Congenital Neutropenia Research Articles

Mitochondrial CLPB is a pro-survival factor at the onset of granulocytic differentiation of mouse myeloblastic cells.

Loss-of-function mutations in the CLPB gene lead to congenital neutropenia due to impaired neutrophil differentiation. CLPB, a member of the AAA+ family of proteins, resides in the intermembrane space of mitochondria. The mechanism by which a loss of CLPB elicits defects in the differentiation program of neutrophil precursor cells is not understood. Here, we used 32D clone 3 (32Dcl3) cells, an interleukin-3 (IL-3)-dependent mouse myeloblastic cell line model, to investigate the effects of CLPB knockout on myeloblast-to-neutrophil differentiation in vitro. We found that CLPB-deficient 32Dcl3 cells showed a decreased mitochondrial membrane potential and increased levels of insoluble HAX1 aggregates in mitochondria, as compared to control cells. Despite those abnormalities, CLPB loss did not affect cell proliferation rates in the presence of IL-3 but it increased apoptosis after IL-3 withdrawal and simultaneous induction of cell differentiation with granulocytic colony stimulating factor (G-CSF). CLPB-deficient cells that survived the stress associated with IL-3 withdrawal/G-CSF treatment expressed the same levels of differentiation markers as control cells. Moreover, we found that increased apoptosis of CLPB-deficient cells is linked to production of reactive oxygen species (ROS). N-acetylcysteine, exogenous free fatty acids, or exogenous citrate protected CLPB-deficient 32Dcl3 cells from apoptosis at the onset of differentiation. The protective effect of citrate was abolished by inhibition of ATP-citrate lyase (ACLY), an enzyme that converts cytosolic citrate into acetyl-CoA, a substrate for protein acetylation. We propose that citrate supplementation may help mitigate the effects of CLPB loss by facilitating ACLY-dependent ROS detoxification in granulocytic precursor cells.

A New Severe Congenital Neutropenia Syndrome Associated with Autosomal Recessive COPZ1 Mutations.

We have identified a new inherited bone marrow (BM) failure syndrome with severe congenital neutropenia (CN) caused by autosomal recessive mutations in the coatomer protein complex I (COPI) subunit zeta 1 (COPZ1) gene. A stop-codon COPZ1 mutation and a missense mutation were found in three patients from two unrelated families. While two affected siblings with a stop-codon COPZ1 mutation suffered from congenital neutropenia (CN) that involves other hematological lineages, and non-hematological tissues, the patient with a missense COPZ1 mutation had isolated neutropenia. Both COPZ1 mutations were localized to a highly evolutionarily conserved region. The resulting truncated COPZ1 protein was predicted to display diminished interaction with its COPI complex partner, COPG1. These findings were consistent with the observed block in retrograde protein transport from the Golgi to the ER in human fibroblasts carrying truncated COPZ1. Human CD34+ cells with truncated or missense COPZ1 had significantly impaired granulocytic differentiation and in zebrafish embryos, truncated Copz1 also resulted in defective myelopoiesis. Intracellularly, truncated COPZ1 downregulated JAK/STAT/CEBPE/G-CSFR signaling and hypoxia-responsive pathways while inducing STING, interferon-stimulated genes, stimulating oxidative phosphorylation activity, and increasing reactive oxygen species (ROS) levels in CD34+ cells. Missense COPZ1 deregulated interferon and JAK/STAT signaling but less than the truncated protein. Finally, treatment with the small molecule HIF1α activator IOX2 or transduction of cells with COPZ2 cDNA restored defective granulopoiesis in COPZ1-mutated CD34+ cells, offering potential therapeutic options.

