Bone marrow and peripheral blood involvement of relapsed diffuse large B-cell lymphoma after prior cold agglutinin disease.

The author report an interesting case of relapsed diffuse large B-cell lymphoma (DLBCL) with bone marrow (BM) and peripheral blood (PB) involvement after prior cold agglutinin disease (CAD). A minority of patients with DLBCL present with CAD, and BM or PB involvement with CAD are quite rare. Differential diagnosis of patients with CAD should include DLBCL, considering the possibility of BM or even PB involvement.

Targeted next-generation sequencing revealed a novel homozygous mutation in the LRBA gene causes severe haemolysis associated with Inborn Errors of Immunity in an Indian family.

OBJECTIVES: LPS-responsive beige-like anchor protein (LRBA) deficiency abolishes LRBA protein expression due to biallelic mutations in the LRBA gene that lead to autoimmune manifestations, inflammatory bowel disease, hypogammaglobulinemia in early stages, and variable clinical manifestations. MATERIALS AND METHODS: Mutational analysis of the LRBA gene was performed in Indian patients using targeted Next Generation Sequencing (t-NGS) and confirmed by Sanger sequencing using specific primers of exons 53. Then, bioinformatics analysis and protein modeling for the novel founded mutations were also performed. The genotype, phenotype correlation was done according to the molecular findings and clinical features. RESULTS: We report an unusual case of a female patient born of a consanguineous marriage, presented with severe anaemia and jaundice with a history of multiple blood transfusions of unknown cause up to the age of 5 yrs. She had hepatosplenomegaly with recurrent viral and bacterial infections. Tests for hemoglobinopathies, enzymopathies, and hereditary spherocytosis were within the normal limits. The t-NGS revealed a novel homozygous missense variation in exon 53 of the LRBA gene (chr4:151231464C > T; c.7799G > A) (p.C2600Y), and the parents were heterozygous. The further immunological analysis is suggestive of hypogammaglobulinaemia and autoimmune haemolytic anaemia. The bioinformatics tools are suggestive of deleterious and disease-causing variants. CONCLUSION: This study concludes the importance of a timely decision of targeted exome sequencing for the molecular diagnostic tool of unexplained haemolytic anaemia with heterogeneous clinical phenotypes.

T-follicular helper cell expansion and chronic T-cell activation are characteristic immune anomalies in Evans syndrome.

Pediatric Evans syndrome (pES) is increasingly identified as the presenting manifestation of several inborn errors of immunity. Despite an improved understanding of genetic defects in pES, the underlying immunobiology of pES is poorly defined, and characteristic diagnostic immune parameters are lacking. We describe the immune characteristics of 24 patients with pES and compared them with 22 patients with chronic immune thrombocytopenia (cITP) and 24 healthy controls (HCs). Compared with patients with cITP and HC, patients with pES had increased circulating T-follicular helper cells (cTfh), increased T-cell activation, and decreased naïve CD4+ T cells for age. Despite normal or high immunoglobulin G (IgG) in most pES at presentation, class-switched memory B cells were decreased. Within the cTfh subset, we noted features of postactivation exhaustion with upregulation of several canonical checkpoint inhibitors. T-cell receptor ß chain (TCR-ß) repertoire analysis of cTfh cells revealed increased oligoclonality in patients with pES compared with HCs. Among patients with pES, those without a known gene defect had a similar characteristic immune abnormality as patients with defined genetic defects. Similarly, patients with pES with normal IgG had similar T-cell abnormalities as patients with low IgG. Because genetic defects have been identified in less than half of patients with pES, our findings of similar immune abnormalities across all patients with pES help establish a common characteristic immunopathology in pES, irrespective of the underlying genetic etiology.

In vitro Immunomodulatory Effects of Human Umbilical Cord-Derived Mesenchymal Stem Cells on Peripheral Blood Cells from Warm Autoimmune Hemolytic Anemia Patients.

