Idiopathic Pure Red Cell Aplasia Research Articles

Idiopathic pure red cell aplasia: A case series from India

Pure red cell aplasia (PRCA) is a rare hematologic disorder characterized by normocytic normochromic anemia with reticulocytopenia and depleted erythroid precursors. It is reported mainly from western countries. We report our experience in two patients of PRCA from India, the clinical characteristics, differential diagnosis, and their management. Our patients showed improvements following treatment with corticosteroids, blood transfusion, and/or rituximab.

口腔癌治療中に発症した重度貧血が契機となり診断に至った特発性赤芽球癆の1例

口腔癌治療中に発症した重度貧血が契機となり診断に至った特発性赤芽球癆の1例

Somatic mutations in acquired pure red cell aplasia

Acquired pure red cell aplasia (PRCA) is a syndrome characterized by anemia and a marked reduction of erythroid progenitor cells with various etiologies. The 3 major subtypes of PRCA are idiopathic PRCA, large granular lymphocytic leukemia-associated PRCA and thymoma-associated PRCA, which are thought to be caused by a T-cell-mediated mechanism. In these 3 subtypes, an expansion of clonal cytotoxic T cells is often detected. In addition, those T cells recurrently harbor somatic mutations of STAT3, a gene coding one of the important signal transducers in the JAK/STAT system. Somatic mutations of clonal hematopoiesis (CH)-related genes, including epigenetic modifying genes, have also been reported, however, the data are still not mature enough upon which to draw conclusion, Somatic mutations of STAT3 and CH-related genes may be unique characteristics of acquired PRCA. However, their involvement in dyserythropoiesis or clinical relevance to the clinical course of those somatic mutations. Mutational landscapes, their involvements in dyserythropoiesis and clinical relevance in acquired PRCA remains unclear, and further investigation is needed.

Daratumumab for treatment-refractory acquired idiopathic pure red cell aplasia.

Not available.

Correction: Idiopathic pure red cell aplasia and nonregenerative immune-mediated anemia in dogs: 43 cases (1988-1999).

Correction: Idiopathic pure red cell aplasia and nonregenerative immune-mediated anemia in dogs: 43 cases (1988-1999).

Clinical and pathophysiological features of acquired pure red cell aplasia: based on the concept of T-cell dysregulations

Acquired pure red cell aplasia (PRCA) develops in a variety of contexts and thus, should be regarded as a syndrome. The three major subtypes of acquired PRCA are idiopathic PRCA, T cell large granular lymphocytic leukemia (T-LGLL)-associated PRCA, and thymoma-associated PRCA. Although the Japanese National Research Group on Idiopathic Bone Marrow Failure Syndromes of the Ministry of Health, Labor and Welfare of Japan has made significant contributions to our understanding of PRCA, details of its clinical characteristics, and pathophysiological mechanisms remain largely unknown. A recent epidemiological analysis using the JSH Hematologic Disease Registry revealed that approximately 100 new cases with acquired PRCA were diagnosed annually in Japan, which was higher than previously thought to be. The median age of the patients was 73 years. A prospective observational study on chronic PRCA (PRCA2016) is currently ongoing, and it may provide new clinical insights into acquired PRCA. Dysregulation of T cells has been shown to play a central role in PRCA. We studied T cell clonalities and STAT3 mutations in 90 PRCA patients and discovered that clonal T cell expansions were frequently recognized and closely associated with STAT3 mutations in the three major types of PRCA.

Identification of Adverse Risk Factors for Survival in Acquired Pure Red Cell Aplasia Receiving Immunosuppressive Therapy By the Japan PRCA2004/2006 Study and Introduction to the New Cohort Study PRCA2016