Molecular and Clinical Characterization of a Founder Mutation Causing G6PC3 Deficiency

G6PC3 deficiency is a monogenic immunometabolic disorder that causes severe congenital neutropenia type 4. Patients display heterogeneous extra-hematological manifestations, contributing to delayed diagnosis. Here, we investigated the origin and functional consequence of the G6PC3 c.210delC variant found in patients of Mexican descent. Based on the shared haplotypes amongst mutation carriers, we estimated that this variant originated from a founder effect in a common ancestor. Furthermore, by ancestry analysis, we concluded that it appeared in the indigenous Mexican population. At the protein level, we showed that this frameshift mutation leads to an aberrant protein expression in overexpression and patient-derived Epstein-Barr Virus-immortalized B (EBV-B) cells. The neutropenia observed in G6PC3-deficient patients is driven by the intracellular accumulation of the metabolite 1,5-anhydroglucitol-6-phosphate (1,5-AG6P) that inhibits glycolysis. We characterized how the c.210delC variant impacts glycolysis by performing extracellular flux assays on patient-derived EBV-B cells. When treated with 1,5-anhydroglucitol (1,5-AG), the precursor to 1,5-AG6P, patient cells exhibited markedly reduced engagement of glycolysis. Finally, we compared the clinical presentation of patients with the mutation c.210delC and all other G6PC3-deficient patients reported in the literature, and we found that the c.210delC carriers display all prominent clinical features observed in prior patients. In conclusion, G6PC3 c.210delC is a loss-of-function mutation that arose from a founder effect in the indigenous Mexican population. These findings may facilitate the diagnosis of additional patients in this geographical area. Moreover, the in vitro 1,5-AG-dependent functional assay used in our study could be employed to assess the pathogenicity of additional G6PC3 variants.

Flavopiridol restores granulopoiesis in experimental models of severe congenital neutropenia.

Severe congenital neutropenia (CN) patients require life-long treatment with recombinant human granulocyte colony-stimulating factor (rhG-CSF), but some show no response. We sought to establish a therapy for CN that targets signaling pathways causing maturation arrest of granulocytic progenitors. We developed an isogenic induced pluripotent stem cell (iPSC) invitro model of CN associated with ELANE mutations (ELANE-CN) and performed an in silico drug repurposing analysis of the transcriptomics of iPSC-generated hematopoietic stem and progenitor cells. We identified flavopiridol, a Food and Drug Administration (FDA)-approved pan-cyclin-dependent kinase inhibitor, as a potential therapeutic. Treatment with low-dose flavopiridol rescued defective granulopoiesis in primary CD34+ cells of CN patients with different inherited gene mutations invitro and in two zebrafish CN models invivo without any toxic effects and leading to functional granulocytes. Flavopiridol also restored granulopoiesis caused by diminished CEBPA expression, a known defective signaling molecule in CN. Thus, we described for the first time a potential therapy for CN with flavopiridol that could be potentially used to treat patients with different types of neutropenia.

G-CSF resistance of ELANE mutant neutropenia depends on SERF1 containing truncated neutrophil elastase aggregates.

Severe congenital neutropenia (SCN) is frequently associated with dominant point mutations in ELANE, the gene encoding neutrophil elastase (NE). Chronic administration of granulocyte colony-stimulating factor (G-CSF) is a first-line treatment of ELANE-mutant (ELANEmut) SCN. However, some ELANEmut patients including patients with ELANE start codon mutations do not respond to G-CSF. Here, through directed granulopoiesis of gene-edited isogenic normal and patient-derived iPSCs, we demonstrate that ELANE start codon mutations suffice to induce G-CSF resistant granulocytic precursor cell death and refractory SCN. ELANE start codon mutated neutrophil precursors express predominantly nuclear N-terminal truncated alternate NE. Unlike G-CSF sensitive ELANE mutations that induce endoplasmic reticulum and unfolded protein response stress, we found that the mutation of the ELANE translation initiation codon resulted in NE aggregates and activated pro-apoptotic aggrephagy as determined by downregulated BAG1 expression, decreased BAG1/BAG3 ratio, NE co-localization with BAG3, and localized expression of autophagic LC3B. We found that SERF1, an RNA-chaperone protein, known to localize in misfolded protein aggregates in neurodegenerative diseases, was highly upregulated and interacted with cytoplasmic NE of mutant neutrophil precursors. Silencing of SERF1 enhanced survival and differentiation of iPSC-derived neutrophil precursors, restoring their responsiveness to G-CSF. These observations provide a mechanistic insight of G-CSF-resistant ELANEmut SCN, revealing targets for therapeutic intervention.