INTRODUCTION: Autoimmune hemolytic anemia is a potentially lethal disease characterized by autoimmune hemolysis. Although human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) have been reported as a promising therapy, there is limited evidence regarding warm autoimmune hemolytic anemia (wAIHA) patients. This study aimed to investigate the potential therapeutic effects of hUC-MSCs via immune regulation in wAIHA patients. METHODS: Peripheral blood mononuclear cells (PBMCs) from 10 wAIHA patients and 8 healthy controls were isolated from peripheral blood and cultured for 3 days with or without the presence of hUC-MSCs; PBMCs were co-cultured with hUC-MSCs using Transwell assays. The supernatant cytokine levels were measured after culture through AimPlex Multiple Immunoassays for Flow, including IL-2, IL-4, IL-10, IFN-γ, TNF-α, and IL-17A. The percentages of regulatory T cells, regulatory B cells, and Th1/Th2 in PBMCs were also assessed before and after culturing. RESULTS: In the wAIHA group, hUC-MSCs could upregulate the Treg and Breg proportions after culturing for 3 days, and the Treg and Breg percentages increased after co-culturing with hUC-MSCs in the wAIHA group compared with PBMC cultured alone for 3 days (8.29 ± 8.59 vs. 6.82 ± 1.32, 3.82 ± 1.87 vs. 1.75 ± 1.20, respectively). Compared with the PBMC wAIHA group, the levels of TNF-α (2.13 ± 2.07 vs. 16.20 ± 21.13 pg/mL, p = 0.019) and IL-10 (10.51 ± 18.42 vs. 37.78 ± 44.20 pg/mL, p = 0.012) were significantly elevated in the PBMC + hUC-MSCs wAIHA group. CONCLUSION: The hUC-MSCs contributed to the increasing proportion of regulatory cell populations in PBMCs of wAIHA patients, thereby potentially regulating autoimmune response; thus, hUC-MSCs may be a promising approach for wAIHA treatment.

Autoimmune Cytopenia as an Early and Initial Presenting Manifestation in Activated PI3 Kinase Delta Syndrome: Case Report and Review.

Activated PI3 kinase delta syndrome (APDS) is a combined immunodeficiency characterized by recurrent sinopulmonary infections, increased risk of herpesvirus infections, lymphoproliferation, autoimmunity, and increased risk of lymphoid malignancies. Gain-of-function mutations in PIK3CD and PIK3R1 result in increased phosphoinositide-3-kinase-delta activity which causes hyperactivation of lymphocytes and abnormal development and activation of T and B cells. Cytopenias are the most common autoimmune process occurring in patients with APDS and typically occur as a later manifestation of the disease. Here we present a female patient with an early autoimmune hemolytic anemia, hepatosplenomegaly, and frequent infections presenting in infancy, followed by development of significant lymphadenopathy before her diagnosis with APDS type 1. She had significant improvement in her infectious history with immunoglobulin replacement, and control of autoimmune hemolytic anemia with initiation of sirolimus after her diagnosis with APDS type 1. We utilize this case to review the literature on APDS and present the novel finding of early-onset autoimmune disease in the setting of APDS. Autoimmune cytopenias are seen in many primary immunodeficiencies, and workup of autoimmune cytopenias in young patients should include evaluation for underlying immune disorder.

Immunotactoid glomerulonephritis in a patient with cold agglutinins: causal association or mere coincidence?

We report a case of immunotactoid glomerulonephritis (ITG) in a patient with cold agglutinins. An 86-year-old Japanese male with a history of hypertension, dyslipidemia, and gastric malignancy presented to our hospital for the evaluation of proteinuria and hematuria. He had an elevated blood pressure of 200/77 mmHg and edema of the lower extremities. Initial blood test results revealed an impaired renal function (creatinine, 1.37 mg/dL) and hypoalbuminemia (albumin, 2.6 g/dL). His estimated daily urinary protein was 5.89 g/g creatinine, meeting the diagnostic criteria for nephrotic syndrome. The selectivity index for proteinuria indicated low selectivity (0.329). We conducted a renal biopsy to identify the cause of nephrotic syndrome. Immunofluorescence microscopy demonstrated positive staining of IgM, C4, and C1q. Electron microscopy exhibited mesangial expansion with inflammatory cells and a lobular structure, suggesting membranoproliferative glomerulonephritis. Subendothelial deposits containing microtubular structures with a diameter of approximately 30-200 nm were found, concurrent with the criteria for the diagnosis of ITG. Screening for lymphoproliferative diseases and immunological abnormalities revealed a positive direct Coombs test result and the presence of cold agglutinins. Paraproteinemia was absent. The similarities between cold agglutinin disease and ITG, including the production of autoantibodies and involvement of complement pathways, raise the possibility that cold agglutinins played a role in the development of ITG; however, we were unable to prove it due to difficulties in detecting cold agglutinins on renal histology. We discuss the possible implications for pathogenesis considering prior reports on nephrotic syndrome being potentially associated with cold agglutinins.

