Page 43 - Haematologica August 2018

GATA2 deficiency French/Belgian cohort
patients with nonsense or frameshift mutations (n=2 of 28). We also identify new features of the disease: acute lymphoblastic leukemia, juvenile myelomonocytic leukemia, fatal progressive multifocal leukoen- cephalopathy related to the JC virus, and immune/inflammatory diseases. A revised International Prognostic Scoring System (IPSS) score allowed a distinction to be made between a stable disease and hematologic transformation. Chemotherapy is of limited efficacy, and has a high toxicity with severe infectious complications. As the mortality rate is high in our cohort (up to 35% at the age of 40), hematopoietic stem cell transplantation (HSCT) remains the best choice of treatment to avoid severe infectious and/or hematologic complications. The timing of HSCT remains difficult to determine, but the earlier it is performed, the better the outcome.
Introduction
GATA2 gene encodes a transcription factor critical to hematopoiesis characterized by the presence of two zinc finger domains. Since 2011, heterozygous germline mutations in GATA2 have been reported to cause a com- plex and heterogeneous syndrome consisting of myelodysplasia (MDS), acute myeloid leukemia (AML),1 monocytopenia mycobacterial infections/dendritic cell,2 monocyte, B and natural killer (NK) cell deficiency (MonoMAC3,4/DMLC),5 and lymphoedema (Emberger syndrome).6 The mutational spectrum of GATA2 is hetero- geneous, consisting of missense mutations mostly located within the highly conserved C-terminal zinc finger domains, null mutations mostly located upstream the zinc finger domains, splice site defects, mutations in the enhancer located in the intron 4,7 and, more rarely, exonic and whole-gene deletions.
Apart from hematologic and infectious phenotypes, additional clinical presentations have been described in the last six years, such as aplastic anemia,8 pulmonary alveolar proteinosis,9 dermatological,10 autoimmune or vascular features. So far, a total of 158 patients with a germline GATA2 mutation have been reported in 4 large surveys.11-14 Except for lymphoedema which is more fre- quent in patients with null or regulatory mutations, no correlation between the type or location of the GATA2 mutation and the clinical/biological phenotype have been established in previous reports.
We now report a large multicenter survey which brings together all the patients that have been diagnosed in France and Belgium since 2011, i.e. 79 patients with a het- erozygous germline GATA2 mutation from 53 pedigrees. These 79 patients include 14 previously identified patients through the French Chronic Neutropenia Registry;11 their follow up has been up-dated. This survey allows human GATA2 deficiency to be more accurately defined, taking advantage of a very long follow-up period (2249 patients/years). We describe the initial manifestations, their evolution, and their outcome regarding the onset of severe manifestations (leukemia, severe infections, vascu- lar defects). This large cohort also allows new features of the disease to be described.
Methods
Patients
All patients with heterozygous germline GATA2 mutations diagnosed between 2011 and 2016 in France and Belgium were enrolled in this survey, secondary to their identification through
the laboratories which performed their genetic diagnosis in France and Belgium (JB, CBC, HC-AC, ED, CP). Fourteen patients with chronic neutropenia, and who were registered in the French Severe Chronic Neutropenia Registry, had been reported previ- ously.11 This registry has been recognized as a national registry by the French health authorities since 2008, and has contributed to several studies.15,16 The database was approved by the French national data protection agency (CNIL, certificate n. 97.075). This registry was primary established to enrol all the patients with chronic neutropenia in France. By extension, all patients identified with a given genetic disease (e.g. GATA2 ) occasionally associated with a chronic neutropenia can be enrolled in the registry. With regards to GATA2 mutations, we systemically seek additional sources of enrollment, extending the borders of the initial network to internal medicine, infectious diseases and genetics, as well as from adult hematopoietic stem cell transplantation (HSCT) units.
Genetic analysis
The patient or his/her parents gave their written informed consent to undergo genetic testing and participate in the study. Genomic DNA was extracted from a blood sample. Genetic analysis included the Sanger sequencing of exons 2 to 6, the intronic regulatory region (intron 4) of the GATA2 gene (NM_032631.4), and the search for exonic or large genomic deletions by quantitative PCR and/or MLPA. The germline sta- tus of the identified GATA2 mutation was confirmed by analyz- ing non-hematopoietic tissue (cultured skin fibroblasts, hair fol- licles or nails) in 30 probands. Null mutations (nonsense, frameshift, multi-exon deletion) were considered to be disease- causing. The pathogenicity of missense mutations and splice- site variants that did not affect the canonical +1 and +2 splice site bases were based on the following criteria: frequency in the general-population database [Exome Aggregation Consortium (ExAC): http://exac.broadinstitute.org], literature that took into account mutations that were previously reported as a GATA2- associated defect, functional studies supporting a damaging effect, a de novo occurrence, family segregation analysis, and finally pre- dictive algorithms of pathogenicity for missense mutations [SIFT, Align GVGD, PolyPhen-2 and Combined Annotation-Dependent Depletion (CADD) score, and for splice-site defects (MaxEntScan and Human Splicing Finder)].17
Clinical investigation
Demographics, immuno-hematologic parameters and infec- tious status were recorded. Septicemia, cellulitis, pneumonia, osteitis, and liver abscess were considered to be severe infections. Computed tomography (CT) scans, bronchioalveolar lavage and pulmonary function investigations were performed in patients with lung disease. Profuse skin or genito-anal warts were consid- ered to be a specific event. Mycobacterial infections were consid- ered if mycobacteria were identified in a pathological tissue (Ziehl
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