A large cohort from an immunology reference center and an algorithm for the follow-up of chronic neutropenia.

Chronic neutropenia causes involve nutritional deficiencies and inborn errors of immunity(IEI), such as severe congenital neutropenia. To classify common chronic neutropenia causes in a pediatric immunology unit. We enrolled 109 chronic neutropenia patients admitted to a pediatric immunology department between 2002-2022. We recorded clinical/laboratory features and genetic characteristics. The male/female ratio was 63/46. Fifty-eight patients had parental consanguinity(57.4%). 26.6% (n = 29) patients had at least one individual in their family with neutropenia. Common symtpoms at presentation were upper respiratory tract infections(URTI)(31.1%), oral aphthae(23.6%), skin infections(23.6%), pneumonia(20.8%), and recurrent abscesses(12.3%). Common infections during follow-up were URTI(56.8%), pneumonia(33%), skin infections(25.6%), gastroenteritis(18.3%), and recurrent abscesses(14,6%). Common long-term complications were dental problems(n = 51), osteoporosis(n = 22), growth retardation(n = 14), malignancy(n = 16)[myelodysplastic syndrome(n = 10), large granulocytic leukemia(n = 1), acute lymphoblastic leukemia(n = 1), Hodgkin lymphoma(n = 1), EBV-related lymphoma(n = 1), leiomyosarcoma(n = 1), and thyroid neoplasm(n = 1)]. We performed a genetic study in 86 patients, and 69(71%) got a genetic diagnosis. Common gene defects were HAX-1(n = 26), ELA-2 (ELANE)(n = 10), AP3B1(n = 4), and ADA-2(n = 4) gene defects. The IEI ratio(70.6%) was high. GCSF treatment(93.4%), immunoglobulin replacement therapy(18.7%), and HSCT(15.9%) were the treatment options. The mortality rate was 12.9%(n = 14). The most common long term complications were dental problems that is three times more common in patients with known genetic mutations. We prepared an algorithm for chronic neutropenia depending on the present cohort. An important rate of inborn errors of immunity, especially combined immunodeficiency(11.9%) was presented in addition to congenital phagocytic cell defects. Early diagnosis will allow us tailor the disease-specific treatment options sooner, preventing irreversible consequences.

Protein Network Alterations in G-CSF Treated Severe Congenital Neutropenia Patients and Beneficial Effects of Oral Health Intervention.

Severe congenital neutropenia (SCN) is a raredisorder characterized by diminished neutrophil levels. Despite granulocytecolony-stimulating factor (G-CSF) treatment, SCN patients remain still prone tosevere infections, including periodontal disease-a significant oral healthrisk. This study investigates the host proteome and metaproteome in saliva andgingival crevicular fluid (GCF) of G-CSF-treated patients. We used label-free quantitative proteomics on saliva and GCF samples from SCN patients before (n=10, mean age: 10.7±6.6 years) and after a 6-month oral hygiene intervention (n=9,mean age: 11.6±5.27 years), and from 12 healthy controls. We quantified 894 proteins in saliva (648 human,246 bacterial) and 756 proteins in GCF (493 human, 263 bacterial). Predominant bacterial genera included Streptococcus, Veillonella, Selenomonas, Corynebacterium, Porphyromonas, and Prevotella. SCN patients showed reduced antimicrobial peptides (AMPs) and elevated complement proteins compared tohealthy controls. Oral hygiene intervention improved oral epithelial conditionsand reduced both AMPs and complement proteins. SCN patients have aunique proteomic profile with reduced AMPs and increased complement proteins, contributing to infection susceptibility. Oral hygiene intervention not onlyimproved oral health in SCN patients but also offers potential overall therapeuticbenefits.

