Paroxysmal Nocturnal Hemoglobinuria and Cancer: High Incidence Of Cancer In a Large Series Of PNH Patients In a Single Center

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4871-4871 ◽  
Author(s):  
Cristina Muñoz-Linares ◽  
Emilio Ojeda ◽  
Rafael Fores ◽  
Martin Cabero ◽  
Daniel Morillo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Bone Marrow Failure ◽  
Bone Marrow Failure Syndrome ◽  
Final Cause ◽  
Incidence Of Cancer ◽  
Hematological Cancers ◽  
High Incidence ◽  
Allogeneic Hematopoietic Transplantation

Abstract Since 1964, a total of 56 patients with Paroxysmal Nocturnal Hemoglobinuria clone (PNH) were evaluated in our Hematology Unit. The PNH was evaluated with Ham’s and/or sucrose tests until 1993, when the firsts cytometric analysis of PNH were performed in our Laboratory with CD55 and CD59 markers on granulocytes mainly, and since 2011 also with the FLAER technique. According with PNH Parker´s Classification, most of the patients were Classical PNH type (28 patients), and the remaining included in the other subsets such as 21 PNH in the setting of another bone marrow failure syndrome (BMFS) and 7 PNH subclinical. Most of the patients (70%) displayed an Aplastic Anemia (AA) before or concomitantly with the diagnosis of PNH, and received immunosuppressant drugs (Steroids with/out antithymocyte globulin & Cyclosporine). In four patients an Allogeneic Hematopoietic Transplantation was performed due to a Severe Aplastic Anemia (2 patients), a Classical Severe PNH (before Eculizumab era) or a Myelodysplastic Syndrome. Another patient received a Liver Transplantation because of advanced Hepatitis C related liver failure. In our PNH series, an unexpected high incidence of cancer has appeared, with 8 patients (14,5%) displaying different hematological or non-hematological cancers in the lasts years:SexAge Diagnosis PNHParker’s ClassificationYear Diagnosis PNHYear Diagnosis CancerPrevious ImmunosuppressionCancerYear Death♂16Classical19692011YesLymphoma2011AA & Liver Tx♀30Classical19732003NonePancreatic2006♂38Classical19741995NoneGastric20122012Pulmonary♂26Classical19892013Yes, SteroidsCerebralAlive♀25Classical19942005YesLymphoma2006AA & Cord-Blood Tx♂40BMFS19951995Yes, SteroidsLiver1995♂75BMFS20112009NoneSeminoma2012*♂56Subclinical20101999Yes, SteroidsProstaticAlive*Dead because bone marrow failure. In our PNH series, cancer reports as one of the most frequent final cause of death, with thrombosis with similar incidence (11 patients of the 56 are dead, with 5 patients dead because thrombosis), although the high incidence and severity of thrombosis episodes in this cohort of patients (20 patients experienced thrombosis with a total of 40 events). As displayed supra, some of the cancers could be attributed to the therapy applied in particular patients: The two secondary lymphomas after organ transplantation could be explained by the immunosuppression employed in these procedures. Only three patients did not received immunosuppressant drugs before cancer diagnosis. This high mortality cancer rate precludes the indiscriminate use of steroids in PNH patients. This result, never reported before in PNH, merits an investigational survey of cancer incidence in PNH patients in the PNH International Registry. Disclosures: Ojeda: Alexion Pharmaceuticals: Consultancy, Speakers Bureau.

Molecular Classification of Bone Marrow Failure Syndromes: Protein Signatures As Surrogate Biomarker for Accurate Diagnosis of Severe Aplastic Anemia, Hypoplastic Myelodysplastic Syndrome and Paroxysmal Nocturnal Hemoglobinuria Patients

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2414-2414
Author(s):  
Ayodele Alaiya ◽  
Hazza A Alzahrani ◽  
Zakia Shinwari ◽  
Tarek Owaidah ◽  
Fahad Al Mohareb ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myelodysplastic Syndrome ◽  
Aplastic Anemia ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Bone Marrow Failure ◽  
Accurate Diagnosis ◽  
Bone Marrow Failure Syndrome ◽  
Bone Marrow Failure Syndromes ◽  
Disease Specific

