Morbidity and Mortality Associated With Respiratory Virus Infections in Allogeneic Hematopoietic Cell Transplant: Too Little Defense or Harmful Immunity?

The impact on morbidity and mortality of Community Acquired Respiratory Virus (CARV) infections in patients undergoing Allogeneic Hematopoietic Cell Transplant (HCT) is widely studied. Here we give an overview of the current literature on the incidence and chance of progression to severe disease in this highly immune compromised population. We discuss the issue whether it is predominantly direct viral damage that causes clinical deterioration, or that it is in fact the allogeneic immuneresponse to the virus that is most important. This is an important question as it will guide therapeutic decision making. It asks for further collaborative studies focusing on sensitive surveillance with PCR techniques and relating clinical data with parameters of immune reconstitution

A "No-Touch" Antibody-Staining Method of Adherent Cells for High-Throughput Flow Cytometry in 384-Well Microplate Format for Cell-Based Drug Library Screening

In the last decade, screening compound libraries on live cells has become an important step in drug discovery. The abundance of compounds in these libraries requires effective high-throughput (HT) analyzing methods. Although current cell-based assay protocols are suitable for HT analyses, the analysis itself is often restrained to simple, singular outcomes. Incorporation of HT samplers on flow cytometers has provided an interesting approach to increase the number of measurable parameters and increase the sensitivity and specificity of analyses. Nonetheless, to date, the labor intensive and time-consuming strategies to detach and stain adherent cells before flow cytometric analysis has restricted use of HT flow cytometry (HTFC) to suspension cells. We have developed a universal "no-touch" HTFC antibody staining protocol in 384-well microplates to bypass washing and centrifuging steps of conventional flow cytometry protocols. Optimizing culture conditions, cell-detachment and staining strategies in 384-well microplates resulted in an HTFC protocol with an optimal stain index with minimal background staining. The method has been validated using six adherent cell lines and simultaneous staining of four parameters. This HT screening protocol allows for effective monitoring of multiple cellular markers simultaneously, thereby increasing informativity and cost-effectiveness of drug screening

Immune Reconstitution Kinetics as an Early Predictor for Mortality using Various Hematopoietic Stem Cell Sources in Children

AbstractThe severity of complications of allogeneic hematopoietic stem cell transplantation (HSCT) is governed mainly by the status of immune reconstitution. In this study, we investigated differences in immune reconstitution with different cell sources and the association between the kinetics of immune reconstitution and mortality. Immunophenotyping was performed every 2 weeks in children who had undergone HSCT between 2004 and 2008 at University Medical Center Utrecht. Lymphocyte reconstitution in the first 90 days after HSCT was studied in relation to mortality in 3 HSCT groups: matched sibling bone marrow (BM) recipients (35 patients), unrelated BM recipients (32 patients), and unrelated cord blood recipients (36 patients). The median age of recipients was 5.9 years (range, 0.1-21 years). The nature and speed of T cell, B cell, and natural killer (NK) cell reconstitution were highly dependent on the cell source. In the first 90 days after HSCT, faster B cell and NK cell reconstitution and delayed T cell reconstitution were shown in unrelated cord blood recipients compared with matched sibling BM and unrelated BM recipients. Of the lymphocyte subsets investigated, a large number of NK cells and a more rapid CD4+ immune reconstitution over time, resulting in sustained higher CD4+ counts, were the only predictors of a lower mortality risk in all cell sources. The final model showed that during the first 90 days, patients with an area under the CD4+ cell receiver- operating curve of >4,300 cells/day and no peak in CD4+ cell counts had the highest likelihood of survival (hazard ratio for mortality, 0.2; 95% confidence interval, 0.06-0.5). Our data indicate that CD4+ kinetics may be used to identify patients at greatest risk for mortality early after HSCT

Preeclampsia is Associated with lower Percentages of Regulatory T Cells in Maternal Blood