Pure red cell aplasia (PRCA) is characterized by normocytic normochromic anemia associated with reticulocytopenia and erythroid hypoplasia in otherwise normal bone marrow. The acquired chronic PRCA may present as a primary hematological disorder in the absence of any other diseases or develop associated with certain conditions such as thymoma, lymphoproliferative disorders, infections, collagen vascular diseases, or after exposure to a wide variety of drugs. Idiopathic PRCA and secondary PRCA not responding to the treatment of the underlying diseases are generally treated by immunosuppressive therapy and the most patients require long-term treatment. Glucocorticoids, cyclosporine and cyclophosphamide are commonly used for initial treatment as monotherapy or combination therapy and majority of the patients respond to immunosuppression. However, most patients treated with these medications relapse during tapering of the treatment. Because of the extreme rarity of this disease, the long-term outcome of chronic PRCA following immunosuppression remains unclear and the adverse risk factors for survival have to be elucidated.In order to identify the adverse risk factors for survival in acquired chronic PRCA following immunosuppression and improve the prognosis, we have conducted nationwide surveys for adult acquired chronic PRCA in Japan. We conducted the first study for adult chronic PRCA in japan in 2004 and 2006 that had been diagnosed between 1990 and 2006 across 109 institutions, and collected the data on a total of 185 patients (PRCA2004/2006 study). We have also started another cohort study in 2016 in collaboration with the Japanese Society of Hematology (JSH) (PRCA2016 study) that was approved by the institutional review board and the ethical committee of the JSH.In PRCA2004/2006 study 73 patients (39%) were classified as having idiopathic PRCA and 112 patients (61%) as having secondary PRCA, including 42 thymoma- and 14 large granular lymphocyte leukemia (LGL)-associated PRCA. The details on idiopathic, thymoma-associated and LGL-leukemia-associated PRCA have been previously reported (Sawada K, et al. Haematologica 2007;92:1021; Hirokawa M, et al. Haematologica 2008;93:27; Fujishima N, et al. Haematologica 2008;93:1555). The last questionnaires for the patients with these three subtypes of PRCA were sent to the institutions in 2012 for collecting the data on relapse, subsequent treatment and efficacy and outcome.The PRCA2004/2006 study revealed that the median overall survival (OS) has not yet been reached in idiopathic PRCA following immunosuppression and the estimated median OS times in thymoma-associated and LGL leukemia-associated PRCA were 142 and 148 months, respectively. Twenty-two deaths were reported and the major causes of death were infection and organ failure (Hirokawa M, et al. Brit J Haematol 2015;169:879). Poor response to induction therapy and relapse of anemia were demonstrated to be associated with death and this was also confirmed by the Mantel-Byar analysis (p<0.001) (Fig. 1). The efficacy of re-induction therapy for relapsing PRCA was inferior to the initial inductive therapy (p<0.001) (Fig. 2).With the results of this study, we have proposed the clinical guidelines for management of PRCA in 2011 and we conducted another cohort study in 2016 in collaboration with the Japanese Society of Hematology (JSH) (PRCA2016 study). The primary endpoint of this study is the OS in chronic PRCA following immunosuppression and the principal aim is to learn whether or not the prognosis would be improved. Between 2006 and 2014, 554 PRCA patients were registered in the JSH hematological disorder registry and we asked the physicians to participate in this study (Fig. 3). The potential size of patient cohort is 181, including 116 idiopathic and 65 secondary PRCA and these patients will be followed up during the next ten years. The PRCA2016 cohort study is expected to reveal the unbiased results regarding the long-term survival and quality of life in PRCA following immunosuppression. [Display omitted] DisclosuresFujisawa:Bristol myers Squibb: Honoraria; Beckman Coulter: Honoraria; Eisai: Honoraria; Janssen Pharmaceutical: Honoraria; Novartis: Honoraria; Astellas: Honoraria, Research Funding; Nippon Shinyaku: Honoraria, Research Funding; Sumitomo Dainippon Pharma: Honoraria; Otsuka Pharmaceutical: Honoraria, Research Funding; SHIONOGI CO, LTD: Honoraria, Research Funding; Shire plc: Honoraria; Alexion Pharmaceutical: Honoraria; Pfizer: Honoraria, Research Funding; Ono Pharmaceutical: Honoraria, Research Funding; Takeda Pharmaceuticals: Honoraria, Research Funding; Celgene: Honoraria; Chugai Pharmaceutical: Honoraria, Research Funding; Kyowa Hakko Kirin: Research Funding; MSD: Research Funding. Yonemura:Alexion Pharma GK: Honoraria, Research Funding. Kurokawa:Pfizer Inc.: Research Funding; Celgene Corporation: Speakers Bureau; Nippon Shinyaku Co., Ltd.: Other: Scholarship donations for the laboratory, Speakers Bureau; Bristol-Myers Squibb: Speakers Bureau; Takeda Pharmaceutical Company Limited.: Research Funding, Speakers Bureau; Astellas Pharma Inc.: Research Funding.

Clonal hematopoiesis in adult pure red cell aplasia

Idiopathic pure red cell aplasia (PRCA) and secondary PRCA associated with thymoma and large granular lymphocyte leukemia are generally considered to be immune-mediated. The PRCA2004/2006 study showed that poor responses to immunosuppression and anemia relapse were associated with death. PRCA may represent the prodrome to MDS. Thus, clonal hematopoiesis may be responsible for treatment failure. We investigated gene mutations in myeloid neoplasm-associated genes in acquired PRCA. We identified 21 mutations affecting amino acid sequences in 11 of the 38 adult PRCA patients (28.9%) using stringent filtering of the error-prone sequences and SNPs. Four PRCA patients showed 7 driver mutations in TET2, DNMT3A and KDM6A, and 2 PRCA patients carried multiple mutations in TET2. Five PRCA patients had mutations with high VAFs exceeding 0.3. These results suggest that clonal hematopoiesis by stem/progenitor cells might be related to the pathophysiology of chronic PRCA in certain adult patients.

