Good engraftment after reduced intensity targeted busulfan-based conditioning and matched related donor hematopoietic cell transplantation in hemoglobinopathies.

BACKGROUND: Hematopoietic cell transplantation (HCT) is an established curative therapy for transfusion-dependent thalassemia (TDT) and sickle cell disease (SCD). The latest American Society of Hematology guidelines recommend myeloablative preparative regimen in patients under 18 years of age. PROCEDURE: The objective was to demonstrate safety and efficacy of a reduced intensity conditioning (RIC) regimen including high-dose fludarabine, anti-thymocyte globulin, and targeted busulfan as a single alkylator to sub-myeloablative exposures. RESULTS: Between 2012 and 2021, 11 patients with SCD and five patients with TDT and matched related donor (MRD) HCT were included. The median age at transplantation was 8.3 years (range: 3.7-18.8 years). The median administered busulfan AUC was 67.4 mg/L×h (range: 60.7-80 mg/L×h). Overall survival was 93.8% and event-free survival 87.5% with one engrafted SCD patient with pre-existing moyamoya disease succumbing after drainage of a subdural hematoma. One SCD patient developed a secondary graft failure and was treated with a second HCT. Myeloid chimerism was full in all other patients with a median follow-up time of 4.1 years (range: 2.0-11.1 years), whereas T-cell donor chimerism was frequently mixed. CONCLUSION: This RIC conditioning followed by MRD HCT is sufficiently myeloablative to cure pediatric patients with hemoglobinopathies without the need for additional total body irradiation or thiotepa.

Reducing Mortality and Morbidity in Children with Severe Combined Immunodeficiency in Switzerland: the Role of Newborn Screening.

Newborn screening (NBS) for severe combined immunodeficiency (SCID) has been introduced in various countries with the aim of reducing morbidity and mortality. However, studies analyzing outcomes before and after the implementation of NBS programs remain limited. This study sought to compare the outcomes of SCID patients identified through Switzerland's national SCID NBS program, introduced in January 2019, with those of a historical cohort diagnosed between 2007 and 2019. The study included seven patients (32%) identified through NBS, and 15 (68%) born before NBS implementation and diagnosed based on clinical signs. Children in the NBS group were younger at diagnosis (median age 9 days vs 9 months, P = .002) and at hematopoietic stem cell transplantation (HSCT, median age 5 months vs 11 months, P = .003) compared to the clinical group. The NBS group had a lower incidence of infections before HSCT (29% vs 93%, P = .004). Although not statistically significant, the overall survival rate on last follow-up was higher in the NBS group (86% vs 67%, P = .62). Importantly, patients with active infections undergoing HSCT had a significantly lower overall survival probability compared to those without (P = .01). In conclusion, the introduction of NBS in Switzerland has led to earlier and often asymptomatic diagnosis of affected children, enabling timely intervention, infection prevention, and prompt treatment. These factors have contributed to higher survival rates in the NBS group. These findings underscore the critical importance of NBS for SCID, offering potential life-saving benefits through early detection and intervention.

The hyperinflammatory spectrum: from defects in cytotoxicity to cytokine control.

Cytotoxic lymphocytes kill target cells through polarized release of the content of cytotoxic granules towards the target cell. The importance of this cytotoxic pathway in immune regulation is evidenced by the severe and often fatal condition, known as hemophagocytic lymphohistiocytosis (HLH) that occurs in mice and humans with inborn errors of lymphocyte cytotoxic function. The clinical and preclinical data indicate that the damage seen in severe, virally triggered HLH is due to an overwhelming immune system reaction and not the direct effects of the virus per se. The main HLH-disease mechanism, which links impaired cytotoxicity to excessive release of pro-inflammatory cytokines is a prolongation of the synapse time between the cytotoxic effector cell and the target cell, which prompts the former to secrete larger amounts of cytokines (including interferon gamma) that activate macrophages. We and others have identified novel genetic HLH spectrum disorders. In the present update, we position these newly reported molecular causes, including CD48-haploinsufficiency and ZNFX1-deficiency, within the pathogenic pathways that lead to HLH. These genetic defects have consequences on the cellular level on a gradient model ranging from impaired lymphocyte cytotoxicity to intrinsic activation of macrophages and virally infected cells. Altogether, it is clear that target cells and macrophages may play an independent role and are not passive bystanders in the pathogenesis of HLH. Understanding these processes which lead to immune dysregulation may pave the way to novel ideas for medical intervention in HLH and virally triggered hypercytokinemia.

