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We previously reported that granulocytic myeloid-derived suppressor cells (G-MDSCs) suppressed T-cell activation and attenuated bone marrow failure (BMF) in a minor histocompatibility (minor-H) antigen mismatched murine aplastic anemia (AA) model. In the current study, we tested the hypothesis that exosomes, a subset of extracellular vesicles, are responsible at least partially for G-MDSCs' therapeutic efficacy. Indeed, exosomes isolated from GMDSCs (G-MDSC-exos) suppressed CD4+ and CD8+ T-cell proliferation in vitro and mildly attenuated immune BMF in the minor-H mismatched AA model. G-MDSC-exos treatment significantly increased red blood cells, hemoglobin, and total bone marrow (BM) cells, and moderately reduced BM CD8+ T cells. G-MDSC-exos' effects were associated with upregulations in an array of lymphocyte-suppression-related miRNAs such as hsa-miR-142-5p, miR-19a-3p, and miR-19b-3p in both BM CD4+ and CD8+ T cells. We concluded that G-MDSC-exos attenuate immune BMF via modulating the delivery of immunosuppressive miRNAs into activated T lymphocytes. [ABSTRACT FROM AUTHOR]

COVID-19 did not result in SAA relapse but these results cannot be generalised to patients who may have more severe COVID-19 manifestations due to other underlying comorbidities.13 The timing of relapse post viral infection is not established therefore longer follow-up may lead to better knowledge. The coronavirus disease 2019 (COVID-19) pandemic has caused concern among physicians caring for immunosuppressed patients. No patients required COVID-19 infection-directed therapies or had interruption of ongoing immunosuppression. In patients with concurrent SAA and COVID-19 infection, continuation or initiation of CsA and/or EPAG during SARS-CoV-2 infection does not appear to complicate infectious course. [Extracted from the article]

Summary: 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). [ABSTRACT FROM AUTHOR]

The article provides information on the diagnosis and treatment of aplastic anemia. Damage to the bone marrow mostly takes place as a result of chemotherapy or radiation therapy. It has been observed that genetic lesions reduce the capacity for hematopoietic stem cells to repair any damage experienced by DNA.

Paroxysmal nocturnal hemoglobinuria ( PNH) is an acquired bone marrow disorder caused by expansion of a clone of hematopoietic cells lacking glycosylphosphatidylinositol ( GPI)-anchored membrane proteins. Multiple lines of evidence suggest immune attack on normal hematopoietic stem cells provides a selective growth advantage to PNH clones. Recently, frequent loss of HLA alleles associated with copy number-neutral loss of heterozygosity in chromosome 6p ( CN-6p LOH) in aplastic anemia ( AA) patients was reported, suggesting that AA hematopoiesis 'escaped' from immune attack by loss of HLA alleles. We report here the first case of CN-6p LOH in a Japanese PNH patient only in GPI-anchored protein positive (59%) granulocytes, but not in GPI-anchored protein negative (41%) granulocytes. CN-6p LOH resulted in loss of the alleles A*02:06- DRB1*15:01- DQB1*06:02, which have been reported to be dominant in Japanese PNH patients. Our patient had maintained nearly normal blood count for several years. Our case supports the hypothesis that a hostile immune environment drives selection of resistant hematopoietic cell clones and indicates that clonal evolution may occur also in normal phenotype (non- PNH) cells in some cases. [ABSTRACT FROM AUTHOR]

Survival in severe aplastic anemia (SAA) has markedly improved in the past 4 decades because of advances in hema-topoietic stem cell transplantation, immu-nosuppressive biologies and drugs, and supportive care. However, management of SAA patients remains challenging, both acutely in addressing the immediate consequences of pancytopenia and in the long term because of the disease's natu-ral history and the consequences of therapy. Recent insights into pathophysi-ology have practical implications. We re-view key aspects of differential diagno-sis, considerations in the choice of first-and second-line therapies, and the management of patients after immunosup-pression, based on both a critical review of the recent literature and our large per-sonal and research protocol experience of bone marrow failure in the Hematology Branch of the National Heart, Lung, and Blood Institute. [ABSTRACT FROM AUTHOR]

The telomeropathies are a newly described group of human diseases based on the genetics and molecular biology of the telomeres, the ends of chromosomes. Telomeres are repeated hexanucleotides and their associated proteins; the protect chromosomes from recognition as damaged DNA, and their inevitable gradual loss with DNA replication is harmless as they are noncoding. However, when telomeres become critically short in a cell, senescence, apoptosis, or, rarely malignant transformation results. In individuals with mutations in genes involved in telomere repair, especially the enzymatic telomerase complex, telomere attrition is accelerated. Severe deficiencies result in dyskeratosis congenita, a congenital aplastic anemia with associated mucocutaneous abnormalities. Mutations in TERT, the catalytic component, and TERC, the RNA template, can behave as risk factors for the development of bone marrow failure, pulmonary fibrosis, and hepatic cirrhosis. Both penetrance and organ specificity are variable and not well understood. Chromosome instability is a result of critical shortening of telomeres and cancer. For example, short telomeres are the major prognostic risk factor for clonal evolution to myelodysplasia and acute leukemia. Practically, hematologists need to recognize the multisystem presentation of telomere disease, implications for outcomes, and options for therapy. [ABSTRACT FROM AUTHOR]

