Safety and efficacy of one and two booster doses of SARS-CoV-2 mRNA vaccines in kidney transplant recipients: A randomized clinical trial.

BACKGROUND: Kidney transplant recipients are at risk for a severe course of COVID-19 with a high mortality rate. A considerable number of patients remains without a satisfactory serological response after the baseline and adjuvant SARS-CoV-2 vaccination schedule. METHODS: In this prospective, randomized study, we evaluated the efficacy and safety of one and two booster doses of mRNA vaccines (either mRNA-1273 or BNT162b2) in 125 COVID-19 naive, adult kidney transplant recipients who showed an insufficient humoral response (SARS-CoV-2 IgG <10 AU/ml) to the previous 2-dose vaccination schedule. The primary outcome was the difference in the rate of a positive antibody response (SARS-CoV-2 IgG ≥10 AU/ml) between one and two booster doses at 1 month after the final booster dose. RESULTS: A positive humoral response was observed in 36 (62%) patients receiving two booster doses and in 28 (44%) patients receiving one booster dose (odds ratio [OR], 2.10, 95% confidence interval [CI], 1.02-4.34, p = .043). Moreover, median SARS-CoV-2 IgG levels were higher with two booster doses (p = .009). The number of patients with positive virus neutralizing antibody (VNA) levels was numerically higher with two booster doses compared to one booster dose, but without statistical significance (66% vs. 50%, p = .084). There was no significant difference in positive seroconversions rate and antibody levels between mRNA-1273 and BNT162b2. CONCLUSION: A higher number of kidney transplant recipients achieved a positive antibody response after two booster doses compared to one booster dose.

A Randomized Trial of Valganciclovir Prophylaxis Versus Preemptive Therapy in Kidney Transplant Recipients.

SIGNIFICANCE STATEMENT: Although cytomegalovirus (CMV) infection is an important factor in the pathogenesis of kidney allograft rejection, previous studies have not determined the optimal CMV prevention strategy to avoid indirect effects of the virus. In this randomized trial involving 140 kidney transplant recipients, incidence of acute rejection at 12 months was not lower with valganciclovir prophylaxis (for at least 3 months) compared with preemptive therapy initiated after detection of CMV DNA in whole blood. However, prophylaxis was associated with a lower risk of subclinical rejection at 3 months. Although both regimens were effective in preventing CMV disease, the incidence of CMV DNAemia (including episodes with higher viral loads) was significantly higher with preemptive therapy. Further research with long-term follow-up is warranted to better compare the two approaches. BACKGROUND: The optimal regimen for preventing cytomegalovirus (CMV) infection in kidney transplant recipients, primarily in reducing indirect CMV effects, has not been defined. METHODS: This open-label, single-center, randomized clinical trial of valganciclovir prophylaxis versus preemptive therapy included kidney transplant recipients recruited between June 2013 and May 2018. After excluding CMV-seronegative recipients with transplants from seronegative donors, we randomized 140 participants 1:1 to receive valganciclovir prophylaxis (900 mg, daily for 3 or 6 months for CMV-seronegative recipients who received a kidney from a CMV-seropositive donor) or preemptive therapy (valganciclovir, 900 mg, twice daily) that was initiated after detection of CMV DNA in whole blood (≥1000 IU/ml) and stopped after two consecutive negative tests (preemptive therapy patients received weekly CMV PCR tests for 4 months). The primary outcome was the incidence of biopsy-confirmed acute rejection at 12 months. Key secondary outcomes included subclinical rejection, CMV disease and DNAemia, and neutropenia. RESULTS: The incidence of acute rejection was lower with valganciclovir prophylaxis than with preemptive therapy (13%, 9/70 versus 23%, 16/70), but the difference was not statistically significant. Subclinical rejection at 3 months was lower in the prophylaxis group (13% versus 29%, P = 0.027). Both regimens prevented CMV disease (in 4% of patients in both groups). Compared with prophylaxis, preemptive therapy resulted in significantly higher rates of CMV DNAemia (44% versus 75%, P < 0.001) and a higher proportion of patients experiencing episodes with higher viral load (≥2000 IU/ml), but significantly lower valganciclovir exposure and neutropenia. CONCLUSION: Among kidney transplant recipients, the use of valganciclovir prophylaxis did not result in a significantly lower incidence of acute rejection compared with the use of preemptive therapy. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: Optimizing Valganciclovir Efficacy in Renal Transplantation (OVERT Study), ACTRN12613000554763 .

