEs. Nevertheless, the detection of autoantibodies post anti-SARS-CoV-2 vaccination warrants further investigation (NCT03702309).Despite more than 2 years having elapsed since the onset of SARS-CoV-2 pandemic, a level of hesitation around increased SARS-CoV-2 vaccine toxicity in cancer patients receiving immunotherapy (IO) remains. This hesitation stems from the idea that IO agents could elicit an overwhelming immune stimulation post vaccination and therefore increase the risk of vaccine-related toxicity. The aim of our study was to explore serological responses to SARS-CoV-2 vaccination in patients treated with IO and describe the level of immune stimulation using parameters such as blood cytokines, autoantibody levels and immune related adverse events (irAEs) post vaccination. Fifty-one evaluable patients were enrolled in this longitudinal study. Absolute levels and neutralization potential of anti-SARS-CoV-2 antibodies were not significantly different in the IO group compared to non-IO. Chemotherapy adversely affected seroconversion when compared to IO and/or targeted treatment. Following vaccination, the prevalence of grade ≥2 irAEs in patients treated with IO was not higher than the usual reported IO toxicity. We report, for the first time, that anti-SARS-CoV-2 vaccination, elicited the generation of five autoantibodies. The significantly increased autoantibodies were IgM autoantibodies against beta-2 glycoprotein ( For more than 2 years, the world has faced tremendous public health, economic, and ethical challenges due to the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic . Cancer The development of vaccines preventing severe disease, hospitalization and death from COVID-19 has marked a significant achievement , 6. DespCancer patients have been prioritized for receipt of initial vaccination series and boosters around the globe. Despite this, there is ongoing concern as to whether vaccine efficacy is impacted by increased immune vulnerability in the setting of malignancy or secondary to immune-modulating therapies (such as immuno-oncology (IO) agents). The heterogeneous nature of neoplastic diseases and diverse therapies applied has further complicated efforts to interrogate this relationship.Emerging evidence suggests that the use of certain anti-cancer therapies during SARS-CoV-2 vaccination may alter immune correlates of protection. In some cancer patients, including a high proportion of those receiving cytotoxic or B-cell depleting therapies, lower rates of seropositivity and T-cell responses have been reported after two doses of mRNA vaccine –15. VaccIn the present study, we comprehensively explored peripheral immune responses to SARS-CoV-2 vaccination in IO-treated cancer patients. We also described the immune-mediated toxicities in IO-treated patients who received anti-SARS-CoV-2 vaccination and lastly, we conducted an in-depth analysis of auto-antibody generation following vaccination.Immune Response to COVID-19 Vaccine in immunotherapy (IO) and non-IO Treated Cancer Patients  between March 2021 and July 2022. Patients were allowed to have any of the approved vaccines in Canada at the time of study recruitment, such as Pfizer (BNT162b2), Moderna (mRNA-1273), AstraZeneca [AZD1222 (ChAdOx1)] or their approved combinations.Patients with a diagnosis of solid malignancy, treated at Princess Margaret Cancer Centre, University Health Network (PM-UHN), with either a standard of care (SOC) therapy or within a clinical trial were recruited. Blood samples from patients enroled were banked under the study oC. After processing, serum samples were aliquoted and stored in liquid nitrogen vapor phase for long term storage until time of downstream analysis.Sampling at 6 time points per patient were undertaken: at baseline (BASE), between 1st and 2nd vaccine doses (PRE2D), at 1 week (1 W), 1 month (1 M), 4–6 months (4–6 M) and 10–12 months (10–12 M) after the 2nd dose, as per Fig. Patient demographics, cancer diagnosis, past medical history, treatment details, concomitant medications, and immune related adverse events (irAEs) of ≥grade 2 severity by Common Terminology Criteria for Adverse events Version 5.0 (CTCAE 5.0) were collected prospectively and entered into a Medidata Rave database. This de-identified database is password protected and encrypted, to be compliant with PM-UHN standards for all investigator-initiated studies.The most updated Charlson Co-morbidity Index (CCI) was used to describe patients’ overall health status . CCI