Although recent advances in therapy offer the promise for improving survival in patients with severe aplastic anemia (SAA), the small size of the patient population, lack of a mechanism in North America for longitudinal follow-up of patients, and inadequate cooperation among hematologists, scientists, and transplant physicians remain obstacles to conducting large studies that would advance the field. To address this issue, the Blood and Marrow Transplant Clinical Trials Network (BMT CTN) convened a group of international experts in March 2010 to define the most important questions in the basic science, immunosuppressive therapy (IST), and bone marrow transplantation (BMT) of SAA and propose initiatives to facilitate clinical and biologic research. Key conclusions of the working group were: (1) new patients should obtain accurate, expert diagnosis and early identification of biologic risk; (2) a population-based SAA outcomes registry should be established in North America to collect data on patients longitudinally from diagnosis through and after treatment; (3) a repository of biologic samples linked to the clinical data in the outcomes registry should be developed; (4) innovative approaches to unrelated donor BMT that decrease graft-versus-host disease are needed; and (5) alternative donor transplantation approaches for patients lacking HLA-matched unrelated donors must be improved. A partnership of BMT, IST, and basic science researchers will develop initiatives and partner with advocacy and funding organizations to address these challenges. Collaboration with similar study groups in Europe and Asia will be pursued. [Copyright &y& Elsevier]

The article discusses a study which investigated the association between telomere length and clinical outcomes in patients with severe aplastic anemia. The study included 183 patients with severe aplastic anemia who were treated at the U.S. National Institutes of Health from 2000 to 2008. Findings showed no relationship between hematologic response and telomere length. Study authors concluded that the length of telomere is not related to therapeutic response but it is linked with the risk of relapse, clonal evolution and overall survival.

Horse anti-thymocyte globulin (h-ATG) and ciclosporin are the initial therapy for most patients with severe aplastic anaemia (SAA), but there is no practical and reliable method to predict response to this treatment. To determine whether pretreatment blood counts discriminate patients with SAA who have a higher likelihood of haematological response at 6 months to immunosuppressive therapy (IST), we conducted a single institution retrospective analysis on 316 SAA patients treated with h-ATG-based IST from 1989 to 2005. In multivariate analysis, younger age, higher baseline absolute reticulocyte count (ARC), and absolute lymphocyte count (ALC) were highly predictive of response at 6 months. Patients with baseline ARC ≥ 25 × 109/l and ALC ≥ 1 × 109/l had a much greater probability of response at 6 months following IST compared to those with lower ARC and ALC (83% vs. 41%, respectively; P < 0·001). This higher likelihood of response translated to greater rate of 5-year survival in patients in the high ARC/ALC group (92%) compared to those with a low ARC/ALC (53%). In the era of IST, the baseline ARC and ALC together serve as a simple predictor of response following IST, which should guide in risk stratification among patients with SAA. [ABSTRACT FROM AUTHOR]

In aplastic anaemia, T cells have a central role in the pathophysiology of bone marrow destruction. This study showed that T cells from patients with aplastic anaemia expressed decreased T-cell receptor (TCR) ζ-chain protein and mRNA levels compared to healthy controls. Patients with decreased TCR ζ-chain showed an abnormal response in intracellular calcium following stimulation through the TCR. We also observed an altered pattern of the transcription factors CREM α and Elf-1 that are implicated in ζ-chain transcription. We concluded that TCR ζ-chain expression was decreased in the majority of patients with aplastic anaemia, regardless of disease activity or treatment status. [ABSTRACT FROM AUTHOR]

BACKGROUND: Most patients with severe aplastic anemia (sAA) respond to immunosuppression, but a significant number relapse or develop clonal abnormalities such as paroxysmal nocturnal hemoglobinuria, myelodysplasia, or leukemia. In principle, patients without matched sibling donors and older patients might benefit from transplantation of autologous hematopoietic peripheral blood progenitor cells (PBPCs) obtained during remission. Even patients who have clinically recovered from aplastic anemia have diminished hematopoietic progenitor cells, so the practicability of PBPC mobilization in these individuals is unknown. STUDY DESIGN AND METHODS: The feasibility of PBPC mobilization in nine patients with a history of sAA was evaluated. Granulocyte–colony-stimulating factor (10 µg/kg) was administered subcutaneously for 5 days and followed by a 12-L leukapheresis procedure. RESULTS: Only two of the nine patients had sufficient mobilization of CD34 cells to merit collection; in these cases sufficient CD34 cells were obtained for autologous transplantation should the need arise. CONCLUSION: PBPC collection is feasible only in a fraction of recovered AA patients. [ABSTRACT FROM AUTHOR]

