The CREBBP lysine acetyltransferase (KAT) is frequently mutated in follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL) and has been studied using gene knock-out (KO) in murine and human cells. However, the majority of CREBBP mutations encode amino acid substitutions within the catalytic KAT domain (CREBBP KAT-PM) that retain an inactive protein and have not been extensively characterized. Using CRISPR gene editing and extensive epigenomic characterization of lymphoma cell-lines, we found that CREBBP KAT-PM lead to unloading of CREBBP from chromatin, loss of enhancer acetylation and prevention of EP300 compensation. These enhancers were enriched for those that are dynamically loaded by CREBBP in the normal centroblast-to-centrocyte transition in the germinal center, including enhancers activated in response to CD40 signaling, leading to blunted molecular response to CD40 ligand in lymphoma cells. We provide evidence that CREBBP KAT-PM inhibits EP300 function by binding limiting quantities nuclear transcription factor, thereby preventing its compensatory activity. This effect can be experimentally overcome by expressing saturating quantities of transcription factor, or biologically attenuated by strong stimulation of CD40 signaling that increases nuclear transcription factor abundance. Importantly, epigenetic responses to CD40 signaling can be induced by enforcing CD4 T-cell engagement using a bispecific antibody, leading to CD40-dependent restoration of antigen presentation machinery in CREBBP KAT-PM cells and cell death. We therefore provide a mechanistic basis for enhancer deregulation by CREBBP KAT-PM and highlight enforced CD4 T-cell engagement as a potential approach for overcoming these effects.

Chronic granulomatous disease (CGD) is an inborn error of immunity characterized by defective NADPH oxidase function, leading to impaired microbial killing, recurrent infections and granulomatous inflammation. Allogenic hematopoietic stem cell transplantation (HSCT) is a curative treatment for CGD, particularly effective when a fully HLA-matched donor is available. However, the place of HLA-haploidentical HSCT remains less established. This retrospective, multicenter study analyzed outcomes of 64 CGD patients (53 males, 46 with X-linked CGD) who underwent a first HSCT with HLA-haploidentical family donors either with in vitro TCRαβ/CD19 depletion or in vivo depletion using post-transplant cyclophosphamide (PTCY). The mean age at transplant was 5.8 years (0-33 years). Patients exhibited a high disease burden prior to HSCT, with 45% experiencing infections in the 6 months prior to HSCT and 67% exhibiting inflammation. Outcomes in the entire cohort showed a 3-year overall survival (OS), event-free survival (EFS) and GvHD grade III to IV-free, event-free survival (GEFS) of 75.9%, of 70.2%, and of 56.1% respectively and were not impacted by the type of depletion or age. The cumulative incidence (CI) of primary graft failure was 20.6%. The CI of grade II to IV acute GvHD was higher in the PTCY group (p=0.04) whereas the CI of GVH grade III to IV was not. These results indicate that HLA-haploidentical HSCT is a feasible transplant option for CGD patients lacking HLA-matched donors. Further refinement of transplant protocols is necessary to mitigate graft failure and acute GvHD, ultimately improving access and outcomes for this life-saving therapy.

Multiple myeloma (MM) is a complex hematological malignancy characterized by genomic changes and transcriptomic dysregulation. Initial exome sequencing approaches have failed to identify any single frequent (>25%) mutation in the coding genome. However, using whole-genome sequencing (WGS), we found that one of the genomic regions most frequently mutated (62% of the MM patients) was the 5' untranslated (UTR) region and/or intron 1 of the BCL7A gene. RNA-seq data from a large cohort suggests a loss of BCL7A expression in a large majority of MM patients as compared to normal plasma cells. BCL7A loss of function in a panel of MM cell lines led to a highly proliferative phenotype in vitro and in vivo, while its ectopic expression significantly reduced cell viability, suggesting a tumor suppressor function for BCL7A in MM. We studied the cellular and molecular effects of BCL7A loss and observed that it endows myeloma cells with proliferative potential in cooperation with the plasma cell-defining transcription factor IRF4. BCL7A is involved in a direct protein-protein interaction with IRF4, limiting its DNA binding activity. Loss of BCL7A thus enhances the expression of IRF4-associated cytokines and reduces mitochondrial metabolism and ROS levels. Our study therefore suggests that BCL7A loss provides the necessary molecular change to allow IRF4-mediated transcriptional activity and MM cell growth and survival.