Abstract Background/Purpose: Bone marrow failure syndrome is an example of disease entity where accurate diagnosis of Severe Aplastic Anemia (SAA), Paroxysmal Nocturnal Hemoglobinuria (PNH) and Hypoplastic Myelodysplastic Syndrome (MDS) is very challenging. The aim of this study was to identify panels of disease-specific /disease-associated proteins biomarkers to be used for more objective diagnosis and better prediction of disease prognosis of patients presenting with features of bone marrow failure syndromes. Methodology: Bone marrow plasma (MBP) and peripheral blood plasma (PBP) samples from 20 patients with bone marrow hypoplasia; including AA/MDS/PNH were subjected to expression proteome analysis using label-free quantitative liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Results: Approximately 300 unique protein species were identified of which 107 and 218 were significantly differentially expressed (> 2- ∞- fold change & p < 0.05) in BMP and PBP respectively. These protein fingerprints independently discriminates patients into three distinct clusters; AA/MDS/PNH. Furthermore, only approx. 25% of the proteins were common between the two datasets from BMP and PBP. Some of the identified proteins were filtered and mapped using Ingenuity Pathway Analysis, and were associated with five different networks. The top two of these networks involved cell-to-cell signaling interaction, hematological system development and function, and immune cell trafficking. Only three of the differentially expressed proteins were uniquely expressed in SAA and MDS but absent in PNH, thus making these proteins potential biomarkers. The probable diagnostic utility of these proteins would be validated in large archival clinical samples. Our data indicates the utility of multivariate analysis of quantitative proteome data as a means of discovery of disease related or disease specific biomarkers for bone marrow syndromes. Conclusions: We have identified protein signatures capable of objective classification of bone marrow failure syndromes patients. Our expression proteomics strategy is very promising for identification of clinically useful biomarkers. These proteins once validated, on a larger cohort of patients, might be valuable to complement the currently existing parameters for reliable and objective disease diagnosis, monitoring treatment response and clinical outcome of bone marrow failure syndrome patients. Disclosures Owaidah: King abdulaziz city for science, Novo Nordisk, Bayer: Honoraria, Research Funding.

Hepatitis-associated Aplastic Anemia Presenting as a Familial Bone Marrow Failure Syndrome

2009 ◽  
Vol 31 (11) ◽  
pp. 884-887 ◽  
Author(s):  
Vicky Rowena Breakey ◽  
Stephen Meyn ◽  
Vicky Ng ◽  
Christopher Allen ◽  
Inderjeet Dokal ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Bone Marrow Failure ◽  
Bone Marrow Failure Syndrome

scholarly journals Aplastic anemia: immunosuppressive therapy in 2010

2011 ◽  
Vol 3 (2s) ◽  
pp. 7 ◽  
Author(s):  
Antonio M. Risitano ◽  
Fabiana Perna
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
State Of The Art ◽  
Bone Marrow Failure ◽  
Standards Of Care ◽  
Bone Marrow Failure Syndrome ◽  
Clinical Observations ◽  
Immune Mediated ◽  
Experimental Works ◽  
Current Standards

Acquired aplastic anemia (AA) is the typical bone marrow failure syndrome characterized by an empty bone marrow; an immune-mediated pathophysiology has been demonstrated by experimental works as well as by clinical observations. Immunusuppressive therapy (IST) is a key treatment strategy for aplastic anemia; since 20 years the standard IST for AA patients has been anti-thymocyte globuline (ATG) plus cyclosporine A (CyA), which results in response rates ranging between 50% and 70%, and even higher overall survival. However, primary and secondary failures after IST remain frequent, and to date all attempts aiming to overcome this problem have been unfruitful. Here we review the state of the art of IST for AA in 2010, focusing on possible strategies to improve current treatments. We also discuss very recent data which question the equality of different ATG preparations, leading to a possible reconsideration of the current standards of care for AA patients.