Features of serum complement C3 and C4 levels in autoimmune hemolytic anemia patients.

INTRODUCTION: Abnormally activated complement system induces erythrolysis in a part of autoimmune hemolytic anemia (AIHA) patients. However, the alterations in serum complement levels in these patients are seldom reported. In this study, we aimed to evaluate the serum complement features of AIHA patients according to different clinical and laboratory characteristics and to find relationships between complement levels and hemolysis-associated laboratory indexes. METHODS: A retrospective analysis of 146 AIHA patients was performed, and serum complement C3 and C4 levels were compared between control subjects and AIHA patients with different subtypes. Correlations of serum C3/C4 levels with titers of cold agglutinin test (CAT), direct antiglobulin test (DAT), and serological indexes were assessed. Spearman correlation analysis was performed to analyze the relationship between serum complement levels and other laboratory indexes. RESULTS: Autoimmune hemolytic anemia patients showed reduced serum C3 levels, while serum C4 levels tended to be lower in DAT-positive AIHA patients but not in DAT-negative AIHA patients. Patients with warm AIHA secondary to connective tissue diseases and cold agglutinin disease/cold agglutinin syndrome had the lowest serum C3/C4 levels. Serum C4 levels were negatively correlated with CAT (P = .004) and DAT (anti-C3d) (P = .007) titers. In patients with positive CAT and/or DAT (anti-C3d) but negative DAT (anti-IgG), serum C3/C4 levels were negatively correlated with indirect bilirubin (P = .017 and =.026, respectively). CONCLUSION: The study findings may be helpful in not only unraveling the mechanism underlying hemolysis in AIHA but also diagnosing AIHA and selecting targeted treatment strategies.

How I treat warm autoimmune hemolytic anemia.

Warm autoimmune hemolytic anemia (wAIHA) is caused by increased erythrocyte destruction by immunoglobulin G (IgG) autoantibodies, with or without complement activation. Antibody-dependent cell-mediated cytotoxicity by macrophages/activated lymphocytes occurs in the lymphoid organs and spleen (extravascular hemolysis). The ability of the bone marrow (BM) to compensate determines clinical severity. The different pathogenic mechanisms, their complex interplay, and changes over time may explain wAIHA's great clinical heterogeneity and unpredictable course. The disease may be primary, drug induced, or associated with lymphoproliferative neoplasms, autoimmune and infectious diseases, immunodeficiencies, solid tumors, or transplants. Therapeutic interventions include steroids, splenectomy, immunosuppressants, and rituximab; the latter is increasingly used in steroid-refractory cases based on evidence from the literature and a few prospective trials. We present 5 patient case studies highlighting important issues: (1) the diagnosis and proper use of steroid therapy, (2) the concerns about the choice between rituximab and splenectomy in second-line treatment, (3) the need of periodical re-evaluation of the disease to assess the possible evolution of relapsed/refractory cases in myelodysplastic and BM failure syndromes, and (4) the difficulties in managing cases of severe/acute disease that are at high risk of relapse. Incorporating novel targeted therapies into clinical practice will be an exciting challenge in the future.

Refractory and Fatal Presentation of Severe Autoimmune Hemolytic Anemia in a Child With the DNASE1L3 Mutation Complicated With an Additional DOCK8 Variant.