Relapsed or refractory (R/R) B-cell acute lymphoblastic leukemia (B-ALL) remains a challenging disease with a dismal prognosis. Despite the revolutionary impact of CD19-directed chimeric antigen receptor (CAR19)-T cell therapy, >50% of patients relapse within a year. Both leukemia cell-intrinsic factors favoring immune escape and poor CAR-T cell persistence contribute significantly to clinical failure. Moreover, the expression of immune checkpoint receptors (ICRs) and their ligands within the complex bone marrow (BM) microenvironment may contribute to leukemia progression and therapy resistance. Here, we comprehensively characterized the expression of ICRs and their ligands in leukemic blasts, T cells, and mesenchymal stromal cells (MSCs) from B-ALL BM samples at diagnosis (n=47) and relapse (n=38), comparing them with age-matched healthy BM controls. Our findings reveal a significant upregulated expression of TIM-3 in T cells, and its ligand galectin-9 in both blasts and MSCs throughout disease progression. The expression levels of galectin-9 in B-ALL blasts and TIM-3 in CAR19-T cells negatively correlate with clinical outcome. Furthermore, we demonstrate that galectin-9 impairs CAR19-T cell homeostasis and cytotoxicity. Notably, an engineered TIM-3-Fc decoy receptor, delivered either by primary T cells co-administered with CAR19-T cells or via a bicistronic all-in-one CAR19-TIM-3-Fc construct, improved the anti-leukemia efficacy and persistence of CAR19-T cells in B-ALL patient-derived xenograft models. Mechanistically, CAR19-TIM-3-Fc-T cell treatment promotes the in vivo expansion of both transduced and bystander effector and memory T cells, as determined by spectral flow cytometry. Collectively, these potent and persistent TIM-3-Fc decoy-armored CAR19-T cells offer a promising therapeutic strategy for R/R B-ALL patients.

Cytokine release syndrome (CRS) and immune effector cell (IEC)-associated neurotoxicity syndrome (ICANS) are common complications following IEC therapy for hematologic malignancies. This two-part, phase 2 study (INCB 39110-211) investigated safety and efficacy of itacitinib, a potent, highly selective Janus kinase 1 inhibitor with broad anti-inflammatory activity, for prevention of CRS and ICANS in patients receiving commercial CD19-directed IEC therapy. Patients in part 1 received once-daily itacitinib 200 mg 3 days before IEC therapy (axicabtagene ciloleucel [axi-cel], brexucabtagene autoleucel, or tisagenlecleucel) through Day 26, with guidelines for use of other CRS/ICANS interventions. In part 2 (double-blind), patients were randomized to receive twice-daily (bid) itacitinib 200 mg or placebo 3 days before IEC therapy with axi-cel. The primary endpoint was proportion of patients with CRS grade ≥2 by Day 14 using ASTCT consensus grading system. Overall, 111 patients were enrolled (63 in part 1; 48 in part 2), with 109 patients analyzed for efficacy and 110 for safety. Itacitinib 200 mg bid resulted in a significantly lower proportion of patients with grade ≥2 CRS by Day 14 versus placebo (17.4% vs 56.5%; P=0.003). The proportion of patients with grade ≥2 ICANS by Day 28 was lower than with placebo (8.7% vs 21.7%). Itacitinib was well tolerated, with pyrexia the most common TEAE (itacitinib 200 mg bid: 43.5%; placebo: 50.0%) and itacitinib-related cytopenias manageable. Importantly, itacitinib did not impact IEC therapy efficacy (objective response rate at 6 months: 39.1% for itacitinib 200 mg bid vs 26.1% for placebo). Trial registration: clinicaltrials.gov; #NCT04071366.

This study identified an increased risk of lymphoid malignancy in Shwachman-Diamond syndrome with an observed risk 38-fold higher than expected based on population data. Increased toxicity was observed with standard therapies in patients with SDS.