Bone Marrow Histology in Patients with Paroxysmal Nocturnal Hemoglobinuria.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4215-4215
Author(s):  
Sandra van Bijnen ◽  
Konnie Hebeda ◽  
Petra Muus
Keyword(s):  
Bone Marrow ◽  
Mast Cells ◽  
Aplastic Anemia ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Plasma Cells ◽  
Bone Marrow Failure ◽  
Clone Size ◽  
Immune Mediated ◽  
Lymphoid Nodules ◽  
Pnh Clone

Abstract Abstract 4215 Introduction Paroxysmal Nocturnal Hemoglobinuria (PNH) is a disease of the hematopoietic stem cell (HSC) resulting in a clone of hematopoietic cells deficient in glycosyl phosphatidyl inositol anchored proteins. The clinical spectrum of PNH is highly variable with classical hemolytic PNH at one end, and PNH in association with aplastic anemia (AA/PNH) or other bone marrow failure states at the other end. It is still largely unknown what is causing these highly variable clinical presentations. Immune-mediated marrow failure has been suggested to contribute to the development of a PNH clone by selective damage to normal HSC. However, in classic PNH patients with no or only mild cytopenias, a role for immune mediated marrow failure is less obvious. No series of trephine biopsies has been previously documented of patients with PNH and AA/PNH to investigate the similarities and differences in these patients. Methods We have reviewed a series of trephine biopsies of 41 PNH patients at the time the PNH clone was first detected. The histology was compared of 27 patients with aplastic anemia and a PNH clone was compared to that of 14 patients with classic PNH. Age related cellularity, the ratio between myeloid and erythroid cells (ME ratio), and the presence of inflammatory cells (mast cells, lymphoid nodules and plasma cells) were evaluated. The relation with clinical and other laboratory parameters of PNH was established. Results Classic PNH patients showed a normal or hypercellular marrow in 79% of patients, whereas all AA/PNH patients showed a hypocellular marrow. Interestingly, a decreased myelopoiesis was observed not only in AA/PNH patients but also in 93% of classic PNH patients, despite normal absolute neutrophil counts (ANC ≥ 1,5 × 109/l) in 79% of these patients. The number of megakaryocytes was decreased in 29% of classic PNH patients although thrombocytopenia (< 150 × 109/l) was only present in 14% of the patients. Median PNH granulocyte clone size was 70% (range 8-95%) in classic PNH patients, whereas in AA/PNH patients this was only 10% (range 0.5-90%). PNH clones below 5% were exclusively detected in the AA/PNH group. Clinical or laboratory evidence of hemolysis was present in all classical PNH patients and in 52% of AA/PNH patients and correlated with PNH granulocyte clone size. Bone marrow iron stores were decreased in 71% of classic PNH patients. In contrast, increased iron stores were present in 63% of AA/PNH patients, probably reflecting their transfusion history. AA/PNH patients showed increased plasma cells in 15% of patients and lymphoid nodules in 37%, versus 0% and 11% in classic PNH. Increased mast cells (>2/high power field) were three times more frequent in AA/PNH (67%) than in PNH (21%). Conclusion Classic PNH patients were characterized by a more cellular bone marrow, increased erythropoiesis, larger PNH clones and clinically by less pronounced or absent peripheral cytopenias and more overt hemolysis. Decreased myelopoiesis and/or megakaryopoiesis was observed in both AA/PNH and classic PNH patients, even in the presence of normal peripheral blood counts, suggesting a role for bone marrow failure in classic PNH as well. More prominent inflammatory infiltrates were observed in AA/PNH patients compared to classical PNH patients. Disclosures: No relevant conflicts of interest to declare.