Successful repurposing of empagliflozin to treat neutropenia in a severe congenital neutropenia patient with G6PC3 mutations

Successful repurposing of empagliflozin to treat neutropenia in a severe congenital neutropenia patient with G6PC3 mutations

Severe Congenital Neutropenia with ELANE gene mutation, on G-CSF therapy; a case report

Abstract Introduction/Objective Severe congenital neutropenia (SCN) is a genetic syndrome characterized by a deficiency of mature neutrophils in the bone marrow and peripheral blood. The disease is caused by various gene mutations, with ELANE gene mutations accounting for 50% to 60% of cases. Methods/Case Report We report a case of a 12-year-old male patient with severe congenital neutropenia, positive for the ELANE gene mutation. He has a history of recurrent infections and unexplained fevers since childhood. He has been receiving G-CSF treatment for ten years but is now exhibiting a diminished response to it. Analysis of the bone marrow biopsy reveals trilineage hematopoiesis, maturing myeloid with a rise in eosinophils, maturing erythroid, megakaryocytes normal in number and morphology with significantly reduced iron reserves in normocellular bone marrow. On immunostaining, CD34, CD14, or CD117 stains were negative. Neither monocytes nor granulocytes were CD56 positive. Increased monocyte counts coincide with partial CD14 loss. Lymphocytes make up 10% of all cells and consist of Polytypic B-cells, natural killer cells, and a heterogeneous population of T-cells with a normal CD4 to CD8 ratio. Hence, myelodysplastic syndrome was ruled out. Results (if a Case Study enter NA) NA Conclusion This case report provides valuable insights into the limited knowledge of congenital neutropenia with ELANE mutation on G-CSF therapy. It highlights the importance of ruling out Myelodysplastic syndrome in patients with severe congenital neutropenia and ELANE gene mutation treated with G-CSF therapy.

Inherited Predispositions to Myeloid Neoplasms: Pathogenesis and Clinical Implications.

Myeloid neoplasms with and without preexisting platelet disorders frequently develop in association with an underlying germline predisposition. Germline alterations affecting ANKRD26, CEBPA, DDX41, ETV6, and RUNX1 are associated with nonsyndromic predisposition to the development of myeloid neoplasms including acute myeloid leukemia and myelodysplastic syndrome. However, germline predisposition to myeloid neoplasms is also associated with a wide range of other syndromes, including SAMD9/9L associated predisposition, GATA2 deficiency, RASopathies, ribosomopathies, telomere biology disorders, Fanconi anemia, severe congenital neutropenia, Down syndrome, and others. In the fifth edition of the World Health Organization (WHO) series on the classification of tumors of hematopoietic and lymphoid tissues, myeloid neoplasms associated with germline predisposition have been recognized as a separate entity. Here, we review several disorders from this WHO entity as well as other related conditions with an emphasis on the molecular pathogenesis of disease and accompanying somatic alterations. Finally, we provide an overview of establishing the molecular diagnosis of these germline genetic conditions and general recommendations for screening and management of the associated hematologic conditions.

Comparison of Gene-Editing Approaches for Severe Congenital Neutropenia-Causing Mutations in the ELANE Gene.

Safety considerations for gene therapies of inherited preleukemia syndromes, including severe congenital neutropenia (CN), are paramount. We compared several strategies for CRISPR/Cas9 gene editing of autosomal-dominant ELANE mutations in CD34+ cells from two CN patients head-to-head. We tested universal and allele-specific ELANE knockout, ELANE mutation correction by homology-directed repair (HDR) with AAV6, and allele-specific HDR with ssODN. All strategies were not toxic, had at least 30% editing, and rescued granulopoiesis in vitro. In contrast to published data, allele-specific indels in the last exon of ELANE also restored granulopoiesis. Moreover, by implementing patient-derived induced pluripotent stem cells for GUIDE-Seq off-target analysis, we established a clinically relevant "personalized" assessment of off-target activity of gene editing on the background of the patient's genome. We found that allele-specific approaches had the most favorable off-target profiles. Taken together, a well-defined head-to-head comparison pipeline for selecting the appropriate gene therapy is essential for diseases, with several gene editing strategies available.

De Novo Deep Intron ELANE Mutation Resulting in Severe Congenital Neutropenia.