Diagnosis and management of pure red cell aplasia

Pure red cell aplasia (PRCA) is characterized by normocytic anemia with reticulocytopenia and marked reduction of the bone marrow erythroid precursors. PRCA may be congenital (Diamond-Blackfan anemia) or acquired. Acquired PRCA may present in the context of various backgrounds, the most common type in Japan being idiopathic, thymoma-associated, and large granular lymphocyte leukemia. Idiopathic and secondary PRCA that do not respond to the treatment of the underlying disease are generally treated using immunosuppressive agents. A retrospective study PRCA2004/2006 suggests that maintenance therapy and management of infectious complications is crucial for improving the prognosis in patients with PRCA. Recently, allogeneic stem cell transplantation has been considered as a potential option for the treatment of patients with PRCA who are refractory to immunosuppressive therapy. Sirolimus and roxadustat may be effective for relapsed/refractory PRCA with renal insufficiency and anti-erythropoietin antibody-mediated PRCA, respectively. Some gene mutations were detected in certain patients who had acquired PRCA, and the identification of STAT3 mutations may be useful in PRCA management. A prospective cohort study PRCA2016 has been ongoing in Japan, and novel discoveries provide hope for improving the outcome in patients with PRCA.

Somatic Mutations of Myeloid Malignancy-Associated Genes in Acquired Pure Red Cell Aplasia in Adults

Background. Idiopathic pure red cell aplasia (PRCA) and secondary PRCA not responding to the treatment of the underlying diseases are generally thought be immune-mediated and treated by immunosuppressive therapy. We previously conducted the PRCA2004/2006 study and reported that poor response to induction therapy and relapse of anemia were associated with death. Principal causes of death were infections and organ failure. Based on the literatures, idiopathic PRCA may represents the prodrome to myelodysplastic syndromes. Theoretically, there are two potential mechanisms of unresponsiveness to immunosuppression; the clonal hematopoiesis by the stem/progenitor cells that have undergone somatic mutations during disease progression of PRCA and the clonal changes of auto-aggressive lymphocytes reacting against erythroid progenitors.Objectives. In this study, we investigated the somatic mutations of myeloid malignancy-associated genes in acquired PRCA in order to determine how often clonal hematopoiesis is detected in this disorder.Materials and Methods. This study included 23 patients with chronic acquired PRCA (12 idiopathic, 7 thymoma-, 2 LGL leukemia- and 2 systemic lupus erythematosus-associated PRCA) with a median age of 62 (range: 40-62). Disease status was varying. After obtaining informed consent, heparinized blood was drawn and mononuclear cells were separated by density gradient centrifugation. Extracted genomic DNA samples were subjected to targeted sequencing for 54 myeloid malignancy-associated genes using a TruSight Myeloid Sequencing Panel kit according to the manufacturer's instruction (Illumina). Criteria for the significant somatic mutations of myeloid malignancy-associated genes in the present study were as follows: potential functional consequences such as missense, nonsense or frameshift mutations; exclusion of previously reported SNPs; being recurrently detected in two sequencing runs; variant allele frequency (VAF) exceeding 0.02 and less than 0.40. The institutional review board approved the experimental protocol.Results. We detected some mutations of the targeted genes in 20 out of 23 patients, and the somatic mutations defined by the criteria mentioned above were found in 10 patients including 6 idiopathic, 3 thymoma-associated and one LGL leukemia-associated PRCA (Fig. 1). These 10 patients had 38 distinct mutations in 20 genes. Variant allele frequencies were 0.02 to 0.37 (median, 0.04; average, 0.06, Fig. 2). Four patients had more than one mutated genes and multiple genes were mutated in some patients (Fig. 1). The most frequently mutated gene was CUX1 that was found in four patients, and STAG2, DNMT3A, KDM6A, SMC3A, ASXL1, TET2 and TP53 were mutated in more than one patient.Discussion/Conclusion. This study demonstrated that myeloid malignancy-associated genes were somatically mutated in 43% of acquired chronic PRCA patients. This figure appears to exceed the prevalence rate of clonal hematopoiesis of indeterminate potential (CHIP) in the general population with the age of 60s. These mutations were presumably carried by monocytes, because DNA samples were prepared from PBMCs in this study cohort. Profiles of mutated genes in PRCA appear to be different from those of aplastic anemia that were previously reported by other groups. It is yet to be known whether this could result from the different nature of both diseases, or the difference in the experimental protocols. Our findings strongly encourage conducting a prospective study to confirm our observation and clarify the diagnostic and predictive values of somatic mutations of myeloid malignancy-associated genes in acquired PRCA. This project is ongoing in collaboration with the prospective cohort study PRCA2016 being conducted in Japan. [Display omitted] DisclosuresNakao:Kyowa Hakko Kirin Co., Ltd.: Honoraria; Novartis: Honoraria; Alexion Pharmaceuticals, Inc.: Consultancy, Honoraria. Matsuda:GlaxoSmithKline K.K.: Honoraria; Novartis Pharma K. K.: Honoraria; Chugai Pharmaceutical Co, Ltd.: Honoraria; Kyowa Hakko Kirin Co, Ltd.: Honoraria; Sumitomo Dainippon Pharma Co., Ltd.: Honoraria; Nippon Shinyaku Co., Ltd.: Honoraria; Celgene Corporation: Honoraria; Alexion Pharmaceuticals, Inc.: Honoraria; Sanofi K.K.: Honoraria; Beckman Coulter K.K.: Honoraria. Mitani:Kyowa Hakko Kirin Co., Ltd.: Consultancy, Research Funding, Speakers Bureau; Bristol-Myesr Squibb: Research Funding, Speakers Bureau; Celgene: Speakers Bureau; Chugai: Research Funding; Astellas: Research Funding; Sumitomo Dainippon: Research Funding; Novartis: Research Funding; Toyama Chemical: Research Funding.