Screening Patients with Myelodysplastic Syndrome and Aplastic Anemia for Paroxysmal Nocturnal Hemoglobinuria Clones: A Retrospective Study,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3426-3426 ◽  
Author(s):  
Andrew Shih ◽  
Ian H. Chin-Yee ◽  
Ben Hedley ◽  
Mike Keeney ◽  
Richard A. Wells ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Retrospective Study ◽  
Aplastic Anemia ◽  
Board Of Directors ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Bone Marrow Failure ◽  
Cell Surface Expression ◽  
Advisory Committees ◽  
Hematopoietic Stem ◽  
Pnh Clone

Abstract Abstract 3426 Introduction: Paroxysmal Nocturnal Hemoglobinuria (PNH) is a rare disorder due to a somatic mutation in the hematopoietic stem cell. The introduction of highly sensitive flow cytometric and aerolysin testing have shown the presence of PNH clones in patients with a variety of other hematological disorders such as aplastic anemia (AA) and myelodysplasic syndrome (MDS). It is hypothesized that patients with these disorders and PNH clones may share an immunologic basis for marrow failure with relative protection of the PNH clone, due to their lack of cell surface expression of immune accessory proteins. This is supported by the literature showing responsiveness in AA and MDS to immunosuppressive treatments. Preliminary results from a recent multicenter trial, EXPLORE, notes that PNH clones can be seen in 70% of AA and 55% of MDS patients, and therefore there may be utility in the general screening of all patients with bone marrow failure (BMF) syndromes. Furthermore, it has been suggested that the presence of PNH cells in MDS is a predictive biomarker that is clinically important for response to immunosuppressive therapy. Methods: Our retrospective cohort study in a tertiary care center used a high sensitivity RBC and FLAER assay to detect PNH clones as small as 0.01%. Of all patients screened with this method, those with bone marrow biopsy and aspirate proven MDS, AA, or other BMF syndromes (defined as unexplained cytopenias) were analysed. Results from PNH assays were compared to other clinical and laboratory parameters such as LDH. Results: Overall, 102 patients were initially screened over a 12 month period at our center. 30 patients were excluded as they did not have biopsy or aspirate proven MDS, AA, or other BMF syndromes. Of the remaining 72 patients, four patients were found to have PNH clones, where 2/51 had MDS (both RCMD, IPSS 0) [3.92%] and 2/4 had AA [50%]. The PNH clone sizes of these four patients were 0.01%, 0.01%, 0.02%, and 1.7%. None of the MDS patients with known recurrent karyotypic abnormalities had PNH clones present. Only one of the four patients had a markedly increased serum LDH level. Conclusions: Our retrospective study indicates much lower incidence of PNH clones in MDS patients or any patients with BMF syndromes when compared to the preliminary data from the EXPLORE trial. There is also significant disagreement in other smaller cohorts in regards to the incidence of PNH in AA and MDS. Screening for PNH clones in patients with bone marrow failure needs further study before adoption of widespread use. Disclosures: Keeney: Alexion Pharmaceuticals Canada Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees. Wells:Alexion Pharmaceuticals Canada Inc: Honoraria. Sutherland:Alexion Pharmaceuticals Canada Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Recombinant humanized anti-IL-2 receptor antibody (daclizumab) produces responses in patients with moderate aplastic anemia

Blood ◽  
2003 ◽  
Vol 102 (10) ◽  
pp. 3584-3586 ◽  
Author(s):  
Jaroslaw P. Maciejewski ◽  
Elaine M. Sloand ◽  
Olga Nunez ◽  
Carol Boss ◽  
Neal S. Young
Keyword(s):  
Monoclonal Antibody ◽  
Bone Marrow ◽  
Aplastic Anemia ◽  
Neutrophil Count ◽  
Bone Marrow Failure ◽  
Interleukin 2 ◽  
Immunosuppressive Agent ◽  
Solid Organ ◽  
Bone Marrow Failure Syndrome ◽  
Humanized Monoclonal Antibody