The Csa blood group antigen was identified more than 50 years ago but its genetic basis has yet to be elucidated. All our recent genomic investigation has failed to resolve the genetic basis of this enigmatic antigen. By investigating the association of the HNA-3a/b polymorphism (rs2288904-G/A) in SLC44A2 with clinical features of Sickle Cell Disease (SCD), we incidentally discovered that rare subjects with the homozygous HNA-3b/b genotype also carry the uncommon Cs(a-) phenotype. We genotyped this SNP in a cohort of 25 Cs(a-) subjects and found that all of them showed a HNA-3b/b genotype. This result suggests that the high-prevalence allele with rs2288904 (HNA-3a; 455G) encoding Arg152 encodes the high-prevalence Csa. Accordingly, anti-Csa does not react with SLC44A2null RBCs, SLC44A2 knockout K562 cells, and K562 cells expressing HNA-3b, confirming that the Csa and Csb antigens are carried on this protein. Furthermore, mass spectrometry analysis of SLC44A2 from neutrophils and RBCs, along with serological investigation, showed that, despite HNA-3a and Csa having the same genetic basis, anti-HNA-3a and anti-Csa recognize different epitopes on the SLC44A2 protein. Overall, our data resolve the genetic bases of the Cs(a-) and Cs(b-) blood phenotypes, with new insights on the anti-HNA-3a specificity.

Congenital thrombotic thrombocytopenic purpura (cTTP) is an ultrarare disorder characterized by thrombocytopenia, microangiopathic hemolytic anemia, and ischemic organ damage caused by pathogenic ADAMTS13 (a disintegrin and metalloproteinase with thrombospondin type 1 motif, member 13) variants. ADAMTS13-containing product, including fresh-frozen plasma (FFP), and plasma-derived factor VIII concentrates are commonly used to supply ADAMTS13; however, frequent hospital visits and allergic reactions are major drawbacks. A recombinant ADAMTS13 (rADAMTS13) was recently developed to address these issues. However, real-world evidence has not been reported owing to the rarity of this condition. This study compared the efficacy and safety of FFP and rADAMTS13 in 14 Japanese patients, including 5 patients with end-stage renal disease who were excluded from the phase 3 trial. The median peak level of ADAMTS13 activity 15 minutes after rADAMTS13 administration was significantly higher than that after FFP (68.4% vs 15.9%; P < .001). ADAMTS13 activity 1 week after rADAMTS13 administration was well maintained compared with FFP infusion (11.6% vs 5.1%; P < .001). Patients reported no allergic reactions after rADAMTS13 administration and appreciated the convenience of a single infusion of rADAMTS13, suggesting that rADAMTS13 is a safe and effective alternative to FFP in patients with cTTP. To our knowledge, this is the first publication of patients with cTTP who switched FFP to novel rADAMTS13 from Japanese real-world data.

Hemophagocytic lymphohistiocytosis (HLH) is a severe hyperinflammatory syndrome, and the overall survival of adult patients is poor. Ruxolitinib, a Janus kinase (JAK) 1/2 inhibitor, has shown promise in treating HLH and exerts synergistic effects when combined with dexamethasone. Our pilot study preliminarily demonstrated that the combination of ruxolitinib and dexamethasone (the Ru-D regimen) had a high response rate and led to favorable short-term survival outcomes in adult HLH patients. In this prospective phase 2 clinical trial, we propose the Ru-D regimen as a first-line treatment for adults newly diagnosed with HLH with unknown triggers (chictr.org.cn identifier: ChiCTR2100049996). A total of 28 Chinese patients were enrolled, and the median follow-up time was 25.1 months (range, 0.87-34.0). The 2-month OS rate (the primary endpoint) was 85.7%, which exceeded our expected 2-month OS rate of 75%. The 6-month and 2-year OS rates were 67.9% (19/28) and 53.6% (15/28), respectively. The median OS of lymphoma-associated HLH (LAHS) patients was 5.8 months, and most of these patients had NK/T-cell lymphoma. In contrast, the 2-year OS rate of non-LAHS patients was 75%. The overall response rate (ORR) was 85.7% (24/28); 17.9% (5/28) of patients achieved a complete response (CR) during the Ru-D regimen. Overall, the Ru-D regimen was well tolerated in HLH patients. This study demonstrates the efficacy and safety of the Ru-D regimen in adults newly diagnosed with HLH with unknown triggers and warrants a phase 3 randomized controlled study.

