Clonal haematopoiesis across the age spectrum of vasculitis patients with Takayasu's arteritis, ANCA-associated vasculitis and giant cell arteritis.

OBJECTIVES: Ageing and inflammation are associated with clonal haematopoiesis (CH), the emergence of somatic mutations in haematopoietic cells. This study details CH in patients with systemic vasculitis in association with clinical, haematological and immunological parameters. METHODS: Patients with three forms of vasculitis were screened for CH in peripheral blood by error-corrected sequencing. Relative contributions of age and vasculitis on CH prevalence were calculated using multivariable logistic regression. Clonal hierarchies were assessed by proteogenomic single-cell DNA sequencing, and functional experiments were performed in association with CH status. RESULTS: Patients with Takayasu's arteritis (TAK; n=70; mean age=33.2 years), antineutrophil cytoplasmic antibody-associated vasculitis (AAV; n=47; mean age=55.3 years) and giant cell arteritis (GCA; n=59; mean age=71.2 years) were studied. CH, most commonly in DNMT3A and TET2, was detected in 34% (60/176) of patients versus 18% (28/151) of age-matched controls (p<0.01). Prevalence of CH was independently associated with age (standardised B=0.96, p<0.01) and vasculitis (standardised B=0.46, p<0.01), occurring in 61%, 32% and 13% of patients with GCA, AAV and TAK, respectively. Both branched and linear clonal trajectories showed myeloid-lineage bias, and CH was associated with markers of cellular activation. In GCA, mutations were detected in temporal artery biopsies, and clinical relapse correlated with CH in a dose-dependent relationship with clone size. CONCLUSIONS: Age was more strongly associated with CH prevalence than inflammation in systemic vasculitis. Clonal profile was dominated by DNMT3A mutations which were associated with relapse in GCA. CH is not likely a primary causal factor in systemic vasculitis but may contribute to inflammation.

When to consider inherited marrow failure syndromes in adults.

The inherited bone marrow failure syndromes (IBMFS) are a heterogenous group of disorders caused by germline mutations in related genes and characterized by bone marrow failure (BMF), disease specific organ involvement, and, in most cases, predisposition to malignancy. Their distinction from immune marrow failure can often be challenging, particularly when presentations occur in adulthood or are atypical. A combination of functional (disease specific assays) and genetic testing is optimal in assessing all new BMF patients for an inherited etiology. However, genetic testing is costly and may not be available worldwide due to resource constraints; in such cases, clinical history, standard laboratory testing, and the use of algorithms can guide diagnosis. Interpretation of genetic results can be challenging and must reflect assessment of pathogenicity, inheritance pattern, clinical phenotype, and specimen type used. Due to the progressive use of genomics, new IBMFS continue to be identified, widening the spectrum of these disorders.

Spectrum of clonal hematopoiesis in VEXAS syndrome.

Vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic (VEXAS) syndrome is caused by somatic mutations in UBA1 (UBA1mut) and characterized by heterogenous systemic autoinflammation and progressive hematologic manifestations, meeting criteria for myelodysplastic syndrome (MDS) and plasma cell dyscrasias. The landscape of myeloid-related gene mutations leading to typical clonal hematopoiesis (CH) in these patients is unknown. Retrospectively, we screened 80 patients with VEXAS for CH in their peripheral blood (PB) and correlated the findings with clinical outcomes in 77 of them. UBA1mut were most common at hot spot p.M41 (median variant allele frequency [VAF] = 75%). Typical CH mutations cooccurred with UBA1mut in 60% of patients, mostly in DNMT3A and TET2, and were not associated with inflammatory or hematologic manifestations. In prospective single-cell proteogenomic sequencing (scDNA), UBA1mut was the dominant clone, present mostly in branched clonal trajectories. Based on integrated bulk and scDNA analyses, clonality in VEXAS followed 2 major patterns: with either typical CH preceding UBA1mut selection in a clone (pattern 1) or occurring as an UBA1mut subclone or in independent clones (pattern 2). VAF in the PB differed markedly between DNMT3A and TET2 clones (median VAF of 25% vs 1%). DNMT3A and TET2 clones associated with hierarchies representing patterns 1 and 2, respectively. Overall survival for all patients was 60% at 10 years. Transfusion-dependent anemia, moderate thrombocytopenia, and typical CH mutations, each correlated with poor outcome. In VEXAS, UBA1mut cells are the primary cause of systemic inflammation and marrow failure, being a new molecularly defined somatic entity associated with MDS. VEXAS-associated MDS is distinct from classical MDS in its presentation and clinical course.