AbstractIn contrast to severe aplastic anemia (sAA), the appropriate management of patients with moderate pancytopenia is unclear. In this study, we examined the efficacy of a humanized monoclonal antibody recognizing interleukin-2 receptor (daclizumab), which has proven to be a successful immunosuppressive agent in solid organ and bone marrow transplantation. We treated 17 patients with moderate aplastic anemia (mAA) with 1 mg/kg every 2 weeks for 3 months. mAA was defined as depression of 2 of the 3 blood counts: absolute neutrophil count 1200/mm3 or less, platelet count 70 000/mm3 or less, hemoglobin level 8.5 g/dL or lower, and absolute reticulocyte count 60 000/mm3 or less. The primary end point of our protocol was a hematologic response in at least one affected peripheral blood value. Daclizumab had little toxicity. Six of the 16 (38%) evaluable patients responded to treatment. Two patients with previously chronic disease showed complete return of normal counts, which were sustained for more than 2 years following treatment. Four patients had single-lineage responses. Two previously transfusion-dependent patients became transfusion independent; one patient with many neutropenia-related infections had a normal neutrophil count following treatment. Daclizumab appears safe; its efficacy in this pilot protocol suggests that expanded study of this monoclonal antibody in immune-mediated bone marrow failure syndrome is warranted. (Blood. 2003; 102:3584-3586)

Mutations in the RNA Component of Telomerase, TERC, in Children with Aplastic Anemia and Myelodysplastic Syndrome: An NMDP Study.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3736-3736
Author(s):  
Joshua J. Field ◽  
Philip J. Mason ◽  
Yvonne J. Barnes ◽  
Allison A. King ◽  
Monica Bessler ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Myelodysplastic Syndrome ◽  
Aplastic Anemia ◽  
Stem Cell Transplant ◽  
Bone Marrow Failure ◽  
Juvenile Myelomonocytic Leukemia ◽  
Cell Transplant ◽  
Bone Marrow Failure Syndrome ◽  
Base Pair Deletion

Abstract Mutations in TERC, the RNA component of telomerase, result in autosomal dominant dyskeratosis congenita (DC), a rare bone marrow failure syndrome. DC is clinically heterogeneous and TERC mutations have been detected in a subset of patients previously diagnosed with idiopathic aplastic anemia (AA) and myelodysplastic syndrome (MDS). Unrecognized TERC mutations are clinically relevant as patients with DC respond poorly to immunotherapy and have an increased risk of complications following conventional conditioning for stem cell transplant (SCT). We aimed to determine the frequency of TERC mutations in pediatric patients with AA and MDS who require a SCT. We obtained 315 blood or bone marrow samples from the National Donor Marrow Program Registry from children under age 18 with bone marrow failure who underwent an unrelated stem cell transplant. We screened these samples for mutations in the TERC gene using direct DNA sequencing. To exclude polymorphisms, we also screened 537 racially diverse healthy controls. The study group was composed of patients with MDS (n=151), AA (n=123), and juvenile myelomonocytic leukemia (JMML) (n=41), which may be difficult to distinguish from MDS. The mean age at the time of transplant was 9 years. We found sequence alterations in the promoter region of TERC in 2 patients. A 2 base pair deletion (-240delCT) was identified in a 4 year-old child with MDS and a 1 year-old child with JMML was found to have a point mutation (-99C→G), which was identified previously in an 18 year-old patient with paroxysmal nocturnal hemoglobinuria and is known to affect the Sp1 binding site. The pathogenicity of this mutation is unclear. In summary, our findings suggest that screening for TERC gene mutations is unlikely to diagnose occult DC in children with severe bone marrow failure who require a stem cell transplant but have no clinical features or history to suggest a familial bone marrow failure syndrome.

Telomerase Gene Mutation Screening and Telomere Length Detection in Chinese Patients with Bone Marrow Failure Syndrome.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1047-1047
Author(s):  
Bing Han ◽  
Bo Liu ◽  
Yongqiang Zhao
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Small Group ◽  
Telomere Length ◽  
Gene Mutation ◽  
Bone Marrow Failure ◽  
Chinese Patients ◽  
Chinese People ◽  
Bone Marrow Failure Syndrome ◽  
Tert Mutation