Granulocyte colony-stimulating factor (GCSF) administration has been linked to the development of monosomy 7 in severe congenital neutropenia and aplastic anemia. We assessed the effect of pharmacologic doses of GCSF on monosomy 7 cells to determine whether this chromosomal abnormality developed de novo or arose as a result of favored expansion of a preexisting clone. Fluorescence in situ hybridization (FISH) of chromosome 7 was used to identify small populations of aneuploid cells. When bone marrow mononuclear cells from patients with monosomy 7 were cultured with 400 ng/ml GCSF, all samples showed significant increases in the proportion of monosomy 7 cells. In contrast, bone marrow from karyotypically normal aplastic anemia, myelodys-plastic syndrome, or healthy individuals did not show an increase in monosomy 7 cells in culture. In bone marrow CD34 cells of patients with myelodysplastic syndrome and monosomy 7, GCSF receptor (GCSFR) protein was increased. Although no mutation was found in genomic GCSFR DNA, CD34 cells showed increased expression of the GCSFR class IV mRNA isoform, which is defective in signaling cellular differentiation. GCSFR signal transduction via the Jak/Stat system was abnormal in monosomy 7 CD34 cells, with increased phosphorylated signal transducer and activation of transcription protein, STAT1-P, and increased STAT5-P relative to STAT3-P. Our results suggest that pharmacologic doses of GCSF increase the proportion of preexisting monosomy 7 cells. The abnormal response of monosomy 7 cells to GCSF would be explained by the expansion of undifferentiated monosomy 7 clones expressing the class IV GCSFR, which is defective in signaling cell maturation. [ABSTRACT FROM AUTHOR]

Severe aplastic anaemia (SAA) can be successfully treated with immunosuppressive therapies or haematopoietic stem cell transplantation (HSCT). Response rates with horse anti-thymocyte globulin (h-ATG) plus ciclosporin (CsA) are about 60–70%, and robust responders have an excellent long-term survival. We introduced a third immunosuppressive agent to standard h-ATG/CsA, mycophenolate mofetil (MMF), in an attempt to improve the response rate and survival, and to decrease the relapse rate and clonal evolution to myelodysplasia. A total of 104 consecutive patients with SAA were treated with h-ATG/CsA/MMF between May 1999 and June 2003 at the National Institutes of Health Clinical Center. The overall response rate at 6 months was 62%, with 24 (37% of responders) patients relapsing at a median of 389 d from ATG. Nine patients showed evidence of clonal evolution following ATG. After a median follow up of 42 months, the median survival among responders was not reached and among non-responders was 58 months. Over half of the relapses occurred during MMF administration. Despite a strong theoretical rationale for its use, MMF did not result in the improvement of response or relapse rates when compared with historical standard h-ATG/CsA. [ABSTRACT FROM AUTHOR]

The management of patients with severe aplastic anaemia (SAA) who do not have a matched sibling donor and fail a course of horse anti-thymocyte globulin (h-ATG)/ciclosporin (CsA) is uncertain. Repeated courses of ATG-based immunosuppression are often employed; in children and increasingly in adults, alternative donor haematopoietic stem cell transplantation is an option. We analysed the success rate of retreatment with rabbit ATG (r-ATG)/CsA in 43 patients treated at our institution in the last 5 years; 22 were refractory (20 adults; two children) to h-ATG/CsA-based regimens and 21 (17 adults; four children) had relapsed after h-ATG/CsA-based regimens. The overall response rate was 30% in patients who were refractory to h-ATG and 65% in patients who had relapsed following h-ATG. The 1000-d survival in patients who responded to r-ATG was 90% compared with 65% in non-responders. Six patients developed a clonal haematological disorder; two were responders, two were non-responders and in two the evolution occurred before the response could be assessed at 3 months following r-ATG. Thirteen patients died; three were responders, six were non-responders and four patients died prior to 3 months when response was assessed. In our study, the response rate in refractory patients was inferior to what has been previously reported. [ABSTRACT FROM AUTHOR]

Allogeneic haematopoietic cell transplantation (HCT) can cure a variety of non-malignant haematological disorders. Although transplant outcomes for selected patients with severe aplastic anaemia (SAA) and paroxysmal nocturnal haemoglobinuria (PNH) have improved, older age, allo-immunisation from transfusions, prior immunosuppressive therapy and a prolonged time from diagnosis to transplantation are associated with worse outcome. Because of its potent immunosuppressive effects, we investigated a fludarabine-based non-myeloablative conditioning regimen in patients with transfusion-dependent non-malignant haematological disorders at increased risk for graft rejection with conventional transplant conditioning. Twenty-six patients with transfusion dependent/anti-thymocyte globulin (ATG)-refractory SAA, PNH or pure red cell aplasia underwent HCT from a human leucocyte antigen (HLA)-compatible relative. Transplant conditioning consisted of cyclophosphamide (120 mg/kg) and fludarabine (125 mg/m2) with or without ATG. Ciclosporine, alone or combined with mycophenolate mofetil or methotrexate, was used as graft- versus-host disease (GVHD) prophylaxis. All patients achieved durable engraftment and transfusion-independence. Twenty-four of 26 patients are alive at a median of 21 months following transplantation. Although a high cumulative incidence of acute (65% grades II–IV, 54% grades III–IV) and chronic GVHD (56%) was observed, only one patient died from transplant-related causes (cumulative incidence 7%). These data show that HCT following fludarabine-based non-myeloablative conditioning results in durable engraftment and excellent survival in SAA and PNH patients at high risk for graft rejection. [ABSTRACT FROM AUTHOR]