Insufficient response to mRNA SARS-CoV-2 vaccine and high incidence of severe COVID-19 in kidney transplant recipients during pandemic.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mRNA vaccination may fail to sufficiently protect transplant recipients against coronavirus disease 2019 (COVID-19). We retrospectively evaluated COVID-19 in kidney transplant recipients (n = 226) after BNT162b2 mRNA vaccine administration. The control group consisted of unvaccinated patients (n = 194) during the previous pandemic wave. We measured anti-spike protein immunoglobulin G (IgG) levels and cellular responses, using enzyme-linked immunosorbent spot assay, in a prospective cohort after vaccination (n = 31) and recovery from COVID-19 (n = 19). COVID-19 was diagnosed in 37 (16%) vaccinated and 43 (22%) unvaccinated patients. COVID-19 severity was similar in both groups, with patients exhibiting a comparable need for hospitalization (41% vs. 40%, p = 1.000) and mortality (14% vs. 9%, p = .726). Short posttransplant periods were associated with COVID-19 after vaccination (p < .001). Only 5 (16%) patients achieved positive SARS-CoV-2 IgG after vaccination, and 17 (89%, p < .001) recovered from COVID-19 (median IgG levels, 0.6 vs. 52.5 AU/ml, p < .001). A cellular response following vaccination was present in the majority (n = 22, 71%), with an increase in interleukin 2 secreting T cells (p < .001). Despite detectable T cell immunity after mRNA vaccination, kidney transplant recipients remained at a high risk of severe COVID-19. Humoral responses induced by vaccination were significantly lower than that after COVID-19.

Long-Term Cryopreservation Does Not Affect Quality of Peripheral Blood Stem Cell Grafts: A Comparative Study of Native, Short-Term and Long-Term Cryopreserved Haematopoietic Stem Cells.

Cryopreserved haematopoietic progenitor cells are used to restore autologous haematopoiesis after high dose chemotherapy. Although the cells are routinely stored for a long period, concerns remain about the maximum storage time and the possible negative effect of storage on their potency. We evaluated the effect of cryopreservation on the quality of peripheral stem cell grafts stored for a short (3 months) and a long (10 years) period and we compared it to native products.The viability of CD34+ cells remained unaffected during storage, the apoptotic cells were represented up to 10% and did not differ between groups. The clonogenic activity measured by ATP production has decreased with the length of storage (ATP/cell 1.28 nM in native vs. 0.63 in long term stored products, P < 0.05). Only borderline changes without statistical significance were detected when examining mitochondrial and aldehyde dehydrogenase metabolic activity and intracellular pH, showing their good preservation during cell storage. Our experience demonstrates that cryostorage has no major negative effect on stem cell quality and potency, and therefore autologous stem cells can be stored safely for an extended period of at least 10 years. On the other hand, long term storage for 10 years and longer may lead to mild reduction of clonogenic capacity. When a sufficient dose of stem cells is infused, these changes will not have a clinical impact. However, in products stored beyond 10 years, especially when a low number of CD34+ cells is available, the quality of stem cell graft should be verified before infusion using the appropriate potency assays.

Analysis of Sensitization Profiles in Central European Allergy Patients Focused on Animal Allergen Molecules.

INTRODUCTION: Frequently observed multiple sensitizations to several animals highlights the importance of a molecular diagnosis, distinguishing between sensitizations specific to single species and sensitizations due to cross-reactivity. OBJECTIVE: The aim of our study was to assess the usefulness of a molecular diagnosis in the description of sensitization profiles in allergy patients living in Central Europe, with a particular focus on animal-derived molecules. METHODS: The molecular diagnosis was performed using the ImmunoCAP ISAC microarray. Results of 1,255 allergy patients were subjected to statistical analysis. RESULTS: The highest sensitization rates were observed for uteroglobin Fel d 1 (31.8%) and kallikrein Can f 5 (16.4%), followed by animal lipocalins Can f 1 (13.9%), Equ c 1 (6.2%), Fel d 4 (5.3%), Can f 2 (4.2%), and Mus m 1 (4.1%). Sensitization rates to serum albumins Fel d 2, Can f 3, Equ c 3, and Bos d 6 were very low, with the highest being 3.2% to Fel d 2. Detailed subanalysis confirmed the dominant role of Fel d 1 or Can f 5 and/or Can f 1 in cat- or dog-sensitized patients, respectively. Further analysis focused on lipocalins and albumins confirmed a high rate of cosensitizations within both groups. CONCLUSION: The sensitization to animal allergen molecules is very frequent in Central Europe. The most common is sensitization to species-specific cat uteroglobin Fel d 1 and dog kallikrein Can f 5, followed by sensitizations to animal lipocalins. Our data suggest that commonly observed multiple sensitizations detected by extract approach can be explained not only by true cosensitization, but also by cross-reactivity, mainly in the frame of lipocalins. Cross-reactive serum albumins are minor sensitizers and are probably not important from this point of view.