Objective: Immunological mechanisms are involved in the pathophysiology of preeclampsia. During pregnancy there is an increase in regulatory T (Treg) cells, which has an important role in regulating tolerance to the immunologically distinct fetus. We hypothesised that percentages of Treg cells are decreased in preeclamptic patients. Methods: Peripheral blood was obtained from 26 healthy pregnant controls and 18 preeclamptic patients. Treg cells were measured using flow-cytometry. Results: Women with pregnancies complicated by preeclampsia had significantly lower percentages of CD4(+)FOXP3(+) Treg cells. Conclusion: We conclude that a deficiency of regulatory T cells may play a role in the pathophysiology of preeclampsia

HEARING LOSS IN PATIENTS WITH MUCOPOLYSACCHARIDOSES-1 AND -6 AFTER HEMATOPOIETIC CELL TRANSPLANTATION; a longitudinal analysis

Hearing loss is frequently seen in mucopolysaccharidoses (MPS) patients. Although hematopoietic cell transplantation (HCT) increases overall survival, disease progression is observed in certain tissues. This study describes the course of hearing loss (HL) over time in transplanted MPS patients. Transplanted MPS patients between 2003 and 2018 were included and received yearly audiological evaluation, including auditory brainstem response (ABR) or pure tone audiometry (PTA). Twenty-eight MPS-1 and four MPS-6 patients were analyzed with a median follow-up of 5 years (range 11 months–16 years). Air conduction threshold improved significantly over time (P <.001) with a PTA 1-year post-HCT of 50 ± 0.7 dB to 23 ± 11 dB 13 years post-HCT. Bone conduction threshold worsened with a PTA 1 year post-HCT of 10 ± 7 dB to 18 ± 9 dB 13 years post-HCT (P =.34). The degree of HL varied from mainly mild-severe early after HCT to normal-mild at longer follow-up. The type of HL consisted of mainly conductive in the first years post-HCT in contrast to mainly sensorineural at longer follow-up. MRIs of the cerebellopontine angle did not show abnormalities. HL is still seen in patients with MPS despite HCT and consists of a conductive type early after HCT in contrast to a sensorineural type at longer follow-up in the majority of cases. Yearly follow-up of HL is necessary to timely intervene, as hearing is important in the speech and language development of children and their academic achievements

Clinical Grade Production of Wilms' Tumor-1 Loaded Cord Blood-Derived Dendritic Cells to Prevent Relapse in Pediatric AML After Cord Blood Transplantation

Hematopoietic cell transplantation (HCT) is a last resort, potentially curative treatment option for pediatric patients with refractory acute myeloid leukemia (AML). Cord blood transplantation (CBT) results in less relapses and less graft-versus-host disease when compared to other sources. Nevertheless, still more than half of the children die from relapses. We therefore designed a strategy to prevent relapses by inducing anti-AML immunity after CBT, using a CB-derived dendritic cell (CBDC) vaccine generated from CD34+ CB cells from the same graft. We here describe the optimization and validation of good manufacturing practice (GMP)-grade production of the CBDC vaccine. We show the feasibility of expanding low amounts of CD34+ cells in a closed bag system to sufficient DCs per patient for at least three rounds of vaccinations. The CBDCs showed upregulated costimulatory molecules after maturation and showed enhanced CCR7-dependent migration toward CCL19 in a trans-well migrations assay. CBDCs expressed Wilms' tumor 1 (WT1) protein after electroporation with WT1-mRNA, but were not as potent as CBDCs loaded with synthetic long peptides (peptivator). The WT1-peptivator loaded CBDCs were able to stimulate T-cells both in a mixed lymphocyte reaction as well as in an antigen-specific (autologous) setting. The autologous stimulated T-cells lysed not only the WT1+ cell line, but most importantly, also primary pediatric AML cells. Altogether, we provide a GMP-protocol of a highly mature CBDC vaccine, loaded with WT1 peptivator and able to stimulate autologous T-cells in an antigen-specific manner. Finally, these T-cells lysed primary pediatric AML demonstrating the competence of the CBDC vaccine strategy

Prophylactic use of liposomal amphotericin B in children and adolescents undergoing allogeneic hematopoietic cell transplantation: A 10-years single center experience