We analysed endothelial cell membrane microparticles (ECMP) in the peripheral blood of patients with paroxysmal nocturnal haemoglobinuria (PNH) ( n = 9), aplastic anaemia (AA) ( n = 10), sickle cell disease (SCD) ( n = 8), and healthy donors (HD) ( n = 11). There was no clinically manifested thrombosis in the PNH or AA group, except one cured thrombophlebitis (PNH), while all SCD patients had a history of vaso-occlusive crises. We used three-colour flow cytometry with blood cell-specific antibodies and antibodies to endothelial antigens CD105 and CD144. Phosphatidylserine-positive microparticles were detected using the annexin V-binding (AVB) assay. The population of CD105+AVB+ ECMP was significantly ( P < 0·05) higher in SCD (median: 0·568 × 109/l; 25–75th percentile range: 0·351–0·976 × 109/l) and PNH (0·401 × 109/l ; 0·19–0·441 × 109/l) patients when compared with AA (0·122 × 109/l; 0·061–0·172 × 109/l) or HD (0·180 × 109/l; 0·137–0·217 × 109/l) group. Even more pronounced differences were observed in ECMP exhibiting a marker of inflammatory stimulation CD54 (CD105+CD54+). Similarly, ECMP that exhibited endothelial specific and proteolysis-sensitive antigen CD144 were increased in SCD and PNH, but not in AA. Elevated CD54+ ECMP may reflect the inflammatory status of endothelial cells in SCD and PNH, while CD144+ ECMP could indicate continuous endothelial stimulation and/or injury. Analysis of circulating ECMP appears promising to provide useful information on the status of the vascular endothelium in PNH and SCD. [ABSTRACT FROM AUTHOR]

Studies the prevalence of parvovirus B19 in liver-tissue samples from patients with fulminant hepatitis (FH) and hepatitis-associated aplastic anemia (HAA) and from control subjects. Increase in the prevalence of variant erythrovirus sequences in livers of patients with hepatitis B or C infections; Role of parvovirus B19 as possible etiological agent of FH or HAA.

Context: In most patients, aplastic anemia results from T-cell–mediated immune destruction of bone marrow. Aplastic anemia can be effectively treated by stem cell transplantation or immunosuppression. Objective: To assess long-term outcomes after immunosuppressive therapy. Design, Setting, and Patients: Cohort of 122 patients (31 were ≤18 years and 91 were >18 years) with severe aplastic anemia, as determined by bone marrow cellularity and blood cell count criteria, were enrolled in a single-arm interventional research protocol from 1991 to 1998 at a federal government research hospital. Interventions: A dose of 40 mg/kg per day of antithymocyte globulin administered for 4 days, 10 to 12 mg/kg per day of cyclosporine for 6 months (adjusted for blood levels), and a short course of corticosteroids (1 mg/d of methylprednisolone for about 2 weeks). Main Outcome Measures: Survival, improvement of pancytopenia and transfusion-independence, relapse, and evolution to other hematologic diseases. Results: Response rates were 60% at 3 months after initiation of treatment, 61% at 6 months, and 58% at 1 year. The blood cell counts of patients who responded no longer satisfied severity criteria and they were transfusion-independent. Overall actuarial survival at 7 years was 55%. Survival was associated with early satisfaction of response criteria (86% vs 40% at 5 years; P<.001) and by blood counts at 3 months (reticulocyte count or platelet count of >50 × 10[sup 3]/µL predicted survival at 5 years of 90% [64/71] vs 42% [12/34] for patients with less robust recovery [P<.001 by log-rank test]). There were no deaths among responders more than 3 years after treatment. Relapse was common, but severe pancytopenia usually did not recur. Relapse did not influence survival. Thirteen patients showed evolution to other hematologic diseases, including monosomy 7. Conclusions: Approximately half of patients with severe aplastic anemia treated with antithymocyte... [ABSTRACT FROM AUTHOR]

Aplastic anemia (AA) remains an elusive disease. Its pathophysiology is not only fascinating by the seemingly simple findings of cytopenia and marrow hypoplasia, but may also contain key information to the understanding of other fundamental processes such as stem cell regeneration, evolution, and immune control of clonal diseases. Although measurements of blood counts provide an objective tool to assess the disease activity and response to the therapy, immune pathophysiology of AA, as inferred from the successes of immunosuppression, provides only few other clinical clues. Similarly, the current laboratory evidence remains mostly indirect. In spite of the recognition of immune pathways of hematopoietic inhibition and apoptosis in AA, the fundamental question about the nature of the antigen(s) inciting or maintaining the pathologic immune response that ultimately leads to bone marrow failure, remains open. However, recognition of the immune targets may aid in understanding not only the pathogenesis but also many of clinical associations and the late squelae of AA. For example, abnormal cells in AA and myelodysplastic syndrome (MDS) MDS may harbor inciting antigens but the immune response lacks selectivity. Clonal selection pressure may be a result of this process or alternatively, emergence of tolerance could lead to the establishment of abnormal hematopoiesis. Clonal proliferation of large granular lymphocytosis could represent an example of an exaggerated response to an immunodominant hematopoietic antigen. In addition to the traditional functional or phenotypic analysis, pathologic immune response in AA can be studied on molecular level by identifying and quantitating T cell clones based on the presence of unique variable B-chain CDR3 sequences. Detection of clonal expansion is based on the observation that in infections and autoimmune conditions, the presence of antigenic drive will lead to the expansion and overrepresentation of T cell clones recognizing this antigen. However, simple analysis of clonal representation is not sufficient to resolve the complex nature of the immune repertoire in the context of genetic and clinical heterogeneity. Therefore, we analyzed VB and CDR3 repertoire in CD4 and CD8 cells, activated or effector cell subsets. To distinguish truly expanded and likely immunodominant clones, we first studied VB distribution and cloned CDR3 sequences from expanded VB families. Identified clonotypic sequences can be used to design molecular tests to quantitate the strength of pathologic immune response. Clonotype sharing has been confirmed in patients with similar clinical features indicating presence of common antigens. In addition, quantitative analysis showed correlation with the therapy response. Persistence and patterns of clonotypes may be helpful in the classification of immune-mediated marrow failure based on the immune characteristics and will allow inferences into the inciting pathways. [ABSTRACT FROM AUTHOR]