Cross-sectional study on sensitization to mite and cockroach allergen components in allergy patients in the Central European region.

BACKGROUND: The major sources of allergens in the indoor air include house dust mites, dander derived from domestic animals and rodents, cockroach, and several fungi. Mites are the main cause of allergies in some countries with a warmer climate, but the epidemiological significance of mite and cockroach allergens in Central Europe has not been established yet. METHODS: We assessed sensitization profiles of allergy patients in a Central European region in regard to sensitization to mites and cockroach. We used molecular diagnosis by means of the microarray ISAC, and we investigated 1766 patients with clinical suspicion to an allergic disorder. 1255 of them were positive to at least one allergen component, and this group was subjected to statistical analysis. RESULTS: The sensitization to at least one mite-specific molecule (Der p 1, 2, Der f 1, 2) was observed relatively frequently in 32.7% of patients. Specific IgE to mite group 2 molecules is almost fully cross-reactive. Group 1 allergens are also cross-reactive, but in some patients, a species-specific response was observed. Relatively high rate of sensitization both to group 1 and 2 allergens in our patients indicates the greater role of co-sensitizations. Isolated sensitizations to molecules derived from glyciphagid mites Lep d 2 and/or Blo t 5 without sensitization to other mite-derived molecules were observed only exceptionally (in 0.6% of cases). True sensitization to at least one cockroach-specific molecule (Bla g 1, 2, 5) was very rare (in 0.6% of cases), and nearly all of them were co-sensitizations with other noncockroach-derived molecules. Sensitization to an inhaled tropomyosin was observed rarely in 2.2% of patients (Der p 10 in 1.9% and Bla g 7 in 1.5%). Co-sensitization of inhaled tropomyosins with the respective mite- or cockroach-specific molecules was observed only in the minority of patients suggesting the different route of sensitization being more frequent. CONCLUSIONS: The majority of patients are co-sensitized to several molecules of the respective allergen source. The knowledge of this molecular spectrum of sensitization is important for optimal diagnosis and treatment in respect to allergen content in mite extracts used for diagnostic and therapeutic purposes. In regard to the sensitization patterns of Central European patients, it is necessary to point out the importance of quantifying at least three major mite components Der f 1, Der p 1 and Der f 2 (or Der p 2).

Utility of laboratory testing for the diagnosis of Hymenoptera venom allergy.

BACKGROUND: A diagnosis of Hymenoptera venom allergy is based on clinical history and the results of skin tests and/or laboratory methods. OBJECTIVE: To analyze the utility of available laboratory tests in diagnosing Hymenoptera venom allergy. METHODS: Ninety-five patients with Hymenoptera venom allergy with a history of bee (35) or wasp (60) anaphylactic sting reaction and positive skin test with bee or wasp venom were included in this analysis. Specific immunoglobulin E (to bee venom extract, wasp venom extract, available recombinant molecules, and a basophil activation test with venom extracts were assessed in all the patients. Test sensitivity and specificity were calculated by using standard threshold values; then, receiver operating characteristic curve analysis was performed to compute optimal threshold values. Also, statistical analysis of the utility of different combinations of laboratory tests was performed. RESULTS: The optimal threshold values were revealed to be the following: 1.0 kIU/L for bee venom extract (sensitivity, 97.14%; specificity, 100%), 0.35 kIU/L for rApi m 1 (sensitivity, 68.57%; specificity, 100%), 1.22 kIU/L for wasp venom extract (sensitivity, 88.33%; specificity, 95.45%), 0.7 kIU/L for rVes v 5 (sensitivity, 86.67%; specificity, 95.45%), 1.0 kIU/L for rVes v 1 (sensitivity, 56.67%; specificity, 95.45%), 6.5% for basophil activation test with bee venom extract (sensitivity, 80%; specificity, 95.45%), and 4.5% for basophil activation test with wasp venom extract (sensitivity, 91.53%; specificity, 95.45%). The best test combinations were found to be the following: bee venom extract plus rApi m 1 (sensitivity, 97.14%; specificity, 95.45%) in bee and either wasp venom extract plus rVes v 5, or rVes v 5 plus rVes v 1 (both sensitivity, 98.33%; specificity, 95.45%) in patients with wasp venom allergy. CONCLUSION: Our analysis confirmed that currently used laboratory tests represent effective tools in diagnosing Hymenoptera venom allergy. Moreover, our probabilistic approach offered another way to interpret concrete values of laboratory test results and opened possible direction on how to optimize the laboratory diagnostic procedure.

In vitro testing of immunosupressive effects of mesenchymal stromal cells on lymphocytes stimulated with alloantigens.