Hematopoietic cell transplantation in chronic granulomatous disease: a study of 712 children and adults.

Chronic granulomatous disease (CGD) is a primary immunodeficiency resulting in life-threatening infections and inflammatory complications. Allogeneic hematopoietic cell transplantation (allo-HCT) can cure the disease, but the indication to transplant remains controversial. We performed a retrospective multicenter study of 712 patients with CGD who underwent allo-HCT transplantation from March 1993 through December 2018. We studied 635 children (aged <18 years) and 77 adults. Median follow-up was 45 months. Median age at transplantation was 7 years (range, 0.1-48.6). Kaplan-Meier estimates of overall survival (OS) and event-free survival (EFS) at 3 years were 85.7% and 75.8%, respectively. In multivariate analysis, older age was associated with reduced survival and increased chronic graft-versus-host disease. Nevertheless, OS and EFS at 3 years for patients ≥18 years were 76% and 69%, respectively. Use of 1-antigen-mismatched donors was associated with reduced OS and EFS . No significant difference was found in OS, but a significantly reduced EFS was noted in the small group of patients who received a transplant from a donor with a >1 antigen mismatch. Choice of conditioning regimen did not influence OS or EFS. In summary, we report an excellent outcome after allo-HCT in CGD, with low incidence of graft failure and mortality in all ages. Older patients and recipients of 1-antigen-mismatched grafts had a less favorable outcome. Transplantation should be strongly considered at a younger age and particularly in the presence of a well-matched donor.

Targeted busulfan-based reduced-intensity conditioning and HLA-matched HSCT cure hemophagocytic lymphohistiocytosis.

Reduced-intensity/reduced-toxicity conditioning and allogeneic T-cell replete hematopoietic stem cell transplantation are curative in patients with hemophagocytic lymphohistiocytosis (HLH). Unstable donor chimerism (DC) and relapses are clinical challenges . We examined the effect of a reduced-intensity conditioning regimen based on targeted busulfan to enhance myeloid DC in HLH. The European Society for Bone and Marrow Transplantation-approved reduced-intensity conditioning protocol comprised targeted submyeloablative IV busulfan, IV fludarabine, and serotherapy comprising IV alemtuzumab (0.5-0.8 mg/kg) for unrelated-donor and IV rabbit anti-T-cell globulin for related-donor transplants. We assessed toxicity, engraftment, graft-versus-host disease (GHVD), DC in blood cell subtypes, and overall survival/event-free survival. Twenty-five patients from 7 centers were treated (median age, 0.68 year). The median total dose and cumulative area under the curve of busulfan was 13.1 mg/kg (6.4-26.4) and 63.1 mg/L × h (48-77), respectively. Bone marrow, peripheral blood stem cell, or cord blood transplants from HLA-matched related (n = 7) or unrelated (n = 18) donors were administered. Donor cells engrafted in all patients (median: neutrophils d+20/platelets d+28). At last follow-up (median, 36 months; range, 8-111 months), the median DC of CD15+ neutrophils, CD3+ T cells, and CD16+56+ natural killer cells was 99.5% (10-100), 97% (30-100), and 97.5% (30-100), respectively. Eight patients (32%) developed sinusoidal obstruction syndrome, resolving after defibrotide treatment. The 3-year overall survival and event-free survival rates were both 100%. None of the patients developed acute grade III to IV GHVD. Limited chronic GVHD was encountered in 4%. This regimen achieves excellent results with stable DC in patients with HLH.

Life-Threatening Primary Varicella Zoster Virus Infection With Hemophagocytic Lymphohistiocytosis-Like Disease in GATA2 Haploinsufficiency Accompanied by Expansion of Double Negative T-Lymphocytes.