Various autoimmune diseases may be associated with primary immune deficiencies. We reported a case with a loss-of-function mutation in DNASE1L3, a gene described previously in families with systemic lupus erythematosus. In addition, the patient showed a novel homozygous missense variant in DOCK8, a gene known to be responsible for the hyper-IgE recurrent infection syndrome (HIES). A 3-year-old girl born to consanguine parents presented with chronic urticarial rash, hemolytic anemia, pulmonary hemorrhage, and hypovolemic shock findings. She had a low hemoglobin level, a positive direct antiglobulin test, antinuclear antibody and anti-double stranded DNA, low C3 and C4, third-degree tricuspid regurgitation, and severe enlargement of the right ventricle on echocardiography, suggesting pulmonary embolism. Despite treatment with intravenous immunoglobulin, pulse metilprednisolone, rituximab, and supportive treatment for shock, the patient died on the seventh day. Whole-exome sequencing indicated a homozygous stop variant c.537G>A (p. Trp179Ter) in DNASE1L3. In addition, a possibly pathogenic homozygous missense variant in the HIES gene DOCK8 was detected. The occurrence of potentially clinically relevant, genetic variants in several genes posed various challenges with respect to diagnosis, treatment, and prognosis.

Impact of autoimmune cytopenias on severity of childhood-onset systemic lupus erythematosus: A single-center retrospective cohort study.

OBJECTIVE: To assess whether children with autoimmune cytopenias prior to or at diagnosis of systemic lupus erythematosus (cSLE), differ phenotypically from other cSLE patients; and have a lower risk and severity of lupus nephritis (LN) as observed in prior adult studies. To assess the effect of prior immune therapy for autoimmune cytopenias on 2-year risk of LN. METHODS: This was a retrospective cohort study of incident cSLE cases. We included patients aged less than 17 years at diagnosis. We excluded patients with LN at cSLE diagnosis. Our follow-up period was 2 years. We defined autoimmune cytopenias as either autoimmune hemolytic anemia, immune thrombocytopenia or Evan's syndrome. RESULTS: Forty-three (33%) of the 130 patients had autoimmune cytopenias before or at cSLE diagnosis. Those with autoimmune cytopenias had significantly more neuropsychiatric symptoms and higher mean ESR but less arthritis, malar rash and myositis versus those without autoimmune cytopenias. They had lower 2-year incidence proportion of LN compared to other cSLE patients (7% vs 15%). Of the 16 patients who developed LN, those with autoimmune cytopenias had mostly class V (2 of 3 patients) versus mostly class III and IV in those without autoimmune cytopenias (6 of 12 patients). None of the 13 patients pre-treated for autoimmune cytopenias developed LN. CONCLUSION: Patients with autoimmune cytopenias before or at cSLE diagnosis have intriguing differences from other cSLE patients. They may represent a unique sub-type of cSLE patients and should be further explored.

The Wide Spectrum of Pathophysiologic Mechanisms of Paraproteinemic Neuropathy.

Monoclonal gammopathy is encountered quite frequently in the general population. This type of hematologic abnormality may be mild, referred to as monoclonal gammopathy of undetermined significance or related to different types of hematologic malignancies. The association of a peripheral neuropathy with monoclonal gammopathy is also fairly common, and hemopathy may be discovered in an investigation of peripheral neuropathy. In such a situation, it is essential to determine the exact nature of the hematologic process in order not to miss a malignant disease and thus initiate the appropriate treatment (in conjunction with hematologists and oncologists). In this respect, nerve biopsy (discussed on a case-by-case basis) is of great value in the management of such patients. We therefore propose to present the objectives and main interests of nerve biopsy in this situation.

Non-invasive prenatal testing for management of haemolytic disease of the fetus and newborn induced by maternal alloimmunisation.

