Progress in the congenital dyserythropoietic anemias: Juicy but high‐hanging fruits?
Authors: Raffaele Renella
Journal: American Journal of Hematology (2010)

In this issue of the American Journal of Hematology, Russo et al. [1] describe a large number of novel variants in SEC23B found in unrelated patients with congenital dyserythropoietic anemia (CDA) Type 2. They make an important contribution to the molecular diagnosis and to our understanding of the possible genotype–phenotype correlation in this condition. These findings add to our comprehension of the most prevalent type of CDA and are the fruit of many years' expert collection of patient samples via the International CDA-2 Registry in Naples, Italy (led by Achille Iolascon) and European collaborations. Because of the rarity of this disorder, a brief introduction and review of the recent progress in this field seems warranted. The model of combining advances in biochemistry, genetics, and molecular cell biology with clinical investigation recently led to the Lasker Prize being awarded to Sir David J. Weatherall [2]. This strategy has been widely applied in erythropoiesis research and clinical hematology and has clearly impacted patient care. Nowadays, it is safely possible to replace red blood cells and their progenitors; thanks to the knowledge generated in the fields of transfusion medicine and hematopoietic stem cell transplantation. Furthermore, targeted therapies have become available (i.e., eculizumab for paroxysmal nocturnal hemoglobinuria [3]) and even gene therapy has been proven a successful tool (in thalassemia [4]). Although the progress in red cell biology and its clinical applications has been impressive, it has not been universal. In fact, although molecular understanding of sickle cell disease started in the 1940s [5], and putative genes and proteins were investigated since the 1950s, only recently has research on inherited conditions of erythropoiesis started to shed some light on the pathogenesis of congenital dyserythropoietic anemias. This delay is largely due to the nature of the disease (reviewed in detail in Ref.6), which can be summarized as follows. From a clinical standpoint, the Congenital Dyserythropoietic Anemias (CDAs) were first recognized as separate entities by Crookston et al. [7], and subsequently formally described by Heimpel and Wendt in 1968 [8]. They constitute a heterogeneous group of rare inborn disorders marked by morphological abnormalities of the erythroblast, which lead to ineffective erythropoiesis by maturation block. They are distinct from other inherited bone marrow failure syndromes, as the non-erythroid hematopoietic lineages remain unaffected, and they neither seem to be associated with cytogenetic abnormalities nor a tendency to develop malignancy [6]. In their landmark paper, Heimpel and Wendt defined three major CDA subtypes (CDA-1, CDA-2, and CDA-3). Today, this classification remains valid, since the additional subgroups and variants that have been added in the meantime have rather limited value [6]. CDA-1 can be distinguished by peripheral macrocytosis, internuclear bridging, and ultrastructural defects in the chromatin, of erythroblasts. CDA-2 is marked by peripheral normocytosis, binuclearity of erythroblasts, and abnormal glycosylation of erythrocyte membrane proteins. In fact, CDA-2 was previously known as HEMPAS (Hereditary erythroblastic multinuclearity with positive acidified serum test), and the abnormal glycosylation of red cell membrane protein band-3 still serves as a valuable diagnostic tool [9]. Finally, CDA-3 is characterized by the multinuclearity of erythroblasts. In CDAs, the bone marrow findings can sometimes be extreme and lead to considerable anemia. However, this is the exception and not the rule. Many patients escape clinical attention as they maintain hemoglobin levels just above the threshold for symptoms or significant impairment in performance. For example, in CDA-2, jaundice from hemolysis can be the only presenting sign, and as little as 10% of patients require transfusions. In fact, generally, CDAs are an exclusion diagnosis. Hence, a long time-lapse from presentation of subtle signs of anemia/hemolysis to suspicion of CDA is common. In addition, many patients with CDA have spent years with an incorrect diagnosis (hemolytic anemia, myelodysplastic syndrome, iron-deficient anemia, thalassemia, erythrocyte membrane abnormality, or hemochromatosis) and have been exposed to potentially harmful iron supplementation, aggressive transfusion, or steroids and cocktails of vitamins. Nonspecific and sometimes confounding clinical features such as mental retardation or congenital malformations (mostly cardiovascular and skeletal), which could be pleiotropic signs of the disease, coincidental findings or by products of consanguinity, add to the diagnostic difficulty. Intuitively, basic research on the CDAs has the potential to identify critical pathways and important players in the process of erythropoiesis. Unfortunately, as with other orphan disorders, the knowledge on CDAs has long derived from an array of case reports or small series [10]. Until the late 1990s, the absence of collaborative registry data and systematic collection of biological samples at diagnosis has prevented the use of more advanced technology to identify the disorders' molecular pathogenesis. In fact, from an ethical standpoint, research purposes might not entirely justify a bone marrow aspirate in patients with CDA and slightly abnormal