In aplastic anemia, hematopoiesis fails: Blood cell counts are extremely low, and the bone marrow appears empty. The pathophysiology of aplastic anemia is now believed to be immune-mediated, with active destruction of blood-forming cells by lymphocytes. The aberrant immune response may be triggered by environmental exposures, such as to chemicals and drugs or viral infections and, perhaps, endogenous antigens generated by genetically altered bone marrow cells. In patients with post-hepatitis aplastic anemia, antibodies to the known hepatitis viruses are absent; the unknown infectious agent may be more common in developing countries, where aplastic anemia occurs more frequently than it does in the West. The syndrome paroxysmal nocturnal hemoglobinuria (PNH) is intimately related to aplastic anemia because many patients with bone marrow failure have an increased population of abnormal cells. In PNH, an entire class of proteins is not displayed on the cell surface because of an acquired X-chromosome gene mutation. The PNH cells may have a selective advantage in resisting immune attack. In contrast, the disease myelodysplasia can be confused with aplasia and can also evolve from aplastic anemia. The occurrence of cytogenetic abnormalities in patients years after presentation implies that genomic instability is a feature of this immune-mediated disease. Aplastic anemia can be effectively treated by stem-cell transplantation or immunosuppressive therapy. Transplantation is curative but is best used for younger patients who have histocompatible sibling donors. Antithymocyte globulin and cyclosporine restore hematopoiesis in approximately two thirds of patients. However, recovery of blood cell count is often incomplete, recurrent pancytopenia requires retreatment, and some patients develop late complications (especially myelodysplasia). [ABSTRACT FROM AUTHOR]

Historically, diverse clinical associations of aplastic anemia with benzene, radiation, medical drug exposure, pregnancy, viral infections and other hematologic diseases have suggested a complex etiology. However, the modern observation that the majority of patients recover with immunosuppressive drug treatment has implicated an immune pathophysiology. The abnormal immune response has been characterized as TH1/TC1, with measurable cytotoxic T-cell activation and excessive production of interferon-γ, tumor necrosis factor and interleukin-2. The result of this aberrant immune response is to induce programmed cell death in hematopoietic target cells through the Fas pathway, resulting in the destruction of a very high proportion of early progenitor and probably also stem cells. While the sequence and character of initiating events have remained elusive, a role for exogenous antigens may be inferred from preceding hepatitis or a convincing history of suspect pharmaceutic drug use; endogenous antigens have been inferred from the strong association of bone marrow hypocellularity with myelodysplasia and deficient marrow function with paroxysmal nocturnal hemoglobinuria. In addition to exposure, host susceptibility factors must influence the aberrant immune response and a few histocompatibility antigens have been linked to aplastic anemia. Early interruption of the process of immune-mediated hematopoietic cell destruction, as well as preservation of residual marrow cells from continuing subclinical destruction by T cells, are the goals of advanced approaches to immunosuppressive therapy. [ABSTRACT FROM AUTHOR]

Summary Uncertainty remains regarding the safety and tolerability of immunosuppressive therapy (IST) with anti‐thymocyte globulin (ATG) and cyclosporine (CSA) in older patients. We retrospectively analysed two prospective clinical trials of IST in treatment‐naïve severe aplastic anaemia (SAA) to assess safety in older compared to younger patients. Patients ≥18 years of age who had received IST with ATG and CSA +/− eltrombopag (EPAG) were included. Pre‐treatment baseline characteristics and co‐morbidities were assessed as predictors of therapy‐related complications in younger (<60 years) versus older (≥60 years) patients. Out of 245 eligible patients, 54 were older and 191 were younger. Older patients had a similar frequency of SAEs, ICU admissions and hospital length of stay compared to younger patients. Older patients had a higher frequency of cardiac events related to IST, but none resulted in death. Older patients had worse long‐term overall survival, and more relapse and clonal evolution post‐IST. However, older patients who responded to IST had a similar survival at a median follow‐up to younger patients. Disease‐related factors and limited therapeutic options in refractory disease likely contribute to poorer outcomes in older patients, not complications of upfront IST. Therefore, IST should be considered first‐line therapy for most older SAA patients. [ABSTRACT FROM AUTHOR]

Distinguishing constitutional from immune bone marrow failure (BMF) has important clinical implications. However, the diagnosis is not always straightforward, and immune aplastic anemia, the commonest BMF, is a diagnosis of exclusion. In this review, we discuss a general approach to the evaluation of BMF, focusing on clinical presentations particular to immune and various constitutional disorders as well as the interpretation of bone marrow histology, flow cytometry, and karyotyping. Additionally, we examine the role of specialized testing in both immune and inherited BMF, and discuss genetic testing, both its role in patient evaluation and interpretation of results. [ABSTRACT FROM AUTHOR]

Discusses acquired aplastic anemia. Case study of a man with profound hypocellularity and virtual absence of hematopoietic cells in his bone marrow. His condition two and half years after the onset of the bone marrow failure; History of aplastic anemia; Pathophysiology; Etiologies; Definitive therapies; Supportive care.