Progress in the clinical management of pure red cell aplasia and future prospects

Pure red cell aplasia (PRCA) is a type of bone marrow failure syndrome (stem cell failure) and is characterized by severe normocytic, normochromic anemia associated with reticulocytopenia and the absence of erythroblasts in otherwise normal bone marrow. The acquired form of chronic PRCA may present as a primary hematological disease in the absence of any other diseases or secondary to thymoma, lymphoproliferative disorders, infections and collagen vascular diseases or after exposure to various drugs or chemicals. Thus, identifying the cause of PRCA is crucial for the optimal management of this disorder. Idiopathic PRCA and secondary PRCA refractory to treatment of the underlying diseases are both generally treated as an immune-mediated disorder. Most chronic PRCA patients successfully treated with immunosuppressants require maintenance immunosuppressive therapy. Refractoriness to induction immunosuppressive therapy and relapse of anemia may be risk factors for death in idiopathic, thymoma-associated and large granular lymphocyte leukemia-associated PRCA. The major causes of death are infections and organ failure. Standard treatment options for refractory and relapsed PRCA patients and the immunopathophysiology of acquired chronic PRCA merit further research.

Primary acquired chronic pure red cell aplasia refractory to standard treatments: remission with rituximab.

TO THE EDITOR: Pure red cell aplasia (PRCA) is a rare syndrome caused by erythropoietic hypoplasia in the absence of leukocytopenia and thrombocytopenia. It is characterized by severe normocytic and reticulocytopenic anemia, with a normally cellular bone marrow (BM) but devoid of erythroblasts [1]. The acquired form of PRCA is a chronic illness that is often diagnosed in conjunction with a variety of diseases [1], such as lymphoproliferative disorders [2], viral infections, autoimmune hemolytic anemia (AIHA) [3], rheumatologic disorders [4], and allogeneic stem cell transplantation [5]. However, this disorder is rarely diagnosed as an idiopathic condition. Acquired PRCA is managed as an immunologically mediated disease, using immunosuppressive therapy (IST) with corticosteroids and cyclosporine A (CSA) as the treatments of first choice [1]. As alternative and salvage treatment, rituximab has been reported to be highly effective [2,3,4,5]; however, to the best of our knowledge, no case of idiopathic PRCA managed with this agent has been reported. A 63-year-old woman was diagnosed in June 2003 as having PRCA after the discovery of isolated normocytic and reticulocytopenic anemia, the course of which had been insidious and progressive. All other possible underlying causes of erythroblastopenia were ruled out by appropriate investigations (Table 1); other laboratory and radiological evaluations revealed no abnormal findings. The patient had required transfusions of almost 2 units of packed red blood cells (RBC) every 2 to 3 weeks. Once the diagnosis was made, she was started on CSA plus corticosteroids, and soon achieved full recovery from BM erythropoiesis and attained normalization of peripheral blood counts. The patient no longer required transfusions. This was considered complete remission (CR) of PRCA. Therefore, the dosage of CSA was gradually reduced and discontinued. However, there was a progressive loss of response, and CSA was resumed in February 2007 due to a full relapse. The patient achieved a second CR, and the dosage of CSA was carefully tapered. However, the patient experienced progressive chronic renal failure (CRF) in January 2011, which fully resolved after discontinuation of CSA. PRCA recurred soon after, and the patient again required frequent RBC transfusions. When she required approximately 4 RBC units per month, steroids were retried, but without any benefit. Azathioprine was tried without any response. By May 2013, the need for transfusions had reached about 6 RBC units/month, and direct and indirect Coombs blood compatibility tests became positive with the appearance of anti-IgG autoantibody; therefore, the availability of required transfusions became a concern due the difficulty of finding compatible RBC units. In view of this life-threatening complication and a severe hematological condition refractory to all standard treatments for PRCA [1], rituximab was started as a salvage measure at a dose of 375 mg/m2/week for a total of 4 cycles, after the patient gave properly informed consent. Given the lack of experience with rituximab in the setting of primary PRCA, this treatment schedule was derived from that reported as safe and effective in patients with PRCA secondary to lymphoproliferative disorders [2]. After the second dose of rituximab, she exhibited a striking rise in her reticulocyte count and an increase in hemoglobin level. Thus, a third CR was achieved and was maintained during the subsequent 24 months (Fig. 1). In conclusion, the present letter describes a case of acquired chronic primary PRCA of idiopathic origin, refractory to standard measures, with remission on rituximab salvage treatment. To the best of our knowledge, this is the first report of idiopathic PRCA managed with rituximab. In our experience, this agent provided important clinical benefits in an elderly patient with a 12-year PRCA history, during which she had become intolerant/unresponsive to the majority of immunosuppressive agents used in this difficult-to-treat disorder. The course of disease and repeated responses to IST pointed to a possible underlying autoimmune pathogenesis. Although this hypothesis has not been proven by extensive clinical and laboratory evaluations; for this case, rituximab was highly effective in inducing long-lasting remission without any adverse effects.