Severe congenital neutropenia (SCN) comprises a diverse range of rare hematological disorders characterized by recurrent, often life-threatening infections that manifest within the first months of life. Mutations in the ELANE gene are the most prevalent cause of SCN. While over 230 mutations in ELANE have been documented, including substitutions, frameshifts, nonsense mutations, and splice site alterations, the occurrence of deep intronic mutations has not been previously reported. Herein, we present the case of a young girl who exhibited recurrent fever, respiratory infections, skin abscesses, and gingivitis shortly after birth. Laboratory analysis revealed markedly diminished neutrophil levels alongside elevated monocyte and eosinophil counts. Bone marrow examination disclosed a halt in myelopoiesis maturation. ELANE gene full-length sequencing identified a novel de novo deep intron mutation in ELANE (c.598 + 79G > T), subsequently confirmed by Sanger sequencing. cDNA sequencing of the patient demonstrated aberrant gene splicing. Utilizing a mini-gene splicing assay for ELANE intronic variants, we identified a mutant ELANE allele (c.597 + 1_597 + 83ins) leading to the creation of a premature termination codon (p.Gly200ValfsTer40). Confocal microscopy revealed heightened expression of myeloperoxidase and neutrophil elastase in the patient, suggesting a potential role for the unfolded protein response in the pathogenesis of the deep intron ELANE mutation. In summary, our findings illustrate the first reported instance of de novo deep intron ELANE mutations associated with SCN, underscoring the importance of exploring deep intronic regions in SCN patients lacking identifiable disease-causing gene mutations.

Abstract 15: Invasive Aspergillus Niger Infection Cured after V Hematopoietic Stem Cell Transplantation in a Pediatric Patient with Severe Congenital Neutropenia

Abstract 15: Invasive Aspergillus Niger Infection Cured after V Hematopoietic Stem Cell Transplantation in a Pediatric Patient with Severe Congenital Neutropenia

Forty years of human G-CSF: A short history in time.

Human G-CSF was identified in 1984 at Memorial Sloan-Kettering Cancer Centre, New York. Based on these findings, recombinant G-CSF was developed by Amgen, Thousand Oaks. In 1987, clinical trials began using recombinant G-CSF in cancer patients following chemotherapy to reduce the duration of neutropenia and in patients with congenital neutropenia (CN) to increase the number of neutrophils. It has changed the quality of life for many cancer patients and saved the lives of many patients with (CN).

Expanding the genetic landscape of congenital neutropenia: CXCR2 mutations in three families revealed through whole exome sequencing.

Not available.

Oral health management in children with severe congenital neutropenia with periodontitis: Case report.

Severe congenital neutropenia (SCN) is a rare and heterogeneous genetic disease. By describing the diagnosis and treatment of a child with SCN and periodontitis, this case provides a reference for the oral health management of a child with SCN and periodontitis. We describe a boy with clinical manifestations of oral bleeding, neutropenia, recurrent fever, and other recurrent infections. The absolute neutrophil count (ANC) was <0.50 × 109/L most of the time. Morphological examination of bone marrow cells showed active granulocyte hyperplasia and dysmaturation. According to the clinical manifestations, hematological examination and gene detection results, the child was diagnosed as SCN with chronic periodontitis. Periodontal treatment was performed after informed consent was obtained from the child guardian. These included supragingival and subgingival cleaning, hydrogen peroxide and saline irrigation, placement of iodoglycerin in the gingival sulcus, and oral hygiene instruction. Hematopoietic stem cell transplantation (HSCT) was performed later. One month after initial periodontal treatment, oral hygiene was well maintained and gingival swelling had subsided. Probing depth (PD) index on periodontal probing and bleeding was significantly reduced. However, there was no significant change in blood routine and other indicators before and after periodontal treatment. Once SCN is diagnosed, individualized treatment plans can be developed according to the characteristics of the disease and its impact on oral health, which can effectively control the interaction between SCN and periodontal disease and reduce the occurrence of serious infection.

Molecular and clinical characterization of a founder mutation causing G6PC3 deficiency.