Genetic screening for severe congenital immuno-hematological diseases offers potential for early intervention, particularly through preemptive allogeneic stem cell transplantation (HSCT). However, the clinical value of such screening depends on precise prognostic predictions based on genotype-phenotype correlations and/or functional confirmation. We investigated familial hemophagocytic lymphohistiocytosis type 2 (FHL2), caused by PRF1 variants. Specifically we evaluated the clinical significance of the frequent PRF1 A91V variant, if present in trans with a predicted loss-of-function (pLOF) PRF1 variant, defined as "disease mutation" listed in the Human Gene Mutation Database. We combined clinical and functional data from our HLH-network registry with UK Biobank data to evaluate disease penetrance and clinical outcomes. Among 52 A91V/pLOF individuals in the registry, 39 (72%) showed FHL2-related manifestations with mean onset at 20 years. Four patients had recurrent disease, 15 were transplanted, and 14 died. Among 14 A91V/pLOF individuals identified by family screening (mean 29 years), however, only one was symptomatic. Moreover, among 21 A91V/pLOF carriers identified in 200,000 UK Biobank participants, 12 with genotypes identical to symptomatic registry patients, none had developed HLH by age 73. Premature stop pLOF alleles appeared more penetrant than missense variants, but functional data including perforin expression or cytotoxicity failed to predict disease manifestation. Our combined registry and population-based approach reveals significant variability in disease penetrance and severity among PRF1 A91V/pLOF carriers, with no clear association between genotype, functional data and clinical outcomes. This complexity illustrates the challenges of genetic screening and highlights the need for careful clinical decision-making regarding preemptive HSCT in asymptomatic carriers.

Follicular lymphoma (FL) represents a heterogeneous group of B-cell neoplasms with distinct genetic, epigenetic, microenvironmental, and clinical features. It is the most prevalent indolent non-Hodgkin lymphoma, characterized by a relapsing course and risk of transformation to aggressive diffuse large B-cell lymphoma. Recent advances in high-throughput sequencing, spatial transcriptomics, and imaging technologies uncovered genetic, epigenetic, and immunogenetic features underpinning FL, offering insights into its biology and potential therapeutic vulnerabilities. While FL is primarily driven by the hallmark t(14;18) translocation involving BCL2, its pathogenesis requires additional oncogenic mutations, particularly in genes regulating chromatin and histone modifications. These early genetic and epigenetic alterations promote the persistence and evolution of cancer precursor cells, setting the stage for lymphomagenesis. The tumor microenvironment is also crucial in FL progression and patient prognosis, with T-cells, stromal cells, and macrophages playing pivotal roles in facilitating tumor immune escape. Targeted therapies, including BCL2 inhibitors, epigenetic modulators, and immunotherapies, have emerged from this deeper understanding of FL biology. Achieving a cure for FL will require targeted therapies that selectively eliminate cancer precursor cells with minimal impact on normal cells, thus preventing relapse and avoiding harmful side effects. Eradicating minimal-residual-disease should be a primary objective rather than waiting for clinical relapse. Future research must prioritize the development of accurate experimental models, the elucidation of FL precursors, and a deeper understanding of its heterogeneity, dependencies, progression, and mechanisms driving transformation. Implementing targeted therapies at FL early stages, instead of the current "watch and wait" approach, will be essential to improve patient outcomes.

We report our single center experience demonstrating that HLA Class I alloimmunization predicts longer time to platelet engraftment, increased bleeding complications and higher transfusion requirements in patients undergoing gene-modified hematopoietic stem cell transplant for transfusion-dependent beta thalassemia.

Inherited bone marrow failure syndromes (IBMFS) are genetic disorders of impaired hematopoiesis that manifest in childhood with both cytopenias and extra-hematologic findings. While several IBMFS are categorized as ribosomopathies due to shared underlying ribosomal dysfunction, there is a broader disruption of the protein homeostasis (proteostasis) network across both classic and emerging IBMFS. Precise regulation of the proteostasis network, including mechanisms of protein synthesis, folding, trafficking, and degradation as well as associated stress response pathways, has emerged as essential for maintaining hematopoietic stem cell (HSC) function, providing new potential mechanistic insights into IBMFS pathogenesis. Furthermore, the varied clinical trajectories of patients with IBMFS with possible divergent outcomes of malignancy and spontaneous remission may reflect developmental and temporal changes in proteostasis activity and be driven by strong selective pressures to restore proteostasis. These new insights are spurring fresh therapeutic approaches to target proteostasis. Thus, further evaluation of proteostasis regulation and the consequences of proteostasis disruption in IBMFS could aid in developing new biomarkers, therapeutic agents, and preventative approaches for patients.