is This study was conducted with the approval of UHN Oncology Research Ethics Board (CAPCR/UHN REB #: 21–5797) and all patients signed informed consent. Participants who were concurrently treated in an interventional study with experimental treatment, were allowed to take part in this observational study (VIVACIOUS), according to their respective study protocols; or were not specifically disallowed to do so, if protocols were silent on this matter. The laboratory exploratory analysis was performed by scientific staff who was blinded to patients’ identity.Serum samples from patients (20 µL) were analyzed using Elecsys® Anti-SARS-CoV-2 S assay on the cobas e411 analyzer (Roche Diagnostics) for the quantitative detection of total antibodies (IgM/IgG/IgA) to the SARS-CoV-2 spike (S) protein receptor binding domain (RBD). This assay is a double-antigen sandwich electrochemiluminescence immunoassay, which uses streptavidin-coated microparticles to separate bound from unbound substances prior to applying a voltage to the electrode. Application of a voltage to the electrode then induces chemiluminescent emission which is measured by a photomultiplier. Samples were run in singleton. The measuring range of this assay is 0.40–250 U/mL (up to 2500 U/mL with on-board 1:10 dilution), with a concentration of ≥0.80 U/mL considered positive. Serum antibody levels against SARS-CoV-2 S protein are the result of immune response to either natural infection or vaccination but cannot distinguish between the two .The GenScript SARS-CoV-2 surrogate Virus Neutralization Test (sVNT) Kit was used to detect circulating neutralizing antibodies against SARS-CoV-2 that block the interaction between the viral S glycoprotein RBD with the angiotensin converting enzyme-2 (ACE2) cell surface receptor. The interaction between RBD and ACE2 leads to viral endocytosis into the host cells and subsequent viral replication. This assay detects any antibodies in serum and/or plasma that neutralize the RBD-ACE2 interaction.oC (30 min) with respective positive and negative controls. Samples were then loaded on plate wells and incubated at 37 oC for 15 min. Absorbance was measured at 450 nm. A result of ≥30% is considered positive for the detection of SARS-CoV-2 neutralizing antibodies . Similarly, the neutralizing antibody level at 1 M was 94.5%  for the IO group and 92.7%  for the non-IO group  and neutralization levels (97.7 vs 89.1 p = 0.003) compared to patients who received only two vaccine doses  after the second dose. Most cytokines  were undetectable for most patients. Overall, there were no statistically significant differences between either the baseline and the intra-dose measurement, nor the baseline and post 1 week measurement Fig. . Figure Patients treated with IO were prospectively followed-up for irAEs of more than grade 2 severity, from the date of first vaccine dose to 90 days following the 4th dose. Two out of 35 (5.7%) patients experienced immune related toxicity after 1st dose and before the 2nd vaccine dose, 8/34 (23.5%) after the 2nd dose and before the 3rd, 2/29 (7%) following the 3rd dose and 2/13 (15.3%) after the 4th dose Fig. . Most ofp < 0.001) and also IgM antibodies against: Human beta-2 glycoprotein 1 , Myeloperoxidase (MPO) , Nucleosome , and against Short Palate, Lung and Nasal epithelium carcinoma associated protein 2 (SPLUNC2)   between IgM anti-MPO autoantibody and IL-2 cytokine levels post-vaccination was found  following vaccination, as defined by a significant increase of their mean log-transformed fluorescent intensity (mean Log2 MFI). These autoantibodies were an IgG antibody against Myosin heavy-chain 6 (MYH6)  Fig. . There w01) Fig. . In viewund Fig. .Fig. 4AuWe conducted the VIVACIOUS study in a Canadian tertiary cancer centre during the height of initial SARS-CoV-2 pandemic waves. We aimed to understand multi-dimensional longitudinal immune correlates after anti-SARS-CoV-2 vaccination in solid cancer patients treated with immunotherapies and to compare these with patients receiving non-IO-based treatments. At that time, there was considerable vaccine hesitancy for some patients receiving cancer immunotherapies owing to emerging data on potentially autoimmune-related complications linked to mRNA vaccines (such as myocarditis). Thus, we further endeavoured to undertake an in-depth analysis of immunological and clinical parameters that could provide insights into this risk. Unlike other large series, we did this by adding multiple timepoint