Differential diagnosis of bone marrow failure syndromes guided by machine learning.

The choice to postpone treatment while awaiting genetic testing can result in significant delay in definitive therapies in patients with severe pancytopenia. Conversely, the misdiagnosis of inherited bone marrow failure (BMF) can expose patients to ineffectual and expensive therapies, toxic transplant conditioning regimens, and inappropriate use of an affected family member as a stem cell donor. To predict the likelihood of patients having acquired or inherited BMF, we developed a 2-step data-driven machine-learning model using 25 clinical and laboratory variables typically recorded at the initial clinical encounter. For model development, patients were labeled as having acquired or inherited BMF depending on their genomic data. Data sets were unbiasedly clustered, and an ensemble model was trained with cases from the largest cluster of a training cohort (n = 359) and validated with an independent cohort (n = 127). Cluster A, the largest group, was mostly immune or inherited aplastic anemia, whereas cluster B comprised underrepresented BMF phenotypes and was not included in the next step of data modeling because of a small sample size. The ensemble cluster A-specific model was accurate (89%) to predict BMF etiology, correctly predicting inherited and likely immune BMF in 79% and 92% of cases, respectively. Our model represents a practical guide for BMF diagnosis and highlights the importance of clinical and laboratory variables in the initial evaluation, particularly telomere length. Our tool can be potentially used by general hematologists and health care providers not specialized in BMF, and in under-resourced centers, to prioritize patients for genetic testing or for expeditious treatment.

Effects of nandrolone decanoate on telomere length and clinical outcome in patients with telomeropathies: a prospective trial.

Androgens have been reported to elongate telomeres in retrospective and prospective trials with patients with telomeropathies, mainly with bone marrow failure. In our single-arm prospective clinical trial (clinicaltrials gov. Identifier: NCT02055456), 17 patients with short telomeres and/or germline pathogenic variants in telomere biology genes associated with at least one cytopenia and/or radiologic diagnosis of interstitial lung disease were treated with 5 mg/kg of intramuscular nandrolone decanoate every 15 days for 2 years. Ten of 13 evaluable patients (77%) showed telomere elongation at 12 months by flow-fluorescence in situ hybridization (average increase, 0.87 kb; 95% confidence interval: 0.20-1.55 kb; P=0.01). At 24 months, all ten evaluable patients showed telomere elongation (average increase, 0.49 kb; 95% confidence interval: 0.24-1.23 kb; P=0.18). Hematologic response was achieved in eight of 16 patients (50%) with marrow failure at 12 months, and in ten of 16 patients (63%) at 24 months. Seven patients had interstitial lung disease at baseline, and two and three had pulmonary response at 12 and 24 months, respectively. Two patients died due to pulmonary failure during treatment. In the remaining evaluable patients, the pulmonary function remained stable or improved, but showed consistent decline after cessation of treatment. Somatic mutations in myeloid neoplasm-related genes were present in a minority of patients and were mostly stable during drug treatment. The most common adverse events were elevations in liver function test levels in 88%, acne in 59%, and virilization in 59%. No adverse events grade ≥4 was observed. Our findings indicate that nandrolone decanoate elongates telomeres in patients with telomeropathies, which correlated with clinical improvement in some cases and tolerable adverse events.

Somatic mutations in VEXAS Syndrome and Erdheim-Chester disease: Inflammatory myeloid diseases.

