Hemophilia A, an X-linked bleeding disorder caused by deficiency of factor VIII (FVIII), is treated by protein replacement. Unfortunately, this regimen is costly due to the expense of producing recombinant FVIII as a consequence of its low-level secretion from mammalian host cells. FVIII expression activates the endoplasmic reticulum (ER) stress response, causes oxidative stress, and induces apoptosis. Importantly, little is known about the factors that cause protein misfolding and aggregation in metazoans. Here, we identified intrinsic and extrinsic factors that cause FVIII to form aggregates. We show that FVIII forms amyloid-like fibrils within the ER lumen upon increased FVIII synthesis or inhibition of glucose metabolism. Significantly, FVIII amyloids can be dissolved upon restoration of glucose metabolism to produce functional secreted FVIII. Two ER chaperone families and their cochaperones, immunoglobulin binding protein (BiP) and calnexin/calreticulin, promote FVIII solubility in the ER, where the former is also required for disaggregation. A short aggregation motif in the FVIII A1 domain (termed Aggron) is necessary and sufficient to seed β-sheet polymerization, and BiP binding to this Aggron prevents amyloidogenesis. Our findings provide novel insight into mechanisms that limit FVIII secretion and ER protein aggregation in general and have implication for ongoing hemophilia A gene-therapy clinical trials.

Daratumumab is a human monoclonal antibody targeting CD38, an antigen uniformly expressed by plasma cells in multiple myeloma and light-chain amyloidosis (AL). We report the results of a prospective multicenter phase 2 study of daratumumab monotherapy in AL (NCT02816476). Forty previously treated AL patients with a difference between involved and uninvolved free light chains (dFLC) >50 mg/L were included in 15 centers between September of 2016 and April of 2018. Patients received 6 28-day cycles of IV daratumumab, every week for cycles 1 and 2 and every 2 weeks for cycles 3 through 6. Median age was 69 years (range, 45-83). Twenty-six patients had ≥2 organs involved, with heart in 24 and kidney in 26. Median time from diagnosis to enrollment was 23 months (interquartile range, 4-122), with a median of 3 prior therapies (range, 1-5). At data cutoff (September of 2019), all patients discontinued therapy; 33 received the planned 6 cycles. Overall, 22 patients had hematological response, and 19 patients (47.5%) achieved very good partial response (dFLC <40 mg/L) or better. Median time to hematological response was 1 week. Patients with no response after 4 doses were unlikely to respond further. Renal and cardiac responses occurred in 8 and 7 patients, respectively. Daratumumab was well tolerated, with no unexpected adverse events. With a median follow-up of 26 months, the 2-year overall survival rate was 74% (95% confidence interval, 62-81). Daratumumab monotherapy is associated with deep and rapid hematological responses in previously treated AL patients, with a good safety profile. Further studies of daratumumab in combination regimens are warranted.

The TEMPI syndrome is a rare and acquired disorder characterized by 5 salient features, which compose its name: (1) telangiectasias; (2) elevated erythropoietin and erythrocytosis; (3) monoclonal gammopathy; (4) perinephric fluid collections; and (5) intrapulmonary shunting. Complete resolution of symptoms following treatment with plasma cell-directed therapy supports the hypothesis that the monoclonal antibody is causal and pathogenic. Understanding the basis of the TEMPI syndrome will depend on the identification of additional patients and a coordinated international effort.

There are unresolved questions regarding the association between persistent leukocytosis and risk of thrombosis and disease evolution in polycythemia vera (PV), as much of the published literature on the topic does not appropriately use repeated-measures data or time-dependent modeling to answer these questions. To address this knowledge gap, we analyzed a retrospective database of 520 PV patients seen at 10 academic institutions across the United States. Taking hematologic laboratory data at ∼3-month intervals (or as available) for all patients for duration of follow-up, we used group-based trajectory modeling to identify latent clusters of patients who follow distinct trajectories with regard to their leukocyte, hematocrit, and platelet counts over time. We then tested the association between trajectory membership and hazard of 2 major outcomes: thrombosis and disease evolution to myelofibrosis, myelodysplastic syndrome, or acute myeloid leukemia. Controlling for relevant covariates, we found that persistently elevated leukocyte trajectories were not associated with the hazard of a thrombotic event (P = .4163), but were significantly associated with increased hazard of disease evolution in an ascending stepwise manner (overall P = .0002). In addition, we found that neither hematocrit nor platelet count was significantly associated with the hazard of thrombosis or disease evolution.