hemoglobin levels. This partly explains why the collection of research material has been rather limited. Nevertheless, the first breakthrough was made in 2002, with the description of the causative gene for CDA-1 (CDAN1 on chromosome 15) in an inbred population of Israeli Bedouins [11]. In parallel, genetic loci for CDA-2 and CDA-3 were reported in the literature [12, 13]. Almost 10 years later, time has come for the gene of CDA-2 to be unveiled. Through elegant investigations, including analysis of a zebrafish mutant model, Schwarz et al. showed that SEC23B, an essential component of the COP-II (coat protein complex) coat of vesicles, is mutated in this condition [14]. It is yet unclear how the abnormalities in the coat of COP-II vesicles, which carry protein cargo from the endoplasmic reticulum to the Golgi apparatus, cause the phenotype of CDA-2. Interestingly, SEC23B had been identified by other researchers to be a component of the cellular midbody, leading the CDA-2 investigators to speculate the incomplete cytokinesis (a key feature observed in CDA-2 erythroblasts) to be caused by a dysfunction of this organelle [15]. Although fascinating, this explanation hardly accounts for the abnormal glycosylation pattern of the erythrocyte membrane proteins classically seen in CDA-2, unless this was found to be a secondary phenomenon. Further functional work will be needed to understand how SEC23B is linked to the molecular pathogenesis of CDA-2. Therefore, clinical investigation of SEC23B mutants with CDA-2 can provide valuable information and assist in the diagnosis of the disorder. Russo and colleagues' work in this issue of the American Journal of Hematology is therefore noteworthy. In fact, these investigators add 19 novel SCE23B changes to the pre-existing map of 34 variants thought to cause CDA-2. Furthermore, their findings confirm previous work by the same group showing that compound heterozygosity for missense and nonsense mutations tends to lead to more severe clinical presentations (including lower reticulocyte counts and higher serum ferritin levels) than homozygosity or compound heterozygosity for bi-allelic SEC23B missense mutations [16]. This suggests that the association of one missense mutation and one nonsense mutation is significantly more deleterious than the association of two missense mutations. Also, as it is the case for CDA-1, homozygosity for two nonsense mutations was not encountered, and the authors, therefore, speculate on its embryonic lethality. In addition, Russo and colleagues performed in silico analysis on the SEC23B variants attempting to predict changes in the structure of the COP-II protein Sec23B or its precursor mRNA. Unfortunately, no functional assay is available to confirm the validity of the latter findings, which nonetheless open further interesting questions to investigate in the “wet” laboratory. In summary, the CDAs are a fascinating group of disorders with the potential to identify and investigate new players and pathways in erythropoiesis. Although recent progress has been steady and has led to the description of the genes causing CDA-1 and CDA-2, little is known on the functional impact of CDAN1/SEC23B mutations, and therefore the molecular mechanisms behind their pathogenesis. Interestingly, recent work on CDA-1 suggests that Codanin-1 causes CDA-1 by inappropriate cellular trafficking of the heterochromatin protein HP1-α (RR & Bill Wood, submitted). Combining experimental findings in CDA-1 and CDA-2, one could speculate that the CDAs are disorders of homeostasis of proteins specifically required in erythroid maturation. In fact, both disorders seem to indicate that effective protein trafficking, which requires functional vesicle transport, might be maturation limiting in erythroblasts. Enucleation, for example, could be one of the erythropoieis-specific events that could require specific protein transport. While the locus is known for CDA-3, it will be very interesting to find out whether the putative gene for this condition would also match this hypothetical model. Questions that will need to be addressed are based on our yet very limited understanding of the role of codanin-1 and Sec23B in normal and CDA-1/2 erythropoiesis. How do CDAN1/Sec23B mutations affect the structure of the codanin-1/Sec23B protein, and how does this impact erythropoiesis? The generation of animal models carrying these mutations and their study will certainly prove useful, although biochemical groundwork including crystallization of the proteins and subsequent identification of functional domains might be necessary at some stage. Solid functional assays resulting from these investigations will be the only way to reliably offer diagnosis and weed-out irrelevant sequence changes from our current mutation database. One essential question CDA investigators will have to answer is: why do widely expressed abnormal codanin-1/Sec23B proteins cause an erythroid-specific phenotype (as it also occurs with rRNA proteins in Diamond-Blackfan Anemia)? In addition, identifying the gene for CDA-3 should allow us to understand whether these disorders truly are molecularly linked. Globally, partnering CDAs with the vast skills and knowledge of vesicle biology and/or other fields via the discovery of putative genes/proteins will certainly boost research in this rather small field, and hopefully, positively impact clinical care of patients with this orphan disease. Thanks to Professor Bill Wood for his critical review and helpful suggestions.