G6PC3 deficiency is a monogenic immunometabolic disorder that causes syndromic congenital neutropenia. Patients display heterogeneous extra-hematological manifestations, contributing to delayed diagnosis. Here, we investigated the origin and functional consequence of the G6PC3 c.210delC variant found in patients of Mexican origin. Based on the shared haplotypes amongst carriers of the c.210delC mutation, we estimated that this variant originated from a founder effect in a common ancestor. Furthermore, by ancestry analysis, we concluded that it originated in the indigenous Mexican population. At the protein level, we showed that this frameshift mutation leads to an aberrant protein expression in overexpression and patient-derived cells. G6PC3 pathology is driven by the intracellular accumulation of the metabolite 1,5-anhydroglucitol-6-phosphate (1,5-AG6P) that inhibits glycolysis. We characterized how the variant c.210delC impacts glycolysis by performing extracellular flux assays on patient-derived cells. When treated with 1,5-anhydroglucitol (1,5-AG), the precursor to 1,5-AG6P, patient-derived cells exhibited markedly reduced engagement of glycolysis. Finally, we compared the clinical presentation of patients with the mutation c.210delC and all other G6PC3 deficient patients reported in the literature to date, and we found that c.210delC carriers display all prominent clinical features observed in prior G6PC3 deficient patients. In conclusion, G6PC3 c.210delC is a loss-of-function mutation that arose from a founder effect in the indigenous Mexican population. These findings may facilitate the diagnosis of additional patients in this geographical area. Moreover, the in vitro 1,5-AG-dependent functional assay used in our study could be employed to assess the pathogenicity of additional G6PC3 variants.

Clinical Profile of Adults with Inherited Bone Marrow Failure Syndromes: Results of an Ambispective Clinical Single-Center Study

BACKGROUND. Inherited bone marrow failure syndromes (IBMFS) is a heterogenous group of rare genetically determined diseases with variable hematologic and nonhematologic manifestations. The implementation of highly specific methods of genetic diagnosis advanced the understanding of IBMFS and allowed its application also beyond pediatrics. That presupposes an awareness of clinical features and reference points for recognizing IBMFS in adults. AIM. To describe the clinical profile of adult IBMFS patients. MATERIALS &amp; METHODS. This ambispective single-center study enrolled 35 patients (10 women and 25 men) with IBMFS. Patients were aged 18–51 years (median 26 years). The following IBMFS were identified: congenital dyskeratosis (n = 10; 28 %), Diamond-Blackfan anemia (n = 9; 26 %), Fanconi anemia (n = 7; 20 %), GATA2 deficiency (n = 3; 8 %), Shwachman-Diamond syndrome (n = 1; 3 %), GATA2 deficiency (n = 1; 3 %), amegakaryocytic thrombocytopenia (n = 1; 3 %), bone marrow failure syndrome type 3 (n = 1; 3 %), severe congenital neutropenia (n = 1; 3 %), bone marrow failure with SAMD9 mutation (n = 1; 3 %). These diseases were analyzed in terms of hematologic and nonhematologic manifestations as well as main diagnosis stages and factors that contribute to recognizing IBMFS. RESULTS. Monolinear cytopenia, bilinear cytopenia, and pancytopenia were identified at hematologic onset in 18 (52 %), 6 (17 %), and 11 (31 %) patients, respectively. The median age of patients by hematologic onset was 15 years (range 0–43 years), in 14 (40 %) patients cytopenia was newly diagnosed at the age of &gt; 18 years. In 23 (63 %) patients hypocellular bone marrow was reported, 7 (20 %) and 5 (14 %) patients had pure red cell aplasia and multilineage myelodysplasia, respectively. Chromosomal aberrations were identified in 2 patients. Paroxysmal nocturnal hemoglobinuria clone was detected in none of 27 examined patients. In 12 (34 %) patients, the criteria for non-severe aplastic anemia were met. Temporary partial or complete spontaneous hematologic recovery was observed in 6 (17 %) patients. Abnormalities with partial or complete organ dysfunctions were identified in 14 patients, whereas all patients showed minor congenital defects. All 7 Fanconi anemia patients and 9 out of 10 congenital dyskeratosis patients demonstrated organ damage specific to these diseases. Family history predominantly showing malignant neoplasms in relatives was reported in 15 (43 %) patients. Initial hematological examination yielded suspect of IBMFS in 12 (34 %) patients with the median time to diagnosis of 6 months. In 23 (66 %) patients, hematologic defects with cytopenia were erroneously accounted for by various acquired diseases, which led to a delayed correct diagnosis (median 7 years). The key factors in suspecting IBMFS were organ abnormalities and positive family history. The IBMFS diagnosis was verified by the next-generation sequencing (NGS) in 29 (83 %) patients and by other specific methods in 4 (11 %) patients. In 2 patients, the diagnosis was established on the basis of complete clinical criteria alone. CONCLUSION. IBMFS is a matter of current concern and a difficult-to-recognize clinical challenge in adult hematology patients. Differential diagnosis of acquired and congenital bone marrow failure needs to be performed irrespective of patient’s age. A detailed physical examination of patients, family history, and critical analysis of clinical profile and disease course allow for early suspicion of IBMFS. Suspected IBMFS is an indication for referral of patients to specialized centers and performing genetic diagnostics including NGS.