Background: Azoles are recommended as antifungal prophylaxis in decreasing the incidence of invasive fungal disease (IFD) in high-risk patients in pediatric oncology, including patients receiving allogeneic hematopoietic cell transplantation (HCT). However, azole related toxicity, pharmacological interactions with immunosuppressive medication and conditioning regimen and growing incidence of azole resistance makes this antifungal agent not ideal in the transplant setting. This study reports on the contemporary incidence and outcome of IFD after allogeneic HCT in children with prophylactic liposomal amphotericin B (L-AMB). Methods: This single-center retrospective study included all patients transplanted between 2012 and 2022. Primary endpoint was the incidence of IFD until hospital discharge post-transplant. Secondary aims were the incidence of IFD and survival 180 days after allogeneic HCT, the evaluation of toxicity of L-AMB and further risk factors for development of IFD during antifungal prophylaxis. Descriptive statistics were performed. Results: 161 pediatric patients received L-AMB. Incidence of breakthrough IFD post-transplant was 7.5 % (12/161). The 12 cases comprised of three invasive yeast infections (1.9 %), three probable (1.9 %) and six possible (3.7 %) mold infections. Adverse events were in 22.4 % of the patients, most of them mild and reversible. Discontinuation of L-AMB occurred in 2.5 % (4/161) of the patients due to severe hypersensitivity reactions. Conclusions: The risk of breakthrough IFD in pediatric patients undergoing allogeneic HCT under L-AMB prophylaxis is comparable with the reported risk under first line recommendation drugs for antifungal prophylaxis. If no hypersensitivity reaction occurs, L-AMB is tolerated with manageable side effects. This antifungal agent should therefore be considered as an alternative option to azoles in pediatric allogeneic HCT recipients

First principles investigation of exchange interactions in quasi-one-dimensional antiferromagnet CaV2O4

The effect of orbital degrees of freedom on the exchange interactions in the spin-1 quasi-one-dimensional antiferromagnet CaV2O4 is systematically studied. For this purpose a realistic low-energy model with the parameters derived from the first-principles calculations is constructed. The exchange interactions are calculated using both the theory of infinitesimal spin rotations near the mean-field ground state and the superexchange model, which provide a consistent description. The obtained behaviour of exchange interactions substantially differs from the previously proposed phenomenological picture based on the magnetic measurements and structural considerations, namely: (i) Despite quasi-one-dimensional character of the crystal structure, consisting of the zigzag chains of edge-sharing VO6 octahedra, the electronic structure is essentially three-dimensional, that leads to finite interactions between the chains; (ii) The exchange interactions along the legs of the chains appear to dominate; and (iii) There is a substantial difference of exchange interactions in two crystallographically inequivalent chains. The combination of these three factors successfully reproduces the behaviour of experimental magnetic susceptibility.Comment: 15 pages, 6 figures, supplementary materia

Predictors of outcomes in hematopoietic cell transplantation for Fanconi anemia

Allogeneic hematopoietic cell transplantation (HCT) remains the only cure for the hematologic manifestations of Fanconi anemia (FA). We performed retrospective predictor analyses for HCT outcomes in FA for pediatric and young adult patients transplanted between 2007 and 2020 across three large referral institutions. Eighty-nine patients, 70 with bone marrow failure +/- cytogenetic abnormalities, 19 with MDS/AML, were included. Five-year overall survival (OS) was 83.2% and event-free survival (EFS) was 74%. Age ≥19, HLA mismatch and year of HCT were multivariable predictors (MVPs) for OS, EFS and treatment-related mortality (TRM). In the pediatric group, TCD was a borderline MVP (P = 0.059) with 5-year OS of 73.0% in TCD vs. 100% for T-replete HCT. The cumulative incidence of day 100 grade II-IV aGvHD and 5-year cGvHD were 5.6% and 4.6%, respectively. Relapse in the MDS/AML subgroup occurred in 4 patients (16%). Graft failure was seen in 9 patients (TCD 6/37 [16%]; T-replete 3/52 [5.7%]). Six patients developed malignancy after HCT. Survival chances after HCT for FA are excellent and associated with high engrafted survival and low toxicity. Age ≥19, HLA mismatch, year of transplant and 'TCD in the <19 years group' (although borderline) were found to be negative predictors for survival