Somatic mutations have been increasingly identified as etiologic for many hematologic and autoinflammatory disorders. VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome and Erdheim-Chester disease (ECD), a type of histiocytosis, can be classified as inflammatory myeloid diseases, characterized by systemic inflammation and multi-organ disease with predisposition to myeloid malignancies. VEXAS is a novel disease caused by UBA1 mutations that was first discovered using a genotype-driven approach (genotype was used to identify patients with undiagnosed inflammatory diseases). Since the initial description, many VEXAS cases have been reported and disease phenotype is expanding rapidly. In contrast, ECD was first characterized in the 1930s based on patients' phenotype, and only recently found to be caused by recurrent somatic mutations in the MAPK pathway (traditional phenotype-driven approach). The discovery of these mutations and development of target therapies have revolutionized the treatment of patients with histiocytosis, particularly ECD. Here we discuss the impact of causal and associated somatic mutations in VEXAS and ECD at both clinical and molecular levels.

Predictors of clonal evolution and myeloid neoplasia following immunosuppressive therapy in severe aplastic anemia.

Predictors, genetic characteristics, and long-term outcomes of patients with SAA who clonally evolved after immunosuppressive therapy (IST) were assessed. SAA patients were treated with IST from 1989-2020. Clonal evolution was categorized as "high-risk" (overt myeloid neoplasm [meeting WHO criteria for dysplasia, MPN or acute leukemia] or isolated chromosome-7 abnormality/complex karyotype without dysplasia or overt myeloid neoplasia) or "low-risk" (non-7 or non-complex chromosome abnormalities without morphological evidence of dysplasia or myeloid neoplasia). Univariate and multivariate analysis using Fine-Gray competing risk regression model determined predictors. Long-term outcomes included relapse, overall survival (OS) and hematopoietic stem cell transplant (HSCT). Somatic mutations in myeloid cancer genes were assessed in evolvers and in 407 patients 6 months after IST. Of 663 SAA patients, 95 developed clonal evolution. Pre-treatment age >48 years and ANC > 0.87 × 109/L were strong predictors of high-risk evolution. OS was 37% in high-risk clonal evolution by 5 years compared to 94% in low-risk. High-risk patients who underwent HSCT had improved OS. Eltrombopag did not increase high-risk evolution. Splicing factors and RUNX1 somatic variants were detected exclusively at high-risk evolution; DNMT3A, BCOR/L1 and ASXL1 were present in both. RUNX1, splicing factors and ASXL1 somatic mutations detected at 6 months after IST predicted high-risk evolution.

Translation of cytoplasmic UBA1 contributes to VEXAS syndrome pathogenesis.

Somatic mutations in UBA1 cause vacuoles, E1 ubiquitin-activating enzyme, X-linked, autoinflammatory somatic (VEXAS) syndrome, an adult-onset inflammatory disease with an overlap of hematologic manifestations. VEXAS syndrome is characterized by a high mortality rate and significant clinical heterogeneity. We sought to determine independent predictors of survival in VEXAS and to understand the mechanistic basis for these factors. We analyzed 83 patients with somatic pathogenic variants in UBA1 at p.Met41 (p.Met41Leu/Thr/Val), the start codon for translation of the cytoplasmic isoform of UBA1 (UBA1b). Patients with the p.Met41Val genotype were most likely to have an undifferentiated inflammatory syndrome. Multivariate analysis showed ear chondritis was associated with increased survival, whereas transfusion dependence and the p.Met41Val variant were independently associated with decreased survival. Using in vitro models and patient-derived cells, we demonstrate that p.Met41Val variant supports less UBA1b translation than either p.Met41Leu or p.Met41Thr, providing a molecular rationale for decreased survival. In addition, we show that these 3 canonical VEXAS variants produce more UBA1b than any of the 6 other possible single-nucleotide variants within this codon. Finally, we report a patient, clinically diagnosed with VEXAS syndrome, with 2 novel mutations in UBA1 occurring in cis on the same allele. One mutation (c.121 A>T; p.Met41Leu) caused severely reduced translation of UBA1b in a reporter assay, but coexpression with the second mutation (c.119 G>C; p.Gly40Ala) rescued UBA1b levels to those of canonical mutations. We conclude that regulation of residual UBA1b translation is fundamental to the pathogenesis of VEXAS syndrome and contributes to disease prognosis.