Diagnosis and management of pure red cell aplasia

Pure red cell aplasia (PRCA) is a type of bone marrow failure syndrome (stem cell failure) and is characterized by severe normocytic, normochromic anemia associated with reticulocytopenia, and the absence of erythroblasts in otherwise normal bone marrow. PRCA may be congenital or acquired, and the acquired form of chronic PRCA may present as a primary hematological disease in the absence of any other diseases or secondary to thymoma, lymphoproliferative disorders, infections, and collagen vascular diseases or after exposure to various drugs or chemicals. The most common types of acquired chronic PRCA in Japan are idiopathic, thymoma-associated and lymphoid neoplasm-associated PRCA. Initial treatment of PRCA includes the cessation of potentially deleterious drugs and careful observation for one month while making efforts to identify the cause of PRCA. Idiopathic PRCA and secondary PRCA refractory to treatment of the underlying diseases are generally treated as immune-mediated disorders. Most chronic PRCA patients successfully treated with immunosuppressants require maintenance immunosuppressive therapy. Thus, identifying the cause of PRCA is crucial for the optimal management of this disorder.

Long-term outcome of patients with acquired chronic pure red cell aplasia (PRCA) following immunosuppressive therapy: a final report of the nationwide cohort study in 2004/2006 by the Japan PRCA collaborative study group.

Immunosuppressive therapy has been employed as the initial treatment for acquired chronic pure red cell aplasia (PRCA), such as idiopathic, thymoma-associated, or large granular lymphocyte (LGL) leukaemia-associated PRCA, which is thought to be immune-mediated. To explore the overall long-term outcome following immunosuppression and to identify the risk factors for death in these disorders, we conducted nationwide surveys in Japan 2004 and 2006, and identified a total of 185 patients with acquired chronic PRCA, including 72 idiopathic, 41 thymoma-associated and 14 LGL leukaemia-associated cases of PRCA for whom data was available. The present study evaluated 127 patients with these three subsets of PRCA. The median overall survival has not yet been reached in idiopathic PRCA. The estimated median overall survival times in patients with thymoma-associated and LGL leukaemia-associated PRCA were 142·1 and 147·8months, respectively. Twenty-two deaths were reported, and the response to induction therapy and relapse of anaemia were found to be associated with death. The major causes of death were infection in seven patients and organ failure in another seven patients. The results suggest that maintenance therapy and the management of infectious complications are crucial for improving the prognosis of chronic PRCA.

A Profound Decrease in FoxP3+Helios+CD4+ T Cells in a Subset of Patients with Acquired Aplastic Anemia and Pure Red Cell Aplasia: A Common Mechanism Underlying a Dependency on Cyclosporine