Abstract Background Acquired bone marrow failure syndrome (BMF) is a group of diseases include aplastic anemia(AA), melodysplastic syndrome (MDS) and paraoxymal nocturnal hemoglobinuria (PNH). Some BMF patients have short telomeres in their peripheral nucleated cells. The length of telomere is maintained by a group of enzymes called telomerase complex. The core components of this complex are a RNA template and a reverse transcriptase, called TERC and TERT, respectively. Recently several studies in the west and Japan have disclosed the presence of telomerase complex gene mutation in a small group of patients with acquired bone marrow failure. They speculated that this small group of patients might represent a subset of cryptogenic Dyskeratosis Congenita (DKC), in which the premature exhaustion of hematopoietic reservoir is caused by mutations in the telomerase gene. This group of patients, though very small in number, would benefit from early bone marrow transplantation instead of traditional immunosuppressive therapy. The incidence of aplastic anemia in Chinese people is relatively high compared with that in the western country. But there has so far been no study in China about the incidence of telomerase gene mutation in acquired bone marrow failure and its relationship with telomere length. Objectives To study the incidence of telomerase gene (namely TERC and TERT ) mutation in Chinese patients with acquired bone marrow failure and explore its relationship with telomere shortening. Methods Blood samples from 90 patients with AA, MDS, and PNH in northern China were collected and performed TERC and TERT mutation analysis. Telomere length was measured by Southern blotting and compared with their normal counterparts. Results 2 TERC mutations (n37 A→G, reported previously ; n66G→C) and 2 TERT mutations (n1870G→T (E/*); n1780G→T (S/I) ) were identified in 90 BMF patients. Among them, 3 mutations are reported first time. 1 patient with TERT mutation, however, was finally diagnosed as DKC instead of acquired AA, making the incidence of telomerase gene mutation in Chinese people with acquired bone marrow failure 3.4%, similar to that of the western people. Southern Blot analysis showed the small group of patients carrying TERC and TERT mutations has very short telomeres, compared with normal controls and with their aplastic counterparts. Conclusions The incidence of telomerase gene mutation in Chinese people with acquired bone marrow failure is 3.4%, similar to that of the western people. This small group of patients has very short telomeres, it is thus clinically important to screen for this small group of patients.

scholarly journals Clinical approach to marrow failure

Hematology ◽  
2009 ◽  
Vol 2009 (1) ◽  
pp. 329-337 ◽  
Author(s):  
Akiko Shimamura
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Medical Management ◽  
Bone Marrow Failure ◽  
Clinical History ◽  
Clinical Approach ◽  
Careful Attention ◽  
Bone Marrow Failure Syndrome ◽  
Bone Marrow Failure Syndromes ◽  
Molecular Pathophysiology

Abstract The treatment and medical management of aplastic anemia fundamentally differ between patients with inherited versus acquired marrow failure; however, the diagnosis of an inherited bone marrow failure syndrome is frequently obscure. Recent exciting advances in our understanding of the molecular pathophysiology of the inherited bone marrow failure syndromes have resulted in a profusion of new tests to aid in diagnosis. This in turn has raised questions regarding the appropriate choice of testing for the patient presenting with aplastic anemia. Important clues to the diagnosis of an inherited marrow failure syndrome may be gleaned from careful attention to the clinical history, physical exam, and laboratory workup.

Development of Paroxysmal Nocturnal Hemoglobinuria in Children with Aplastic Anemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1499-1499 ◽  
Author(s):  
Atsushi Narita ◽  
Hideki Muramatsu ◽  
Yusuke Okuno ◽  
Yuko Sekiya ◽  
Kyogo Suzuki ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Paroxysmal Nocturnal Hemoglobinuria ◽  
Bone Marrow Failure ◽  
Subsequent Development ◽  
Poor Quality ◽  
Targeted Sequencing ◽  
Hematopoietic Stem ◽  
Bone Marrow Failure Syndromes