Corrigendum: Possible Involvement of Hsp90 in the Regulation of Telomere Length and Telomerase Activity During the Leishmania Amazonensis Developmental Cycle and Population Proliferation.

[This corrects the article DOI: 10.3389/fcell.2021.713415.].

Long-term outcomes in patients with severe aplastic anemia treated with immunosuppression and eltrombopag: a phase 2 study.

Patients with severe aplastic anemia (SAA) are either treated with bone marrow transplant (BMT) or immunosuppression (IST) depending on their age, comorbidities, and available donors. In 2017, our phase 2 trial reported improved hematologic responses with the addition of eltrombopag (EPAG) to standard IST for SAA when compared with a historical cohort treated with IST alone. However, the rates and characteristics of long-term complications, relapse, and clonal evolution, previously described in patients treated with IST alone, are not yet known with this new regimen, IST and EPAG. Patients were accrued from 2012 to 2020, with a total of 178 subjects included in this secondary endpoint analysis. With double the sample size and a much longer median follow-up (4 years) since the original publication in 2017, we report a cumulative relapse rate of 39% in responding patients who received cyclosporine (CSA) maintenance and clonal evolution of 15% in all treated patients at 4 years. Relapse occurred at distinct timepoints: after CSA dose reduction and EPAG discontinuation at 6 months, and after 2 years when CSA was discontinued. Most relapsed patients were retreated with therapeutic doses of CSA +/- EPAG, and two-thirds responded. Clonal evolution to a myeloid malignancy or chromosome 7 abnormality (high-risk) was noted in 5.7% of patients and conferred a poorer overall survival. Neither relapse nor high-risk evolution occurred at a higher rate than was observed in a historical comparator cohort, but the median time to both events was earlier in IST and EPAG treated patients. This trial was registered at www.clinicaltrials.gov as #NCT01623167.

Utility of plasma cell-free DNA for de novo detection and quantification of clonal hematopoiesis.

Although cell-free DNA (cfDNA) tests have emerged as a potential non-invasive alternative to bone marrow biopsies for monitoring clonal hematopoiesis in hematologic diseases, whether commercial cfDNA assays can be implemented for the detection and quantification of de novo clonal hematopoiesis in place of blood cells is uncertain. In this study, peripheral plasma cfDNA samples available from patients with aplastic anemia (n=25) or myelodysplastic syndromes (n=27) and a healthy cohort (n=107) were screened for somatic variants in genes related to hematologic malignancies using a Clinical Laboratory Improvement Amendments-certified panel. Results were further compared to DNA sequencing of matched blood cells. In reported results, 85% of healthy subjects, 36% of patients with aplastic anemia and 74% of patients with myelodysplastic syndromes were found to have somatic cfDNA variants, most frequently in DNMT3A, TET2, ASXL1 and SF3B1. However, concordance between cfDNA and blood cell findings was poor for the detection of clonal hematopoiesis when the allele frequency of the variants was <10%, which was mostly observed in the healthy and aplastic anemia cohorts but not in patients with myelodysplastic syndromes. After filtering data for potential artifacts due to low variant allele frequency and sequencing depth, the frequency of clonal hematopoiesis in cfDNA from healthy individuals and patients with aplastic anemia decreased to 52% and 20%, respectively. cfDNA and matched blood cells were not interchangeable for tracking changes in allele burdens as their agreement by Bland-Altman analysis was poor. A commercial cfDNA assay had good performance for de novo detection of clonal hematopoiesis in myelodysplastic syndromes, but showed no advantage over blood cells in diseases with low allele burdens or in healthy individuals.

Association between leukocyte telomere length and sex by quantile regression analysis.