Presents an overview of aplastic anemia. Clinical presentation, pathology and natural history; Epidemiology; Causes; Pathophysiology; Treatment.

Focuses on flaviviruses and bone marrow failure suffered by patients with aplastic anemia. Viruses that infect bone marrow cells; Epidemiology of aplastic anemia; Effect of acute flavivirus infections on the bone marrow.

Provides information on a study that determined gamma-interferon gene expression in the bone marrow of patients with aplastic anemia and controls. Methodology of the study; Results and discussion on the study.

Objectives. A population-based case-control study was conducted to elucidate the incidence and etiology of aplastic anemia in Thailand. Methods. Case patients and hospital control patients were enrolled in three regions from 1989 to 1994; data were collected by interview. Results. Forty-six percent of 81 case patients and 19% of 295 control patients from Khonkaen were grain farmers (estimated relative risk [RR] = 2.7. 95% confidence interval [CI] = 1.4, 5.2). Sixteen percent of case patients and 6% of control patients used agricultural pesticides (estimated RR = 2.7, 95% CI = 1.1, 6.6). The association with grain farming remained among those not exposed to pesticides. In Songkla, 16% of 43 case patients and 2% of 181 control patients were grain farmers (crude RR estimate = 11, 95% CI = 3.4,35). Conclusions. The relation of aplastic anemia to grain fanning may partly explain the high incidence of aplastic anemia in Thailand. [ABSTRACT FROM AUTHOR]

Study Objective: To determine whether Epstein-Barr virus is present in the bone marrow of patients with aplastic anemia.Design: Assay of fresh and fixed bone marrow specimens for Epstein-Barr virus using immunofluorescence for nuclear antigen, Southern analysis with an Epstein-Barr virus specific probe, and in-situ hybridization.Setting: Governmental medical referral center.Patients: Five patients were studied prospectively: three who previously had infectious mononucleosis, one with a recent viral pneumonitis, and one who was asymptomatic. Stored DNA samples from other patients with aplastic anemia were also screened.Measurements and Main Results: Epstein-Barr virus DNA and protein were detected in the bone marrow of 5 patients studied prospectively and in 1 of 40 patients studied retrospectively. As estimated by in-situ hybridization, about 3% to 5% of marrow cells were infected with virus in those patients who had not received acyclovir. In contrast, Epstein-Barr virus DNA was not detected in peripheral blood DNA of these patients, nor were Epstein-Barr virus proteins or DNA found in the bone marrow of normal donors, patients with other hematologic diseases, or in 1 patient with acute infectious mononucleosis. Analysis of DNA fragments by hybridization with Epstein-Barr virus probes showed a pattern dissimilar to the type of Epstein-Barr virus usually associated with infectious mononucleosis.Conclusions: Aplastic anemia may be associated with Epstein-Barr virus more commonly than suspected by history. Localization of the virus in the bone marrow supports a causative role for Epstein-Barr virus in bone marrow failure. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented related to a survey conducted on the use of irradiated blood products by patients with aplastic anaemia taking immunosuppressive therapy.

A response from the authors of the report on somatic mutations and clonal hematopoiesis in aplastic anemia published in July 2 issue is presented.

• Ruxolitinib promotes expansion of Tregs in murine bone marrow failure (BMF). • GITR is a surrogate marker for FoxP3 in ruxolitinib-treated BMF mice. • Ruxolitinib suppresses Tregs in normal and sublethal irradiation conditions. • The effects of ruxolitinib on Tregs are immune-context dependent. Aplastic anemia is a bone marrow failure (BMF) disorder characterized by pancytopenia and hypocellular marrow from an immune-mediated etiology. Regulatory T cells (Tregs) prevent autoimmunity by suppressing autoreactive T cells. We recently demonstrated the efficacy of ruxolitinib (RUX), a JAK 1/2 inhibitor, in attenuating murine BMF. Herein, we investigated the changes of Tregs in the context of RUX treatment for murine BMF. Tregs are conventionally identified by surface expression of CD4 and CD25, in addition to intracellular transcription factor FoxP3. RUX promoted the expansion of Tregs in BMF mice defined by increased expression of FoxP3 in CD4 T cells but suppressed expression of activation marker CD25 in CD4 and CD8 T cells. In this context, CD25 is no longer a reliable surface marker for Tregs. We observed strong co-expression of FoxP3 with surface marker GITR instead of CD25 in RUX-treated BMF mice. Fluorescence-activated cell sorting (FACS)-sorted CD4+GITRhi cells showed high FoxP3 expression and intact suppressive function in vitro, suggesting GITR to be a surrogate marker for Tregs. In contrast to its expansive effect on Tregs in BMF, RUX suppressed Tregs in normal and sublethal irradiation conditions, indicating that the effects of RUX on Tregs are immune-context dependent. [ABSTRACT FROM AUTHOR]