Background: A subset of patients with acquired aplastic anemia (AA) require low-dose cyclosporine (CsA) to maintain good hematopoietic function even after achieving a remission with immunosuppressive therapy (IST). Such CsA-dependent AA offers opportunities to clarify the immune mechanisms underlying bone marrow (BM) failure, but little is known about the mechanism(s) underlying this condition except for its close association with a particular HLA-class II allele DRB1*15:01. Regulatory T cells (Tregs), defined as CD4+CD25highFoxP3+ T cells, have been shown to be decreased in patients with severe AA at diagnosis and to recover to normal levels in association with hematological recovery after successful IST. An insufficient Treg recovery may account for dependency on CsA in a particular subset of AA patients, and also in patients with idiopathic pure red cell aplasia (PRCA), another typical immune-mediated BM failure of which remission is known to depend on CsA in virtually all patients. To test this hypothesis, we determined the percentage and absolute number of Tregs, as well as Th1, Th2 and Th17 cells, in various types of immune-mediated BM failure, using an anti-FoxP3 antibody (Ab) and anti-Helios Ab that allows enumeration of the functional Tregs more precisely than conventional methods. Methods: Different CD4+ Treg subsets were enumerated using a multicolor flow cytometry assay. Peripheral blood mononuclear cells (PBMCs) were stimulated by phorbol myristate acetate (PMA) and ionomycin in the presence of monensin for four hours, then were subjected to flow cytometry to determine the percentage of CD4+ T cells positive for cytoplasmic interferon-γ (Th1), interleukin 4 (Th2) and interleukin 17 (Th17). CsA-dependent AA was defined as AA with two or more episodes of relapse of pancytopenia that occurred during more than one year of therapy, followed by a good response to CsA. All six patients with CsA-dependent AA had normal blood cell counts and HLA-DR15 (DRB1*15:01 in five and DRB1*15:02 in one). Results: The normal ranges of each T cell subset percentage and the absolute numbers determined in 47 healthy volunteers (HVs) were: FoxP3+Helios+ Tregs 6.5±1.6%/ 47.7±13.8/µl, Th1 22.3±6.6%/ 151.2± 45.5/µl, Th2 5.5±1.6%/ 38.3±170.9/µl and Th17 1.7± 0.7%/ 11.2± 4.7/µl. The expression of FoxP3 by CD4+ T cells decreased with time after sampling, and this assay therefore necessitated analyses within 16 hours after sampling using fresh PBMCs. Compared to healthy volunteers, the percentage and absolute number of FoxP3+Helios+ Treg cells were significantly lower in the five patients with severe AA at diagnosis (3.7±1.1%, 22.7±6.8/µl) and in six CsA-dependent AA patients (3.4±0.8%, 13.8±12.3/µl) (Figure). In contrast, the Treg percentages were comparable to those of the HVs in four AA patients on CsA in convalescence (5.8±1.0%) and nine patients in remission off CsA (6.3±1.1%) while the absolute numbers of FoxP3+Helios+Tregs (27.3±19.2/µl, 32.3±13.3/µl) in these groups were lower than those of the HVs. On the other hand, the percentages of all other CD4+ T cell subsets were comparable among the four different groups and HVs. The apparent decrease in FoxP3+Helios+ Tregs despite normal blood cell counts in patients with CsA-dependent AA prompted us to screen nine patients with idiopathic PRCA whose erythropoietic functions were all dependent on CsA. The FoxP3+Helios+ Tregs were decreased in both the percentage and absolute count (3.1±0.5%, 12.1±5.9/µl) in all of these patients, including two patients whose PBMCs were examined before CsA therapy. As a result, the ratios of Th1/Treg and Th17/Treg were markedly higher in newly-diagnosed AA (12.5±3.4, 0.9±0.8), CsA-dependent AA (12.6±12.1, 1.1±1.5) and PRCA (10.6±7.7, 0.7±0.3) patients than in AA patients in convalescence (6.1±4.3, 0.4±0.1) and those in remission (3.8±2.3, 0.4±0.2), which were comparable to the HVs Th1/Treg and Th17/Treg ratios (3.3±0.9, 0.3±0.2). Conclusions: Decreased percentages and absolute counts of Tregs, as well as increased Th1/Treg and Th17/Treg ratios, are common features in patients with CsA-dependent AA and PRCA. A persistent decrease in Tregs may serve as a good marker for predicting AA relapse in patients responding to CsA. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Pure Red Cell Aplasia in a Patient with Cri-Du-Chat

Background: The Cri-Du-Chat syndrome also known as 5p-syndrome is a rare genetic autosomal disorder with the characteristic deletion of the short arm (p) of chromosome 5. To date, hematologic manifestationscharacteristic of BM failure have not been linked to this syndrome. Aims: To evaluate the safety and efficacy of cyclosporine to treat pure red cell aplasia in the clinical setting of 5p -syndrome. Presentation of Case:We report here a patient with 5p -syndrome who subsequently developed idiopathic pure red cell aplasia. He was treated with cyclosporine 1mg/kg twice a day, and achieved a complete remission, with no toxicities. Conclusion: This case suggests that cyclosporine can be used safely and effectively in

Burst-Forming Unit-Erythroid Assays (BFU-E) Can Help Distinguish Cellular Bone Marrow Failure Disorders