Two unrelated patients with GATA2-haploinsufficiency developed a hemophagocytic lymphohistiocytosis (HLH)-like disease during a varicella zoster virus (VZV) infection. High copy numbers of VZV were detected in the blood, and the patients were successfully treated with acyclovir and intravenous immunoglobulins. After treatment with corticosteroids for the HLH, both patients made a full recovery. Although the mechanisms leading to this disease constellation have yet to be characterized, we hypothesize that impairment of the immunoregulatory role of NK cells in GATA2-haploinsufficiency may have accentuated the patients' susceptibility to HLH. Expansion of a double negative T-lymphocytic population identified with CyTOF could be a further factor contributing to HLH in these patients. This is the first report of VZV-triggered HLH-like disease in a primary immunodeficiency and the third report of HLH in GATA2-haploinsufficiency. Since HLH was part of the presentation in one of our patients, GATA2-haploinsufficiency represents a potential differential diagnosis in patients presenting with the clinical features of HLH-especially in cases of persisting cytopenia after recovery from HLH.

Mapping atomic motions with ultrabright electrons: towards fundamental limits in space-time resolution.

The long held objective of directly observing atomic motions during the defining moments of chemistry has been achieved based on ultrabright electron sources that have given rise to a new field of atomically resolved structural dynamics. This class of experiments requires not only simultaneous sub-atomic spatial resolution with temporal resolution on the 100 femtosecond time scale but also has brightness requirements approaching single shot atomic resolution conditions. The brightness condition is in recognition that chemistry leads generally to irreversible changes in structure during the experimental conditions and that the nanoscale thin samples needed for electron structural probes pose upper limits to the available sample or "film" for atomic movies. Even in the case of reversible systems, the degree of excitation and thermal effects require the brightest sources possible for a given space-time resolution to observe the structural changes above background. Further progress in the field, particularly to the study of biological systems and solution reaction chemistry, requires increased brightness and spatial coherence, as well as an ability to tune the electron scattering cross-section to meet sample constraints. The electron bunch density or intensity depends directly on the magnitude of the extraction field for photoemitted electron sources and electron energy distribution in the transverse and longitudinal planes of electron propagation. This work examines the fundamental limits to optimizing these parameters based on relativistic electron sources using re-bunching cavity concepts that are now capable of achieving 10 femtosecond time scale resolution to capture the fastest nuclear motions. This analysis is given for both diffraction and real space imaging of structural dynamics in which there are several orders of magnitude higher space-time resolution with diffraction methods. The first experimental results from the Relativistic Electron Gun for Atomic Exploration (REGAE) are given that show the significantly reduced multiple electron scattering problem in this regime, which opens up micron scale systems, notably solution phase chemistry, to atomically resolved structural dynamics.

Inhibition of novel protein kinase C-epsilon augments TRAIL-induced cell death in A549 lung cancer cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has great potential for cancer treatment since it provokes cell death in most tumor cells while leaving most normal cells unscathed. Some cancers, however, show resistance to TRAIL, indicating that TRAIL alone may be insufficient for cancer therapy. Here we studied whether the apoptotic susceptibility of A549 non-small cell lung cancer cells could be modulated by inhibiting protein kinase C (PKC). We show that an inhibitor with preference for novel PKC isozymes, NPC 15437, significantly augmented TRAIL sensitivity of A549 cells, as judged by assessing cell death and mitochondrial membrane potential. Likewise, NPC 15437 also significantly potentiated the responsiveness of DAOY medulloblastoma cells to TRAIL. In contrast, an inhibitor with preference for conventional PKC isozymes, Gö6976, did not augment TRAIL sensitivity of A549 cells. To further specify the PKC isozyme responsible for TRAIL sensitization, we used a peptide inhibitor with selectivity for the novel PKC isozyme epsilon, myr-PKC-epsilon V1-2. The inhibition of PKC-epsilon resulted in a significant amplification of the cytotoxic activity of TRAIL in A549 cells. Altogether, our study provides evidence for a considerable role of PKC-epsilon in the apoptotic responsiveness of A549 lung cancer cells, and possibly other malignancies, to TRAIL.