Type 1 plasminogen deficiency (PLGD), an ultra-rare disorder caused by PLG pathogenic variants, results in decreased levels of immunoreactive and functional plasminogen. PLGD can cause fibrin-rich pseudomembranes on mucosa that impair tissue/organ function, impact quality of life, and are potentially life-threatening. Lesion regression/resolution is facilitated by intravenous administration of human plasma-derived Glu-plasminogen (IV PLG concentrate), the first and only FDA-approved specific treatment, licensed in 2021. The diagnosis of PLGD is frequently delayed due to its rarity (1.6 per million) and the variability of the initial medical specialty contact determined by the affected systems. Symptoms are often attributed to more common conditions, like conjunctivitis, recurrent otitis media, reactive airway disease, etc. This manuscript presents clinical vignettes highlighting strategies for PLGD diagnosis and treatment. Initial evaluation includes a detailed history, laboratory assays, and, at times, radiologic or other procedures. Genetic testing can confirm the diagnosis. Consistent, knowledgeable management is required to promptly identify and treat lesions, even in initially asymptomatic individuals. Personalized treatment may include continuous prophylaxis or intermittent treatment with IV PLG concentrate, dependent on disease severity and clinical course. Specialized facilities like Hemophilia Treatment Centers offering multidisciplinary care represent medical homes for this ultra-rare disorder.

Paroxysmal nocturnal hemoglobinuria (PNH) is a non-malignant clonal hematopoietic disorder. There are two components to the pathogenesis of PNH: (i) a mutant stem cell, (ii) expansion of the mutant clone. Component (i) is straightforward: there is almost always an inactivating somatic mutation of the X-linked gene PIGA. As for (ii), different mechanisms may be involved. In rare cases expansion may be driven by independently arisen mutations (e.g. in JAK2): however, in the large majority of PNH patients such mutations are not found. Instead, clonal expansion may result from the escape of GPI-negative (PIGA mutant) stem cells from a T cell-mediated autoimmune attack on non-mutant stem cells. Several lines of evidence support this mechanism. (1) PNH is closely related to aplastic anemia (AA). (2) PIGA mutant microclones exist in normal people but they do not expand. (3) In PNH patients receiving syngeneic bone marrow transplantation PNH remission has occurred only when immunosuppressive conditioning was applied. (4) After targeted inactivation of piga in mice, large populations of GPI-negative blood cells are produced, but they gradually disappear rather than expanding. (5) There is evidence that cytotoxic T cells may spare GPI(-) stem cells, and CD1d-restricted GPI-specific T cells have been demonstrated in patients with PNH and with AA. Thus, the pathogenesis of PNH conforms to a Darwinian model within somatic cell populations: it results from a somatic mutation and from a specific selective environment. The findings in PNH are also highly relevant to the pathogenesis of AA.

The critical role of leukemic initiating cells as a therapy-resistant population in myeloid leukemia is well established, however, the molecular signatures of such cells in acute lymphoblastic leukemia remain underexplored. Moreover, their role in therapy response and patient prognosis is yet to be systematically investigated across various types of acute leukemia. We employed single-cell multiomics to analyze diagnostic specimens from 96 pediatric patients with acute lymphoblastic, myeloid, and lineage ambiguous leukemias. Through the integration of single-cell multiomics with extensive bulk RNA-Seq and clinical datasets, we uncovered a prevalent, chemotherapy-resistant subpopulation that resembles hematopoietic stem and progenitor cells (HSPC-like) and is associated with poor clinical outcomes across all subtypes investigated. We identified a core transcriptional regulatory network (TRN) in HSPC-like blasts that is combinatorically controlled by HOXA/AP1/CEBPA. This TRN signature can predict chemotherapy response and long-term clinical outcomes. We identified shared potential therapeutic targets against HSPC-like blasts, including FLT3, BCL2, and the PI3K pathway. Our study provides a framework for linking intra-tumoral heterogeneity with therapy response, patient outcome and discovery of new therapeutic targets for pediatric acute leukemias.