AbstractAbstract 3475Cytopenias in the context of cellular marrows cause significant morbidity for patients and are challenging for clinicians to diagnose and treat. Often the differential diagnosis includes pure red cell aplasia (PRCA), large granular lymphocyte leukemia (LGL), marrow suppression associated with systemic inflammation, and clonal bone marrow failure disorders such as myelodysplastic syndrome (MDS) or paroxysmal nocturnal hemoglobinuria (PNH). Unfortunately, it can be difficult to differentiate between these possibilities, and correct treatment depends on accurate diagnosis.The diagnostic algorithm for these patients includes multiple studies: cytogenetics, fluorescence in situ hybridization, T cell gene rearrangements, and flow cytometry. These multiple tests rarely provide a definitive diagnosis and do not consistently predict response to therapy. Functional tests of effective hematopoiesis are lacking. Colony growth assays quantifying BFU-E in bursts per 105 marrow mononuclear nuclear cells (MMNC) have been shown to predict response to immunosuppressive therapy (IST) for the bone marrow failure disorder of PRCA. (Charles et al. Blood 1996) However, these data were from an era before it was recognized that PRCA had several etiologies, one of which was LGL.We hypothesize that colony growth assays might provide diagnostic and therapeutic information in the evaluation of patients with bone marrow failure disorders. As part of an IRB-approved study, we performed a retrospective analysis on patients presenting for evaluation of cytopenias in the context of cellular marrows from January 2007-June 2012. Inclusion criteria were a recorded BFU-E assay performed prior to therapy, established diagnosis for the cytopenias and at least 6 months of follow up after treatment initiated. Response was defined as transfusion independence and ANC >1000/μL. For comparison, BFU-E in healthy controls was used.Thirty-six patients meeting the above criteria had BFU-E assessed over the study. All patients’ marrows were cellular or hypercellular for age, with a median cellularity of 60% (range 20–100). The diagnoses included 13 patients with LGL (8 of which had red cell aplasia only); 9 patients with PRCA (non-LGL in etiology); 8 patients with MDS; and 6 patients with cytopenias from systemic inflammation. The median follow up was 10.5 months (range 1–42). There were 8 deaths. There were 25 normal controls for comparison of BFU-E growth. The median growth in the normal controls was 40.3 BFU-E/105 MMNC (range 23.7–87).The eight patients with MDS had the lowest median growth of all categories at 5 BFU-E/105 MMNC (range 0–38). Four of these patients with MDS died from progression to acute leukemia in less than 9 months after initiation of therapy. The 4 patients still alive remain red cell transfusion dependent despite therapy with demethylating agents. Two have increased blasts on restaging marrows.The 13 patients with LGL grew a median of 61.3 BFU-E/105 MMNC (range 24.7–101). Nine patients (69%) responded to first or second line IST with durable remissions of 9 months or greater. One of the nonresponders died at 31 months from infection after multiple relapses and treatments.The 9 non-LGL PRCA patients grew a median of 23 BFU-E/105 MMNC (range 4.5–49). Only three patients (33%) of these patients responded to IST. The etiologies of the PRCA included 2 associated with thymoma, 2 with anti-erythropoietin antibodies (2 deaths at 1 and 11 months), 2 with ABO-incompatible transplants, and 3 idiopathic PRCA patients (1 death at 15 months).The 6 patients with cytopenias from systemic inflammation had documented autoimmunity (4 with Evan’s syndrome and 2 with systemic lupus erythematosus) and grew a median of 105.5 BFU-E/105 MMNC (range 33–137). All 6 (100%) patients responded to IST.In conclusion, patients with cellular marrows and cytopenias can be distinguished by BFU-E growth as seen in Figure 1. MDS patients have very low to undetectable growth. LGL and PRCA associated with LGL have high BFU-E growth, similar to other autoimmune processes, whereas non-LGL PRCA has low BFU-E growth. BFU-E predicts for response to IST in PRCA, as it seems to predict for LGL. In this cohort, nonmalignant diseases are well distinguished from MDS by BFU-E growth. Prospective studies of colony growth assays as part of the diagnostic algorithm for bone marrow failure disorders appear warranted. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Expansion of CD8+/perforin+ T-Cell Subset in Patients Predicts Response to Cyclosporin A Therapy in Patients with Erythroid Hypoplasia/Aplasia