Lymphomatoid granulomatosis (LYG) is a rare Epstein-Barr virus (EBV)-driven B-cell lymphoproliferative disease (LPD). This disease is hypothesized to result from defective immune surveillance of EBV, with most patients showing evidence of immune dysfunction, despite no known primary immunodeficiency. Pathologically, LYG is graded by the number and density of EBV+ atypical B cells, and other characteristic findings include an angioinvasive/angiodestructive reactive T-cell infiltrate and various degrees of necrosis. Clinically, LYG universally involves the lungs with other common extranodal sites, including skin, central nervous system, liver, and kidneys. Nodal and/or bone marrow involvement is extremely rare and, if present, suggests an alternative diagnosis. Treatment selection is based on histologic grade and underlying pathobiology with low-grade disease hypothesized to be immune-dependent and typically polyclonal and high-grade disease to be immune-independent and typically oligoclonal or monoclonal. Methods of augmenting the immune response to EBV in low-grade LYG include treatment with interferon-α2b, whereas high-grade disease requires immunochemotherapy. Given the underlying defective immune surveillance of EBV, patients with high-grade disease may have a recurrence in the form of low-grade disease after immunochemotherapy, and those with low-grade disease may progress to high-grade disease after immune modulation, which can be effectively managed with crossover treatment. In patients with primary refractory disease or in those with multiple relapses, hematopoietic stem cell transplantation may be considered, but its efficacy is not well established. This review discusses the pathogenesis of LYG and highlights distinct histopathologic and clinical features that distinguish this disorder from other EBV+ B-cell LPDs and lymphomas. Treatment options, including immune modulation and combination immunochemotherapy, are discussed.

Daratumumab has shown promising first results in systemic amyloid light-chain (AL) amyloidosis. We analyzed a consecutive series of 168 patients with advanced AL receiving either daratumumab/dexamethasone (DD, n = 106) or daratumumab/bortezomib/dexamethasone (DVD, n = 62). DD achieved a remission rate (RR) of 64% and a very good hematologic remission (VGHR) rate of 48% after 3 months. Median hematologic event-free survival (hemEFS) was 11.8 months and median overall survival (OS) was 25.6 months. DVD achieved a 66% RR and a 55% VGHR rate. Median hemEFS was 19.1 months and median OS had not been reached. Cardiac organ responses were noted in 22% with DD and 26% with DVD after 6 months. Infectious complications were common (Common Terminology Criteria [CTC] grade 3/4: DD 16%, DVD 18%) and likely related to a high rate of lymphocytopenia (CTC grade 3/4: DD 20%, DVD 17%). On univariable analysis, hyperdiploidy and gain 1q21 conferred an adverse factor for OS and hemEFS with DD, whereas translocation t(11;14) was associated with a better hemEFS. N-terminal prohormone of brain natriuretic peptide >8500 ng/L could not be overcome for survival with each regimen. Multivariable Cox regression analysis revealed plasma cell dyscrasia (difference between serum free light chains [dFLC]) >180 mg/L as an overall strong negative prognostic factor. Additionally, nephrotic-range albuminuria with an albumin-to-creatinine-ratio (ACR) >220 mg/mmol was a significantly adverse factor for hemEFS (hazard ratio, 2.1 and 3.1) with DD and DVD. Daratumumab salvage therapy produced good results and remission rates challenging any therapy in advanced AL. Outcome is adversely influenced by the activity of the underlying plasma cell dyscrasia (dFLC) and nephrotic-range albuminuria (ACR).

Erdheim-Chester disease (ECD) is characterized by the infiltration of tissues by foamy CD68+CD1a- histiocytes, with 1500 known cases since 1930. Mutations activating the MAPK pathway are found in more than 80% of patients with ECD, mainly the BRAFV600E activating mutation in 57% to 70% of cases, followed by MAP2K1 in close to 20%. The discovery of BRAF mutations and of other MAP kinase pathway alterations, as well as the co-occurrence of ECD with LCH in 15% of patients with ECD, led to the 2016 revision of the classification of histiocytoses in which LCH and ECD belong to the "L" group. Both conditions are considered inflammatory myeloid neoplasms. Ten percent of ECD cases are associated with myeloproliferative neoplasms and/or myelodysplastic syndromes. Some of the most striking signs of ECD are the long bone involvement (80%-95%), as well as the hairy kidney appearance on computed tomography scan (63%), the coated aorta (40%), and the right atrium pseudo-tumoral infiltration (36%). Central nervous system involvement is a strong prognostic factor and independent predictor of death. Interferon-α seems to be the best initial treatment of ECD. Since 2012, more than 200 patients worldwide with multisystem or refractory ECD have benefitted from highly effective therapy with BRAF and MEK inhibitors. Targeted therapies have an overall, robust, and reproducible efficacy in ECD, with no acquired resistance to date, but their use may be best reserved for the most severe manifestations of the disease, as they may be associated with serious adverse effects and as-yet-unknown long-term consequences.