INTRODUCTION: Telomere length (TL) is a biomarker of cellular proliferative history. In healthy individuals, leukocyte TL shortens with age and associates with the lifespan of men and women. However, most of studies had used linear regression models to address the association of the TL attrition, aging and sex. METHODS: We evaluated the association between the TL, aging and sex in a cohort of 180 healthy subjects by quantile regression. The TL of nucleated blood cells was measured by fluorescent in situ hypridization (flow-FISH) in a cohort of 89 men, 81 women, and 10 umbilical cord samples. The results were validated by quantitative polymerase chain reaction (qPCR) and compared to a linear regression analysis. RESULTS: By quantile regression, telomere dynamics slightly differed between sexes with aging: women had longer telomeres at birth and slower attrition rate than men until the sixth decade of life; after that, TL eroded faster and became shorter than that in men. These differences were not observed by linear regression analysis, as the overall telomere attrition rates in women and men were similar (42 pb per year, p < 0.0001 vs. 45 pb kb per year, p < 0.0001). Also, qPCR did not recapitulate flow-FISH findings, as the telomere dynamics by qPCR followed a linear model. CONCLUSION: The quantile regression analysis accurately reproduced a third-order polynomial TL attrition rate in both women and men, but it depended on the technique applied to measure TL. The Flow-FISH reproduced the expected telomere dynamics through life and, differently from the qPCR, was able to detect the subtle TL variations associated with sex and aging.

Possible Involvement of Hsp90 in the Regulation of Telomere Length and Telomerase Activity During the Leishmania amazonensis Developmental Cycle and Population Proliferation.

The Leishmania developmental cycle comprises three main life forms in two hosts, indicating that the parasite is continually challenged due to drastic environmental changes. The disruption of this cycle is critical for discovering new therapies to eradicate leishmaniasis, a neglected disease that affects millions worldwide. Telomeres, the physical ends of chromosomes, maintain genome stability and cell proliferation and are potential antiparasitic drug targets. Therefore, understanding how telomere length is regulated during parasite development is vital. Here, we show that telomeres form clusters spread in the nucleoplasm of the three parasite life forms. We also observed that amastigotes telomeres are shorter than metacyclic and procyclic promastigotes and that in parasites with continuous in vitro passages, telomere length increases over time. These observed differences in telomere length among parasite's life stages were not due to lack/inhibition of telomerase since enzyme activity was detected in all parasite life stages, although the catalysis was temperature-dependent. These data led us to test if, similar to other eukaryotes, parasite telomere length maintenance could be regulated by Hsp83, the ortholog of Hsp90 in trypanosomatids, and Leishmania (LHsp90). Parasites were then treated with the Hsp90 inhibitor 17AAG. The results showed that 17AAG disturbed parasite growth, induced accumulation into G2/M phases, and telomere shortening in a time-dependent manner. It has also inhibited procyclic promastigote's telomerase activity. Besides, LHsp90 interacts with the telomerase TERT component as shown by immunoprecipitation, strongly suggesting a new role for LHsp90 as a parasite telomerase component involved in controlling telomere length maintenance and parasite life span.

Hematopoiesis under telomere attrition at the single-cell resolution.

The molecular mechanisms that drive hematopoietic stem cell functional decline under conditions of telomere shortening are not completely understood. In light of recent advances in single-cell technologies, we sought to redefine the transcriptional and epigenetic landscape of mouse and human hematopoietic stem cells under telomere attrition, as induced by pathogenic germline variants in telomerase complex genes. Here, we show that telomere attrition maintains hematopoietic stem cells under persistent metabolic activation and differentiation towards the megakaryocytic lineage through the cell-intrinsic upregulation of the innate immune signaling response, which directly compromises hematopoietic stem cells' self-renewal capabilities and eventually leads to their exhaustion. Mechanistically, we demonstrate that targeting members of the Ifi20x/IFI16 family of cytosolic DNA sensors using the oligodeoxynucleotide A151, which comprises four repeats of the TTAGGG motif of the telomeric DNA, overcomes interferon signaling activation in telomere-dysfunctional hematopoietic stem cells and these cells' skewed differentiation towards the megakaryocytic lineage. This study challenges the historical hypothesis that telomere attrition limits the proliferative potential of hematopoietic stem cells by inducing apoptosis, autophagy, or senescence, and suggests that targeting IFI16 signaling axis might prevent hematopoietic stem cell functional decline in conditions affecting telomere maintenance.