An immune basis for most patients with aplastic anemia (AA) provides a rationale for immunosuppressive therapy (IST), using antithmyocyte globulin and cyclosporine as one therapeutic modality; hematologic response is observed in up to 75% of patients. Recent advances in understanding the pathogenesis of AA have identified defective telomere maintenance as an important explanation for the onset of marrow failure, relapse and clonal evolution after IST, in some patients with AA. The finding of inherited mutations in the telomerase gene complex in patients with apparent acquired AA has important implications for clinical management. Hematopoietic stem cell transplantation (HSCT) for acquired AA, whether from an HLA identical sibling or an unrelated donor, provides an excellent chance of long term cure. Current issues with HSCT include graft rejection, chronic GVHD and poor outcome in older patients. The lack of a suitable bone marrow donor for all patients who need a transplant, illustrates the need for novel transplant procedures, such as cord blood transplantation. [Copyright &y& Elsevier]

Suggests that in most patients with acquired aplastic anemia, bone marrow failure results from immunologically mediated, tissue specific organ destruction. Hematopoiesis in bone marrow failure; Medical therapy and inferences about the mechanism of disease; Immune destruction of hematopoietic cells.

Highlights • Circulating S100A8 levels are elevated in the plasma of myelodysplastic patients. • S100A8, S100A9 and S100A8/A9 are not increased in acquired aplastic anemia patients at diagnosis. • Circulating S100A8/A9 increases after therapy in non-responding aplastic anemia patients. • Correlations with blood counts may mirror the higher number of blood cells in some patients. Abstract The alarmin family members S100A8 and S100A9 are acute phase inflammation proteins, but they also have been proposed as biomarkers in many malignant and non-malignant diseases. In this study, circulating S100A8 and S100A9 homodimers and S100A8/A9 heterodimers in plasma were systematically investigated by ELISA in aplastic anemia (AA) and myelodysplastic syndromes (MDS). Plasma was obtained from 58 severe AA (SAA) and 30 MDS patients, and from 47 age- and sex-matched healthy donors. In 40 out of the 58 AA subjects, S100A protein levels were measured before and 6 months after immunosuppressive therapy (IST). No differences were observed in AA patients at diagnosis compared to healthy controls for circulating S100A homodimers and heterodimers. After therapy, SAA-responders showed significantly increased circulating S100A8. Non-responding patients had significantly higher levels of circulating S100A8/A9 compared to responders and healthy controls, but without variations of S100A8 and S100A9 homodimers. In MDS patients, circulating S100A8 was significantly elevated compared to those of AA and/or healthy controls. By Pearson correlation analysis of protein levels and blood counts, multiple correlations were found. However, as S100A8 and S100A9 are abundantly present in white blood cells and platelets, correlations with blood counts likely mirror the higher number of cells in the blood of some patients. In conclusion, our findings indicate that circulating S100A8 is increased in MDS but not in AA, and that may be useful to distinguish these diseases in the differential diagnosis of bone marrow failure syndromes. Clinicaltrials.gov identifiers: NCT00260689, NCT00604201, NCT01328587, NCT01623167, NCT00001620, NCT00001397. [ABSTRACT FROM AUTHOR]

Highlights • SOMAscan is an aptamer-based proteomic technology for large-scale studies. • Nineteen serum proteins are proposed as biomarkers and prognosticators of aplastic anemia (AA). • Serum DKK1, SELL, CCL17, and HGF are validated as novel markers of AA. • Twenty-eight plasma proteins are identified as candidate biomarkers of AA. • More than 600 proteins can be used as biomarkers of AA in both serum and plasma. Single-stranded oligonucleotides containing deoxyuridine are aptamers (SOMAmers) that can bind proteins with high specificity and affinity and slow dissociation rates. SOMAscan, an aptamer-based proteomic technology, allows measurement of more than 1,300 proteins simultaneously for the identification of new disease biomarkers. The aim of the present study was to identify new serum and plasma protein markers for diagnosis of acquired aplastic anemia (AA) and response to immunosuppressive therapies (IST). SOMAscan was used to screen 1,141 serum proteins in 28 AA patients before and after therapy and 1,317 plasma proteins in seven SAA patients treated with standard IST and a thrombopoietin receptor agonist. From our analysis, 19 serum and 28 plasma proteins were identified as possible candidate diagnostic and prognostic markers. A custom immunobead-based multiplex assay with five selected serum proteins (BMP-10, CCL17, DKK1, HGF, and SELL) was used for validation in a verification set (n = 65) of samples obtained before and after IST and in a blinded validation cohort at baseline (n = 16). After technical validation, four biomarkers were employed to predict diagnosis (accuracy, 88%) and long-term response to IST (accuracy, 79%). In conclusion, SOMAscan is a powerful tool for the identification of new biomarkers. We propose further larger studies to validate new candidate serum and plasma diagnostic and prognostic markers of AA. [ABSTRACT FROM AUTHOR]