Abstract Introduction: Paroxysmal nocturnal hemoglobinuria (PNH) is a nonmalignant clonal disease of hematopoietic stem cells resulting from a somatic mutation in the PIGA gene. PNH frequently manifests in association with aplastic anemia (AA), in which PIGA mutations are believed to enable escape from the immune-mediated destruction by pathogenic T cells. Recent studies using next-generation sequencing have revealed that frequent somatic PIGA mutationsin AA patients are associated with a better response to IST and prognosis (Yoshizato et al N Engl J Med. 2015; 373: 35-47). However, clinical PNH is a progressive and life-threatening disease driven by chronic hemolysis that leads to thrombosis, renal impairment, poor quality of life, and death. Large studies in adults have reported that clinical PNH developed in 10%-25% of AA patients; however; the frequency of clinical PNH in children with AA has rarely been described. Here we aimed to elucidate the pathological link between PNH and AA in children. Methods: In total, 57 children (35 boys and 22 girls) diagnosed with acquired AA at our hospital between 1992 and 2010 were retrospectively studied. Patients who underwent hematopoietic stem cell transplantation as first-line treatment within 1 year after AA diagnosis and those with clinical PNH at AA diagnosis were excluded. Flow cytometry (FCM) was used to detect PNH CD13+/CD55−/CD59− granulocytes and PNH glycophorin A+/CD55−/CD59− red blood cells (RBCs). Clinical PNH was defined as the presence of intravascular hemolysis and ≥5% PNH granulocytes or PNH RBCs. Minor PNH clones were defined as those with >0.005% PNH granulocytes or >0.010% PNH RBCs. We performed targeted sequencing of bone marrow samples from patients with clinical PNH that were obtained at 2 time points: at AA diagnosis and after PNH development. The panel of 184 genes for targeted sequencing included most of the genes known to be mutated in inherited bone marrow failure syndromes and myeloid cancers, as well as PIGA. Results: The median patient age at AA diagnosis was 9.3 (1.2-17.8) years, and the median follow-up period was 123 (2-228) months. A total of 43 patients were screened for PNH clones by FCM after AA diagnosis, and 21 of these with minor PNH clones were identified. The median percentages of PNH granulocytes and PNH RBCs were 0.001% (0.000%-4.785%) and 0.000% (0.000%-3.829%), respectively. During follow-up, 5 patients developed clinical PNH after adolescence (15-22 years of age). The median time between AA diagnosis and PNH development was 4.9 (3.3-7.9) years. All clinical PNH patients were treated with IST for AA, and complete and partial response after 6 months were achieved in 1 and 4 patients, respectively. Gross hemoglobinuria was present in all clinical PNH patients, but thrombosis was not observed. The size of PNH clones varied greatly among patients: PNH granulocytes and PNH RBCs were 42.96% (10.04%-59.50%) and 48.87% (15.02%-90.80%), respectively. Oral cyclosporine A and intravenous eculizumab were administered to 3 and 1 patients, respectively; all patients showed sustained response as indicated by improvement in gross hemoglobinuria and normal blood counts after treatment. The remaining 1 patient underwent bone marrow transplantation from the HLA-identical mother and was alive without any complications. Overall, the 10-year probability of developing clinical PNH was 10.2% (95%CI, 3.6-20.7). Among 43 patients screened for PNH clones at AA diagnosis, the 10-year cumulative clinical PNH incidence was significantly higher in patients with minor PNH clones than in those without minor PNH clones at AA diagnosis [29% (95% CI, 10%-51%) vs. 0% (95% CI, 0%-0%); p = 0.015]. Among all clinical PNH patients, a total of 8 somatic PIGA mutations were detected (missense, 2; splice site, 2; and frameshift, 4). However, PIGA mutations were not detected at AA diagnosis even in patients who subsequently developed clinical PNH. Conclusion: In our cohort, the percentage of patients who eventually developed clinical PNH was comparable to that reported in adults in a previous study. Furthermore, the current study showed that the presence of minor PNH clones at AA diagnosis was a risk factor for the subsequent development of clinical PNH, although the clones were not detected by targeted sequencing. Thus, pediatric AA patients with PNH clones at AA diagnosis should undergo long-term periodic monitoring for potential clinical PNH development. Disclosures Kojima: SANOFI: Honoraria, Research Funding.

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