Benign and malignant hematologic manifestations in patients with VEXAS syndrome due to somatic mutations in UBA1.

Somatic mutations in UBA1 involving hematopoietic stem and myeloid cells have been reported in patients with the newly defined VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome. Here, we report clinical hematologic manifestations and unique bone marrow (BM) features in 16 patients with VEXAS. All patients were male and had a history of severe autoinflammatory and rheumatologic manifestations and a somatic UBA1 mutation (p.Met41). Ten patients had hematologic disorders: myelodysplastic syndrome (MDS; 6 of 16), multiple myeloma (2 of 16), monoclonal gammopathy of undetermined significance (2 of 16), and monoclonal B-cell lymphocytosis (2 of 16), and a few of those patients had 2 co-existing clonal processes. Although macrocytic anemia (100%) and lymphopenia (80%) were prevalent in all patients with VEXAS, thrombocytopenia and neutropenia were more common in patients with progression to MDS. All BMs in VEXAS patients had prominent cytoplasmic vacuoles in myeloid and erythroid precursors. In addition, most BMs were hypercellular with myeloid hyperplasia, erythroid hypoplasia, and varying degrees of dysplasia. All patients diagnosed with MDS were lower risk (low blast count, very good to intermediate cytogenetics) according to standard prognostic scoring with no known progression to leukemia. In addition, 10 of 16 patients had thrombotic events, including venous thromboembolism and arterial stroke. Although VEXAS presents symptomatically as a rheumatologic disease, morbidity and mortality are associated with progression to hematologic disease. Given the increased risk of developing MDS and multiple myeloma, surveillance for disease progression is important.

Somatic mosaicism in inherited bone marrow failure syndromes.

Inherited bone marrow failure syndromes (IBMFS) are a heterogenous group of diseases caused by pathogenic germline variants in key pathways associated with haematopoiesis and genomic stability. Germline variants in IBMFS-related genes are known to reduce the fitness of hematopoietic stem and progenitor cells (HSPC), which has been hypothesized to drive clonal selection in these diseases. In many IBMFS, somatic mosaicism predominantly impacts cells by two distinct mechanisms, with contrasting effects. An acquired variation can improve cell fitness towards baseline levels, providing rescue of a deleterious phenotype. Alternatively, somatic mosaicism may result in a fitness advantage that results in malignant transformation. This review will describe these phenomena in IBMFS and delineate their relevance for diagnosis and clinical management. In addition, we will discuss which samples and methods can be used for detection of mosaicism according to clinical phenotype, type of mosaicism, and sample availability.

Eltrombopag added to immunosuppression for children with treatment-naïve severe aplastic anaemia.

Acquired severe aplastic anaemia (SAA) has an immune pathogenesis, and immunosuppressive therapy (IST) with anti-thymocyte globulin and cyclosporine is effective therapy. Eltrombopag (EPAG) added to standard IST was associated with higher overall and complete response rates in patients with treatment-naïve SAA compared to a historical IST cohort. We performed a paediatric subgroup analysis of this trial including all patients aged <18 years who received EPAG plus standard IST (n = 40 patients) compared to a historical cohort (n = 87) who received IST alone. Response, relapse, clonal evolution, event-free survival (EFS), and overall survival were assessed. There was no significant difference in either the overall response rate (ORR) or complete response rate at 6 months (ORR 70% in EPAG group, 72% in historical group, P = 0·78). Adults (≥18 years) had a significantly improved ORR of 82% with EPAG compared to 58% historically (P < 0·001). Younger children had lower response rates than did adolescents. The trend towards relapse was higher and EFS significantly lower in children who received EPAG compared to IST alone. Addition of EPAG added to standard IST did not improve outcomes in children with treatment-naïve SAA. EPAG in the paediatric population should not automatically be considered standard of care. Registration: clinicaltrials.gov (NCT01623167).