blood collections through the initial series and the booster vaccines doses and cytokine/autoantibody assays to assess this risk in a multi-dimensional manner.In performing this analysis, we found that IO-treated cancer patients were largely indistinguishable from non-IO-treated cancer patients with respect to absolute levels and neutralization capability of anti-RBD antibody generation after primary series anti-SARS-CoV-2 vaccination. Although others have reported reduced rates of seroconversion after vaccination amongst patients with haematologic malignancies, we confirm that patients with solid cancer receiving immunotherapy generate robust antibody responses. Both quantitative and functional testing of anti-SaRS-CoV-2 antibodies following vaccination seem to be largely unaffected by cancer immunotherapy, providing surrogate evidence that B cell function remains intact during treatment with immunotherapy. This finding is in keeping with what is reported in the literature so far , 33, 34 et al. showed that third and fourth vaccine doses can restore waning antibody titres in solid cancer patients [et al. recently reported that 3 vaccination doses are imperative for achieving neutralizing antibodies that offer protection against the Omicron variant in patients with solid malignancies [As with healthy subjects, we further observed that serological responses wane over the 4–6 months following the second vaccine dose which highlights the importance of implementing booster vaccinations , 8. A repatients . We notepatients , 37. Intgnancies . In our Although most of our participants generated robust antibody responses after initial primary vaccine series, we identified several patients with low seroconversion. Interestingly, all of these patients were male and 3/4 of them had a head and neck primary. In the literature, there is conflicting data as to whether patient sex or primary tumor type have an impact on seroconversion after vaccination. At least one publication reported higher antibody titres amongst female solid cancer patients, an observation also made in our cohort of patients . In our We next examined whether cytokine production differed after vaccination in IO-treated and non-IO-treated patients. Given the broad immunomodulatory impacts of vaccination and immunotherapies, we were somewhat surprised to find no significant, lasting differences after vaccination in any of the cytokines studied. A moderate positive correlation was observed between IL-2 and anti-MPO autoantibody generation and interestingly, a recent case report on cytokine release syndrome following anti-SARS-CoV-2 vaccination in a patient receiving ICI was accompanied by an increase in IL-2 .Having examined the impact of IO therapies on vaccine efficacy, we further explored whether vaccine administration might affect immunotherapy-related toxicities. Isolated thrombocytopenia (without thrombosis) has been described with mRNA-based COVID-19 vaccines  and a paLastly, we observed an increase of five antibodies against self-antigens after vaccination, an effect that was not treatment dependent. Human beta-2 glycoprotein 1, a critical regulator of complement and coagulation systems, is also a target of anti-phospholipid antibodies implicated in the antiphospholipid syndrome . In our Intriguingly, we observed an elevation of autoantibody against alpha-cardiac myosin heavy chain (MYH6), a component of the contractile system in cardiac muscle . MYH6 haOur study has various limitations, such as the small number of patients included and the lack of characterising immune responses at a cellular level by interrogating IL-2/IFN-γ-expressing T lymphocyte responses and functionality. Moreover, for our neutralization test, we considered 30% as the threshold for positivity, although we note that more recent evidence has suggested that higher neutralization levels are required for adequate protection against infection . In desiIn conclusion, we have found that administration of anti-SARS-CoV-2 vaccines can be safely delivered in patients undergoing cancer immunotherapy, without elevating the risk of immune related adverse events. The absolute and neutralizing levels of anti-RBD antibodies are decreasing with time and repeat booster vaccination is required to achieve desirable levels of immunity. The observation that anti-SARS-CoV-2 vaccination de-represses generation of five autoantibodies, a cardiac myosin amongst them, warrants further investigation.Supplementary Tables and Figures