Clinical evolution, genetic landscape and trajectories of clonal hematopoiesis in SAMD9/SAMD9L syndromes.

Germline SAMD9 and SAMD9L mutations (SAMD9/9Lmut) predispose to myelodysplastic syndromes (MDS) with propensity for somatic rescue. In this study, we investigated a clinically annotated pediatric MDS cohort (n = 669) to define the prevalence, genetic landscape, phenotype, therapy outcome and clonal architecture of SAMD9/9L syndromes. In consecutively diagnosed MDS, germline SAMD9/9Lmut accounted for 8% and were mutually exclusive with GATA2 mutations present in 7% of the cohort. Among SAMD9/9Lmut cases, refractory cytopenia was the most prevalent MDS subtype (90%); acquired monosomy 7 was present in 38%; constitutional abnormalities were noted in 57%; and immune dysfunction was present in 28%. The clinical outcome was independent of germline mutations. In total, 67 patients had 58 distinct germline SAMD9/9Lmut clustering to protein middle regions. Despite inconclusive in silico prediction, 94% of SAMD9/9Lmut suppressed HEK293 cell growth, and mutations expressed in CD34+ cells induced overt cell death. Furthermore, we found that 61% of SAMD9/9Lmut patients underwent somatic genetic rescue (SGR) resulting in clonal hematopoiesis, of which 95% was maladaptive (monosomy 7 ± cancer mutations), and 51% had adaptive nature (revertant UPD7q, somatic SAMD9/9Lmut). Finally, bone marrow single-cell DNA sequencing revealed multiple competing SGR events in individual patients. Our findings demonstrate that SGR is common in SAMD9/9Lmut MDS and exemplify the exceptional plasticity of hematopoiesis in children.

Hematopoietic stem cell transplantation in children and adolescents with GATA2-related myelodysplastic syndrome.

GATA2 deficiency is a heterogeneous multi-system disorder characterized by a high risk of developing myelodysplastic syndrome (MDS) and myeloid leukemia. We analyzed the outcome of 65 patients reported to the registry of the European Working Group (EWOG) of MDS in childhood carrying a germline GATA2 mutation (GATA2mut) who had undergone hematopoietic stem cell transplantation (HSCT). At 5 years the probability of overall survival and disease-free survival (DFS) was 75% and 70%, respectively. Non-relapse mortality and relapse equally contributed to treatment failure. There was no evidence of increased incidence of graft-versus-host-disease or excessive rates of infections or organ toxicities. Advanced disease and monosomy 7 (-7) were associated with worse outcome. Patients with refractory cytopenia of childhood (RCC) and normal karyotype showed an excellent outcome (DFS 90%) compared to RCC and -7 (DFS 67%). Comparing outcome of GATA2mut with GATA2wt patients, there was no difference in DFS in patients with RCC and normal karyotype. The same was true for patients with -7 across morphological subtypes. We demonstrate that HSCT outcome is independent of GATA2 germline mutations in pediatric MDS suggesting the application of standard MDS algorithms and protocols. Our data support considering HSCT early in the course of GATA2 deficiency in young individuals.

Favorable outcomes of hematopoietic stem cell transplantation in children and adolescents with Diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) is a congenital pure red cell aplasia associated with congenital abnormalities and cancer predisposition. Allogeneic hematopoietic stem cell transplantation (HSCT) can correct the hematological phenotype and is indicated in transfusion-dependent patients. In 70 children reported to the German DBA and French HSCT registries, HSCT was performed from 1985 to 2017. Median age at HSCT was 5.5 years (range, 0.9-17.3 years). Two-thirds of patients (64%) were transplanted from a matched sibling donor (MSD), and most procedures were performed after the year 1999 (73%). Primary engraftment was achieved in all patients. One patient developed secondary graft failure. Cumulative incidence of acute graft-versus-host disease (GVHD) was 24% for °II-IV (95% confidence interval [CI], 16% to 37%) and 7% for °III-IV (95% CI, 3% to 17%); cumulative incidence of chronic GVHD was 11% (95% CI, 5% to 22%). The probability of chronic GVHD-free survival (cGFS) was 87% (95% CI, 79% to 95%) and significantly improved over time (<2000: 68% [95% CI, 47% to 89%] vs ≥2000: 94% [95% CI, 87% to 100%], P < .01). cGFS was comparable following HSCT from a MSD and an unrelated donor (UD). Of note, no severe chronic GVHD or deaths were reported following MSD-HSCT after 1999. The difference of cGFS in children transplanted <10 years of age compared with older patients did not reach statistical significance (<10 years: 90% [95% CI, 81% to 99%] vs 10-18 years 78% [95% CI, 58% to 98%]). In summary, these data indicate that HSCT is efficient and safe in young DBA patients and should be considered if a MSD or matched UD is available. HSCT for transfusion dependency only must be critically discussed in older patients.