Eltrombopag monotherapy can improve hematopoiesis in patients with low to intermediate risk-1 myelodysplastic syndrome.

Myelodysplastic syndromes (MDS) are a group of clonal myeloid disorders characterized by cytopenia and a propensity to develop acute myeloid leukemia (AML). The management of lower-risk (LR) MDS with persistent cytopenias remains suboptimal. Eltrombopag (EPAG), a thrombopoietin receptor agonist, can improve platelet counts in LR-MDS and tri-lineage hematopoiesis in aplastic anemia (AA). We conducted a phase 2 dose modification study to investigate the safety and efficacy of EPAG in LR-MDS. EPAG dose was escalated from 50 mg/day, to a maximum of 150 mg/day over a period of 16 weeks. The primary efficacy endpoint was hematologic response at 16-20 weeks. Eleven of 25 (44%) patients responded; five and six patients had uni- or bi-lineage hematologic responses, respectively. The predictors of response were presence of a PNH clone, marrow hypocellularity, thrombocytopenia with or without other cytopenia, and elevated plasma thrombopoietin levels at study entry. The safety profile was consistent with previous EPAG studies in AA; no patients discontinued drug due to adverse events. Three patients developed reversible grade-3 liver toxicity and one patient had increased reticulin fibrosis. Ten patients discontinued EPAG after achieving a robust response (median time 16 months); four of them reinitiated EPAG due to declining counts, and all attained a second robust response. Six patients had disease progression not associated with expansion of mutated clones and no patient progressed to AML on study. In conclusion, EPAG was well-tolerated and effective in restoring hematopoiesis in patients with low to intermediate-1 risk MDS. This study was registered at clinicaltrials.gov as #NCT00932156.

Somatic Mutations in UBA1 and Severe Adult-Onset Autoinflammatory Disease.

BACKGROUND: Adult-onset inflammatory syndromes often manifest with overlapping clinical features. Variants in ubiquitin-related genes, previously implicated in autoinflammatory disease, may define new disorders. METHODS: We analyzed peripheral-blood exome sequence data independent of clinical phenotype and inheritance pattern to identify deleterious mutations in ubiquitin-related genes. Sanger sequencing, immunoblotting, immunohistochemical testing, flow cytometry, and transcriptome and cytokine profiling were performed. CRISPR-Cas9-edited zebrafish were used as an in vivo model to assess gene function. RESULTS: We identified 25 men with somatic mutations affecting methionine-41 (p.Met41) in UBA1, the major E1 enzyme that initiates ubiquitylation. (The gene UBA1 lies on the X chromosome.) In such patients, an often fatal, treatment-refractory inflammatory syndrome develops in late adulthood, with fevers, cytopenias, characteristic vacuoles in myeloid and erythroid precursor cells, dysplastic bone marrow, neutrophilic cutaneous and pulmonary inflammation, chondritis, and vasculitis. Most of these 25 patients met clinical criteria for an inflammatory syndrome (relapsing polychondritis, Sweet's syndrome, polyarteritis nodosa, or giant-cell arteritis) or a hematologic condition (myelodysplastic syndrome or multiple myeloma) or both. Mutations were found in more than half the hematopoietic stem cells, including peripheral-blood myeloid cells but not lymphocytes or fibroblasts. Mutations affecting p.Met41 resulted in loss of the canonical cytoplasmic isoform of UBA1 and in expression of a novel, catalytically impaired isoform initiated at p.Met67. Mutant peripheral-blood cells showed decreased ubiquitylation and activated innate immune pathways. Knockout of the cytoplasmic UBA1 isoform homologue in zebrafish caused systemic inflammation. CONCLUSIONS: Using a genotype-driven approach, we identified a disorder that connects seemingly unrelated adult-onset inflammatory syndromes. We named this disorder the VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome. (Funded by the NIH Intramural Research Programs and the EU Horizon 2020 Research and Innovation Program.).