Although PD-1 blockade has revolutionized cancer immunotherapy, immune-related adverse events (irAEs) present life-threatening complications. Recent reports of aplastic anemia (AA) as irAEs implicate PD-1/PD-L1 as important in preventing immune-mediated destruction of the hematopoietic niche. Infusion of PD-1-deficient (PD-1 knockout [KO]) lymph node (LN) cells into minor-antigen mismatched mice resulted in early mortality, as well as more severe bone marrow (BM) hypoplasia, anemia, and BM microarchitecture disruption in PD-1 KO LN-infused mice relative to mice that received B6 LN cell infusion. Mice that received PD-1 KO LN cells had more CD8 + T-cell infiltration of the BM and greater expansion of H60-specific CD8 + T cells than did their B6 LN-infused counterparts. In the spleen, CD8 + T cells were skewed to an effector memory phenotype, suggesting accelerated differentiation of PD-1 KO T cells. Our data suggest that PD-1 dysregulation has a role in murine BM failure and vigilance in irAE monitoring may be desirable to treat early AA and related cytopenias. [ABSTRACT FROM AUTHOR]

Background: Acquired aplastic anemia results from immune-mediated destruction of bone marrow. Immunosuppressive therapies are effective, but reduced numbers of residual stem cells may limit their efficacy. In patients with aplastic anemia that was refractory to immunosuppression, eltrombopag, a synthetic thrombopoietin-receptor agonist, led to clinically significant increases in blood counts in almost half the patients. We combined standard immunosuppressive therapy with eltrombopag in previously untreated patients with severe aplastic anemia.Methods: We enrolled 92 consecutive patients in a prospective phase 1-2 study of immunosuppressive therapy plus eltrombopag. The three consecutively enrolled cohorts differed with regard to the timing of initiation and the duration of the eltrombopag regimen (cohort 1 received eltrombopag from day 14 to 6 months, cohort 2 from day 14 to 3 months, and cohort 3 from day 1 to 6 months). The cohorts were analyzed separately. The primary outcome was complete hematologic response at 6 months. Secondary end points included overall response, survival, relapse, and clonal evolution to myeloid cancer.Results: The rate of complete response at 6 months was 33% in cohort 1, 26% in cohort 2, and 58% in cohort 3. The overall response rates at 6 months were 80%, 87%, and 94%, respectively. The complete and overall response rates in the combined cohorts were higher than in our historical cohort, in which the rate of complete response was 10% and the overall response rate was 66%. At a median follow-up of 2 years, the survival rate was 97%; one patient died during the study from a nonhematologic cause. Marked increases in bone marrow cellularity, CD34+ cell number, and frequency of early hematopoietic progenitors were noted. Rates of relapse and clonal evolution were similar to our historical experience. Severe rashes occurred in two patients, resulting in the early discontinuation of eltrombopag.Conclusions: The addition of eltrombopag to immunosuppressive therapy was associated with markedly higher rates of hematologic response among patients with severe aplastic anemia than in a historical cohort. (Funded by the National Heart, Lung, and Blood Institute; ClinicalTrials.gov number, NCT01623167 .). [ABSTRACT FROM AUTHOR]

Memory stem T cells (TSCMs) constitute a long-lived, self-renewing lymphocyte population essential for the maintenance of functional immunity. Hallmarks of autoimmune disease pathogenesis are abnormal CD4+ and CD8+ T cell activation. We investigated the TSCM subset in 55, 34, 43, and 5 patients with acquired aplastic anemia (AA), autoimmune uveitis, systemic lupus erythematosus, and sickle cell disease, respectively, as well as in 41 age-matched healthy controls. CD8+ TSCM frequency was significantly increased in AA compared with healthy controls. An increased CD8+ TSCM frequency at diagnosis was associated with responsiveness to immunosuppressive therapy, and an elevated CD8+ TSCM population after immunosuppressive therapy correlated with treatment failure or relapse in AA patients. IFN-γ and IL-2 production was significantly increased in various CD8+ and CD4+ T cell subsets in AA patients, including CD8+ and CD4+ TSCMs. CD8+ TSCM frequency was also increased in patients with autoimmune uveitis or sickle cell disease. A positive correlation between CD4+ and CD8+ TSCM frequencies was found in AA, autoimmune uveitis, and systemic lupus erythematosus. Evaluation of PD-1, CD160, and CD244 expression revealed that TSCMs were less exhausted compared with other types of memory T cells. Our results suggest that the CD8+ TSCM subset is a novel biomarker and a potential therapeutic target for AA. [ABSTRACT FROM AUTHOR]

The pathophysiology of severe aplastic anemia (SAA) is immune-mediated destruction of hematopoietic stem and progenitor cells (HSPCs). Most patients respond to immunosuppressive therapies, but a minority transform to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML), frequently associated with monosomy 7 (-7). Thirteen SAA patients were analyzed for acquired mutations in myeloid cells at the time of evolution to -7, and all had a dominant HSPC clone bearing specific acquired mutations. However, mutations in genes associated with MDS/AML were present in only 4 cases. Patients who evolved to MDS and AML showed marked progressive telomere attrition before the emergence of -7. Single telomere length analysis confirmed accumulation of short telomere fragments of individual chromosomes. Our results indicate that accelerated telomere attrition in the setting of a decreased HSPC pool is characteristic of early myeloid oncogenesis, specifically chromosome 7 loss, in MDS/AML after SAA, and provides a possible mechanism for development of aneuploidy. [ABSTRACT FROM AUTHOR]