Hemophagocytic lymphohistiocytosis (HLH) is a syndrome describing patients with severe systemic hyperinflammation. Characteristic features include unremitting fever, cytopenias, hepatosplenomegaly, and elevation of typical HLH biomarkers. Patients can develop hepatitis, coagulopathy, liver failure, central nervous system involvement, multiorgan failure, and other manifestations. The syndrome has a high mortality rate. More and more, it is recognized that while HLH can be appropriately used as a broad summary diagnosis, many pediatric patients actually suffer from an expanding spectrum of genetic diseases that can be complicated by the syndrome of HLH. Classic genetic diseases in which HLH is a typical and common manifestation include pathogenic changes in familial HLH genes (PRF1, UNC13D, STXBP2, and STX11), several granule/pigment abnormality genes (RAB27A, LYST, and AP3B1), X-linked lymphoproliferative disease genes (SH2D1A and XIAP), and others such as NLRC4, CDC42, and the Epstein-Barr virus susceptibility diseases. There are many other genetic diseases in which HLH is an infrequent complication of the disorder as opposed to a prominent manifestation of the disease caused directly by the genetic defect, including other primary immune deficiencies and inborn errors of metabolism. HLH can also occur in patients with underlying rheumatologic or autoinflammatory disorders and is usually designated macrophage activation syndrome in those settings. Additionally, HLH can develop in patients during infections or malignancies without a known (or as-yet-identified) genetic predisposition. This article will attempt to summarize current concepts in the pediatric HLH field as well as offer a practical diagnostic and treatment overview.

Systemic mastocytosis (SM) has greatly benefited from the broad application of precision medicine techniques to hematolymphoid neoplasms. Sensitive detection of the recurrent KIT D816V mutation and use of next-generation sequencing (NGS) panels to profile the genetic landscape of SM variants have been critical adjuncts to the diagnosis and subclassification of SM, and development of clinical-molecular prognostic scoring systems. Multilineage KIT involvement and multimutated clones are characteristic of advanced SM (advSM), especially SM with an associated hematologic neoplasm (AHN). A major challenge is how to integrate conventional markers of mast cell disease burden (percentage of bone marrow mast cell infiltration and serum tryptase levels) with molecular data (serial monitoring of both KIT D816V variant allele frequency and NGS panels) to lend more diagnostic and prognostic clarity to the heterogeneous clinical presentations and natural histories of advSM. The approval of the multikinase/KIT inhibitor midostaurin has validated the paradigm of KIT inhibition in advSM, and the efficacy and safety of second-generation agents, such as the switch-control inhibitor ripretinib (DCC-2618) and the D816V-selective inhibitor avapritinib (BLU-285) are being further defined in ongoing clinical trials. Looking forward, perhaps the most fruitful marriage of the advances in molecular genetics and treatment will be the design of adaptive basket trials that combine histopathology and genetic profiling to individualize treatment approaches for patients with diverse AHNs and relapsed/refractory SM.

Fanconi anemia (FA) is the most common genetic cause of bone marrow failure and is caused by inherited pathogenic variants in any of 22 genes. Of these, only FANCB is X-linked. We describe a cohort of 19 children with FANCB variants, from 16 families of the International Fanconi Anemia Registry. Those with FANCB deletion or truncation demonstrate earlier-than-average onset of bone marrow failure and more severe congenital abnormalities compared with a large series of FA individuals in published reports. This reflects the indispensable role of FANCB protein in the enzymatic activation of FANCD2 monoubiquitination, an essential step in the repair of DNA interstrand crosslinks. For FANCB missense variants, more variable severity is associated with the extent of residual FANCD2 monoubiquitination activity. We used transcript analysis, genetic complementation, and biochemical reconstitution of FANCD2 monoubiquitination to determine the pathogenicity of each variant. Aberrant splicing and transcript destabilization were associated with 2 missense variants. Individuals carrying missense variants with drastically reduced FANCD2 monoubiquitination in biochemical and/or cell-based assays tended to show earlier onset of hematologic disease and shorter survival. Conversely, variants with near-normal FANCD2 monoubiquitination were associated with more favorable outcome. Our study reveals a genotype-phenotype correlation within the FA-B complementation group of FA, where severity is associated with level of residual FANCD2 monoubiquitination.

Langerhans cell histiocytosis (LCH) is caused by clonal expansion of myeloid precursors that differentiate into CD1a+/CD207+ cells in lesions that leads to a spectrum of organ involvement and dysfunction. The pathogenic cells are defined by constitutive activation of the MAPK signaling pathway. Treatment of LCH is risk-adapted: patients with single lesions may respond well to local treatment, whereas patients with multisystem disease require systemic therapy. Although survival rates for patients without organ dysfunction is excellent, mortality rates for patients with organ dysfunction may reach 20%. Despite progress made in the treatment of LCH, disease reactivation rates remain above 30%, and standard second-line treatment is yet to be established. Treatment failure is associated with increased risks for death and long-term morbidity, including LCH-associated neurodegeneration. Early case series report promising clinical responses in patients with relapsed and refractory LCH treated with BRAF or MEK inhibitors, although potential for this strategy to achieve cure remains uncertain.