LIN28B overexpression defines a novel fetal-like subgroup of juvenile myelomonocytic leukemia.

Juvenile myelomonocytic leukemia (JMML) is a rare and aggressive stem cell disease of early childhood. RAS activation constitutes the core component of oncogenic signaling. In addition, leukemic blasts in one-fourth of JMML patients present with monosomy 7, and more than half of patients show elevated age-adjusted fetal hemoglobin (HbF) levels. Hematopoietic stem cell transplantation is the current standard of care and results in an event-free survival rate of 50% to 60%, indicating that novel molecular-driven therapeutic options are urgently needed. Using gene expression profiling in a series of 82 patient samples, we aimed at understanding the molecular biology behind JMML and identified a previously unrecognized molecular subgroup characterized by high LIN28B expression. LIN28B overexpression was significantly correlated with higher HbF levels, whereas patients with monosomy 7 seldom showed enhanced LIN28B expression. This finding gives a biological explanation of why patients with monosomy 7 are rarely diagnosed with high age-adjusted HbF levels. In addition, this new fetal-like JMML subgroup presented with reduced levels of most members of the let-7 microRNA family and showed characteristic overexpression of genes involved in fetal hematopoiesis and stem cell self-renewal. Lastly, high LIN28B expression was associated with poor clinical outcome in our JMML patient series but was not independent from other prognostic factors such as age and age-adjusted HbF levels. In conclusion, we identified elevated LIN28B expression as a hallmark of a novel fetal-like subgroup in JMML.

Prevalence, clinical characteristics, and prognosis of GATA2-related myelodysplastic syndromes in children and adolescents.

Germline GATA2 mutations cause cellular deficiencies with high propensity for myeloid disease. We investigated 426 children and adolescents with primary myelodysplastic syndrome (MDS) and 82 cases with secondary MDS enrolled in 2 consecutive prospective studies of the European Working Group of MDS in Childhood (EWOG-MDS) conducted in Germany over a period of 15 years. Germline GATA2 mutations accounted for 15% of advanced and 7% of all primary MDS cases, but were absent in children with MDS secondary to therapy or acquired aplastic anemia. Mutation carriers were older at diagnosis and more likely to present with monosomy 7 and advanced disease compared with wild-type cases. For stratified analysis according to karyotype, 108 additional primary MDS patients registered with EWOG-MDS were studied. Overall, we identified 57 MDS patients with germline GATA2 mutations. GATA2 mutations were highly prevalent among patients with monosomy 7 (37%, all ages) reaching its peak in adolescence (72% of adolescents with monosomy 7). Unexpectedly, monocytosis was more frequent in GATA2-mutated patients. However, when adjusted for the selection bias from monosomy 7, mutational status had no effect on the hematologic phenotype. Finally, overall survival and outcome of hematopoietic stem cell transplantation (HSCT) were not influenced by mutational status. This study identifies GATA2 mutations as the most common germline defect predisposing to pediatric MDS with a very high prevalence in adolescents with monosomy 7. GATA2 mutations do not confer poor prognosis in childhood MDS. However, the high risk for progression to advanced disease must guide decision-making toward timely HSCT.

Morphological differentiation of severe aplastic anaemia from hypocellular refractory cytopenia of childhood: reproducibility of histopathological diagnostic criteria.

AIMS: To evaluate the reproducibility and reliability of the histomorphological criteria differentiating severe aplastic anaemia (SAA) and hypoplastic refractory cytopenia of childhood (RCC), the most frequently acquired hypocellular bone marrow conditions of childhood. METHODS AND RESULTS: We performed a double-blind interobserver study of 100 different cases of SAA and RCC among seven haematopathologists of the European Working Group of MDS in Childhood (EWOG-MDS) and the German SAA study. Cases with foci of typical myelodysplastic syndrome (MDS) morphology, such as patchy erythropoiesis with defective maturation, in an otherwise highly hypocellular or adipocytic bone marrow were classified as having RCC. Bone marrow samples without a patchy distribution, few scattered myeloid cells or haematopoietic aplasia were diagnosed as SAA. In only four of 100 cases did the reference pathologists not reach agreement regarding classification as SAA or RCC. The kappa index was 0.79. CONCLUSIONS: Our results show that the vast majority of SAA and RCC cases can be reliably differentiated by morphological means alone. A clear differentiation between SAA and RCC at presentation is mandatory for optimizing therapy strategies, and might be responsible for the fact that, in the German childhood SAA study, the probability of developing clonal disease after immunosuppressive therapy has dropped to 3%.