Anti-D immunoglobulin prophylaxis reduces the risk of RhD negative women becoming alloimmunised to the RhD antigen and is a major preventative strategy in reducing the burden of haemolytic disease of the fetus and newborn (HDFN). HDFN also arises from other maternal red cell antibodies, with the most clinically significant, after anti-D, being anti-K, anti-c and anti-E. Among the 39 human blood group systems advanced genomic technologies are still revealing novel or rare antigens involved in maternal alloimmunisation. Where clinically significant maternal antibodies are detected in pregnancy, non-invasive prenatal testing (NIPT) of cell-free fetal DNA provides a safe way to assess the fetal blood group antigen status. This provides information as to the risk for HDFN and thus guides management strategies. In many countries, NIPT fetal RHD genotyping as a diagnostic test using real-time PCR has already been integrated into routine clinical care for the management of women with allo-anti-D to assess the risk for HDFN. In addition, screening programs have been established to provide antenatal assessment of the fetal RHD genotype in non-alloimmunised RhD negative pregnant women to target anti-D prophylaxis to those predicted to be carrying an RhD positive baby. Both diagnostic and screening assays exhibit high accuracy (over 99 %). NIPT fetal genotyping for atypical (other than RhD) blood group antigens presents more challenges as most arise from a single nucleotide variant. Recent studies show potential for genomic and digital technologies to provide a personalised medicine approach with NIPT to assess fetal blood group status for women with other (non-D) red cell antibodies to manage the risk for HDFN.

Bortezomib treatment of steroid-refractory Evans syndrome in children.

Treatment of refractory Evans syndrome (ES) remains a challenge in hematology practice. Due to rarity of this condition, evidence-based approaches are limited and often treatment choices stem from small case series or anecdotal experiences. There is mounting evidence that some patients have genetic defects that could be targeted with promising preliminary results. Here, we describe three very refractory pediatric ES cases treated on bortezomib without adverse effects. Two of the three patients had dramatic and long-lasting recovery that started following the initial doses of the drug. Clinical trials to assess the role of bortezomib in ES treatment are warranted.

Role of complement in alloimmunization and hyperhemolysis.

PURPOSE OF REVIEW: The purpose of this review is to summarize the role of complement in regulating the removal of a target alloantigen following an incompatible red blood cell (RBC) transfusion, the formation of alloantibodies following RBC alloantigen exposure, and the development of hyperhemolysis in patients with sickle cell disease (SCD). RECENT FINDINGS: Recent studies demonstrate that complement can accelerate alloantibody-mediated removal of target alloantigens from the RBC surface following incompatible transfusion. Complement also influences alloantigen availability during developing alloimmune responses and serves as a unique mediator of CD4 T-cell-independent alloantibody formation following RBC alloantigen exposure. Finally, alternative complement pathway activation appears to play a key role in the development of acute hemolytic episodes in patients with SCD, providing a potential druggable target to prevent acute complications in patients with this disease. SUMMARY: Recent studies suggest that complement can regulate a wide variety of processes germane to hematology, from transfusion complications to baseline hemolysis in patients with SCD. As the role of complement in various disease processes becomes more fully understood, the ability to leverage recently developed complement modulating drugs will only continue to enhance providers' ability to favorably intervene in many hematological diseases.

Sirolimus is effective for primary relapsed/refractory autoimmune cytopenia: a multicenter study.

Autoimmune cytopenia includes autoimmune hemolytic anemia (AIHA), immune thrombocytopenia (ITP), and Evans syndrome (ES) caused by abnormal activation of autoimmunity and has a considerable refractory/relapse rate. To evaluate the efficacy and toxicity of sirolimus of primary relapsed/refractory autoimmune cytopenia, records of 45 patients with primary relapsed/refractory AIHA, ES, or ITP from October 2016 to January 2019 in two institutions, were collected; there were 3 pediatric patients and 42 adult patients. The median age at diagnosis was 31 (1-84) years. Patients were treated for a median of 14 (6-39) months and followed-up for a median of 18 (10-40) months. Thirty-eight patients responded to sirolimus, with 28 complete responses (CRs) and a median of 2 (2-5) months to response. Five patients had mucositis; the incidences of other adverse events were all less than 5%. Four patients relapsed, making the CR and overall response rate 46.7% and 75.6% at the end of follow-up. There were no differences in patient age, sex, time from diagnosis to sirolimus, serum sirolimus concentration, and disease distribution between CR and non-CR patients, or between responders and nonresponders, though AIHA patients were likely to relapse less and respond better. In conclusion, sirolimus is effective for patients with primary relapsed/refractory autoimmune cytopenia with a low relapse rate and good tolerance.