Abstract 1337Erythroid hypoplasia or aplasia is a hematological condition observed in including idiopathic pure red cell aplasia (PRCA), thymoma-associated PRCA and aplastic anemia. Myelodysplastic syndrome (MDS) with erythroid hypoplasia/aplasia in bone marrow is a rare type of MDS that was not included in existing classifications of MDS. Patients with erythroid hypoplasia/aplasia have common characteristics; transfusion dependencies, immunologic abnormalities and successful immunosuppressive therapies with cyclosporin A (CsA). Thus, we may regard erythroid hypoplasia/aplasia as one of hematological disease entities. However, pathogenic mechanisms of erythroid hypoplasia/aplasia have not been fully elucidated, although T-lymphocyte-mediated inhibition of erythropoiesis is suspected to be the most possible mechanism of the pathogenesis. Recently, we reported that oligoclonal expansion of CD8+/perforin+ T cells was observed in patients with thymoma-associated PRCA and the oligoclonality was exclusively detected in CD8+ T cells, but not CD4+ T cells. To clarify the pathogenetic role of the T-cells, we analyzed the T-cell subsets and therapeutic responses in patients with erythroid hypoplasia/aplasia in bone marrow.Among 253 patients with MDS diagnosed at Hiroshima University Hospital between 2000 and June 2011, 12 patients (4.7%) showed erythroid hypoplasia/aplasia. A total of 22 patients with erythroid hypoplasia/aplasia, including 8 MDS with erythroid hypoplasia/aplasia, 3 idiopathic PRCA, 3 thymoma-associated PRCA and 8 aplastic anemia, were enrolled in this study. All patients were treated with CsA and improvement in anemia in this study followed the International Working Group (IWG) 2006 criteria. For T-cell subset analysis, mononuclear cells (MNCs) were purified from bone marrow (BM) or peripheral blood (PB) of the patients. MNCs were stained with fluorescent (FITC, PE, PerCP or APC)-conjugated antibodies for CD8, perforin, CCR7, CD62L, CD27, CD28 and CD45RO, CD45RA and were subjected to flow cytometric analysis. As controls, 30 patients with MDS without erythroid hypoplasia/aplasia and 30 patients without BM abnormalities were also analyzed.Among 22 patients with erythroid hypoplasia/aplasia, 10 patients (4 MDS with erythroid hypoplasia/aplasia, 1 idiopathic PRCA, 3 thymoma-associated PRCA and 2 aplastic anemia) responded to CsA therapy within 2 to 8 weeks. The median blood hemoglobin concentration increased from 6.5 g/dL at the baseline to 9.3 g/dL with treatment, with a median increase of hemoglobin of 2.8 g/dL from the baseline. We attempted to compare the T-cell subsets between CsA-responders and non-responders. All of 3 thymoma-associated PRCA showed good response to CsA therapy, suggesting that the oligoclonal expansion of a CD8+/perforin+ T-cell subset may be associated with the responses to immunosuppressive therapy. Thus, we focused on a T-cell subpopulation expressing CD8+/perforin+. Intriguingly, the CD8+/perforin+ T cells were significantly increased in the CsA-responders (44.3 ± 9.6%, n=10) compared to the non-responders (19.0 ± 9.3%, n=12, P<0.0001), normal BM controls (16.9 ± 7.0%, n=30) and MDS without erythroid hypoplasia/aplasia (15.1 ± 7.0%, n=30). Among the CD8+/perforin+ T cells, CD27+/CD62L+/−/CCR7low/CD28low/CD45RA++/CD45RO+ population was prominent, which is consistent with an effector memory T (TEM) cell subset described by Decrion et al.Our study reveals that CD8+/perforin+ T cell subset is a large population in the patients with CsA-responsive erythroid hypoplasia/aplasia. It is suggested that CD8+/perforin+ T cell subset may have functions to reduce erythroid progenitors via immunological mechanisms. The mechanisms may be easily suppressed by immunosuppressive therapies. In conclusion, expansion of CD8+/perforin+ T cell subset predicts response to cyclosporin A therapy in patients with erythroid hypoplasia/aplasia. The disease entity of “erythroid hypoplasia/aplasia in bone marrow with expansion of CD8+/perforin+ T cell subset”, including MDS, PRCA with or without thymoma and aplastic anemia, may have common pathogenetic mechanisms. Disclosures:No relevant conflicts of interest to declare.

Clinical use of cyclosporine as an adjunctive therapy in the management of feline idiopathic pure red cell aplasia

The clinical use of cyclosporine is described in a group of client-owned cats diagnosed with idiopathic pure red cell aplasia (PRCA). All 10 cats were treated with combinations of glucocorticoids and cyclosporine. Of the 10 cats, the eight for which follow-up data was available achieved and maintained remission for a median of 31 and 406 days, respectively. Therapy was reduced or discontinued in 7/8 cats; 2/7 maintained remission off therapy and 5/7 cats relapsed. Remission was reinduced in four cats, with 3/4 cats maintained long-term on low dose therapy. Adverse effects associated with cyclosporine therapy were responsive to dose reduction or drug withdrawal. Feline idiopathic PRCA was responsive to combination immunosuppressive therapy with glucocorticoids and cyclosporine. Relapse was common, particularly after drug discontinuation; therefore, most cats required maintenance long-term low dose therapy.

Acquired pure red cell aplasia associated with malignant lymphomas: A nationwide cohort study in Japan for the PRCA Collaborative Study Group

Pure red cell aplasia (PRCA) has been reported in association with lymphoma as one of the autoimmune diseases seen during the course of lymphoid malignancies. However, the relation of PRCA with the underlying lymphomas remains unclear. The aim of this study was to clarify the histologic subtypes of lymphomas, the chronological sequence of anemia and lymphoma, and the response to treatment. We conducted a nationwide survey in Japan. From a cohort of 185 PRCA patients, 8 patients with lymphoma were evaluated. Histologic subtypes varied and the lymphoma was of the B-cell type in four cases and of the T-cell type in four. Four patients simultaneously developed PRCA and lymphoma. Three patients developed PRCA following lymphoma, two of whom developed anemia during remission of lymphoma. PRCA preceded lymphoma in one patient. Effective chemotherapy was associated with remission of anemia in concurrent lymphoma and PRCA. Overall, anemia responded to chemotherapy and/or immunosuppressive therapy in seven patients. In four responding patients, PRCA remained in durable remission without maintenance immunosuppressive therapy, which is different from a recurrent feature of idiopathic PRCA. We suggest that the mechanism of lymphoma-associated PRCA is heterogeneous and that durable maintenance-free remission of anemia can be obtained in some patients.