AIMS: Mesenchymal stromal cells (MSC) derived from adult bone marrow or adipose tissue offer the potential to open a new frontier in medicine. MSC are involved in modulating immune response and tissue repair in vitro and in vivo. Experimental evidence and preliminary clinical studies have demonstrated that MSC exhibit an important immunomodulatory function in patients with graft versus host disease (GVHD) following allogeneic hematopoietic stem cell transplantation. The immunosuppressive properties of MSC have already been exploited in the clinical setting. However the precise mechanisms are being still investigated. METHODS: We examined the immunosuppressive function of MSC by coculturing them with stimulated HLA incompatible allogeneic lymphocytes in a mixed lymphocyte culture test. The metabolic and proliferative activity of lymphocytes was determined by MTT test. RESULTS: After stimulation with alloantigens the presence of MSC caused significant decrease of absorbance levels by 62% (P<0.01), 26% (P<0.01) and 6% (P=0.0437) in comparison to positive control depending on the MSC/lymphocyte ratio (1:5, 1:50, 1:500). The mitogenic stimulation of lymphocytes with fMLP or PHA was also significantly reduced during MSC cocultivation. The absorbance was reduced by 42% (P<0.001) and 67% (P<0.001). CONCLUSIONS: Allogeneic bone marrow is an ideal source of MSC for clinical application. The experiments confirmed the dose-dependent inhibitory effect of MSC on lymphocyte proliferation triggered by cellular or mitogenic stimulation. The mixed lymphocyte culture test offers a simple method for characterization and verification of the immunosuppressive potential of MSC, being prepared for clinical use.

A comprehensive analysis of middle-European molecular sensitization profiles to pollen allergens.

Molecular diagnosis of allergy and microarray technology have opened a completely new avenue of insight into sensitization profiles from both the clinical and the epidemiological point of view. We used this innovative tool in the description of sensitization patterns in pollen-sensitized patients in Middle Europe. Immunoglobulin E detection using 112 different allergenic molecules was carried out employing the ImmunoCAP ISAC microarray system. Sera from 826 patients sensitized to at least one pollen-derived molecule were subjected to analysis. The highest observed sensitization rate was 81.0% to grass-specific molecules (the most frequent being Phl p 1; 69.6%). The second most frequent sensitization was 54.8% to Betulaceae-specific molecules (Bet v 1; 54.2%). Together, grasses and Betulaceae components (and their cosensitizations with other components) comprised the vast majority of pollen sensitizations. Unexpectedly frequently observed sensitizations were those to Cupressaceae-specific molecules (14.1%), Oleaceae-specific molecules (10.8%), and the plane tree-derived molecule Pla a 2 (15.5%). The sensitization rates for all other molecules were within the expected range (Art v 1, 13.6%; Pla l 1, 9.6%; Che a 1, 8.4%; Par j 2, 0.9%; Amb a 1, 0.8%, and Sal k 1, 0.5%). Cross-reacting molecule sensitization rates were found to be 12.4% for profilins, 5.0% for polcalcins, and 6.4% for lipid transfer proteins. Molecular diagnosis of allergy gives a more precise and comprehensive insight into pollen sensitization patterns than extract-based testing, allowing a better understanding of the sensitization process and regional differences. The data presented here may help to improve the diagnostic and allergen-specific treatment procedures in the respective region.

Expanded cryopreserved mesenchymal stromal cells as an optimal source for graft-versus-host disease treatment.

Mesenchymal stromal cells (MSC) are fibroblast-like cells present in different types of tissues. Their immunomodulatory potential represents a promising method for post-transplant immunotherapy in the treatment of GVHD (graft-versus-host disease) with suboptimal response to standard immunosuppression. In this study we tested influence of 1-8 month-long cryopreservation on ability of MSC to suppress activation of non-specifically stimulated lymphocytes. We did not observe any changes in proliferation capacity of MSC after thawing. Lymphocytes metabolic activity was inhibited by 30% and number of dividing cells was three times smaller in the presence of MSC. Two activation markers were studied (CD25 and CD69) to confirm preservation of functional cell integrity. Expression of CD25 antigen on CD3(+)CD4(+) and CD3(+)CD4(-) cells was decreased in all co-cultivated samples. Level of CD69 expression on CD3(+)CD4(+) cells was lower in samples with added MSC (10-15% on day +2) but without reaching statistical significance. The lower expression (approximately 5%) was observed also on CD4-cells. The study confirms the preservation of immunomodulatory properties of cryopreserved and re-expanded MSC. Aliquots with cryopreserved cells can represent an optimal source for a quick preparation of MSC cell product with the possibility to apply the same cells repeatedly.