Absence of Neutropenia in Patients With Early Exon Nonsense Mutations in ELANE : Clinical Evidence to Support Gene Therapy Approaches for Severe Congenital Neutropenia.

Severe congenital neutropenia is an inherited bone marrow failure disorder characterized by profoundly low neutrophil counts and promyelocytic maturation arrest in bone marrow. Severe congenital neutropenia is most often caused by heterozygous ELANE mutations. In vitro and mouse xenograft studies using CRISPR/Cas9 have shown that introduction of frameshift/nonsense mutations in mutant ELANE may restore neutrophil counts, providing a model for gene therapy. Here, we present 2 children with inherited nonsense mutations in ELANE analogous to those proposed for gene therapy. Their normal peripheral blood neutrophil counts provide support for this approach through human "experiments of nature."

ДИФФЕРЕНЦИАЛЬНАЯ ДИАГНОСТИКА ПРИОБРЕТЕННОЙ АПЛАСТИЧЕСКОЙ АНЕМИИ У ДЕТЕЙ: АНАЛИЗ ДАННЫХ РЕГИСТРОВОГО ИССЛЕДОВАНИЯ

Acquired aplastic anemia (AA) is still the diagnosis of exclusion. The purpose of the research was to identify clinical and laboratory signs that do not correspond to acquired AA allowing to diagnose other diseases accompanied by cytopenia. Materials and methods used: Authors represent the results of diagnostic search in pediatric patients (0 to 18 y/o) with a suspected diagnosis of acquired AA who underwent remote diagnosis in the laboratories of the National Scientific and Practical Center for Pediatric Hematology, Oncology and Immunology named after Dmitry Rogachev (Moscow, Russia) as part of the Register Study in Sep. 1, 2017-May 1, 2022. Results: in retrospective analysis of 523 children records, 104 (20%) patients had other diagnoses. In 29 (27.9%) patients were diagnosed with inherited bone marrow failure syndromes (IBMFS), in 11 (10.6%) with myelodysplastic syndrome, in 11 (10.6%) with deficiency anemia, in 10 (9.6%) with congenital unspecified AA, in 7 (6.7%) with immune cytopenia, in 7 (6.7%) with inborn defects of immunity, in 6 (5.8%) with secondary AA, in 6 (5.8%) with acute leukemia, in 2 (1.9%) with paroxysmal nocturnal hemoglobinuria and in 15 (14.4%) cases with other unspecified diseases. Fanconi anemia (n=15/29 patients) was the most common disease in the IBMFS group; this group had also included: dyskeratosis congenita (n=9), Schwachman-Diamond syndrome (n=2), amegakaryocytic thrombocytopenia (n=2), Diamond-Blackfan anemia in combination with congenital neutropenia (n=1). Conclusion: differential diagnosis of acquired AA can be difficult though it is undoubtedly must-do for successful treatment. Verification of other alternative diseases, including genetically determined syndromes, contributes to the choice of optimal patient management tactics.