No abstract

To understand the mechanisms that mediate germline genetic leukemia predisposition, we studied the inherited ribosomopathy Shwachman-Diamond syndrome (SDS), a bone marrow failure disorder with high risk of myeloid malignancies at an early age. To define the mechanistic basis of clonal hematopoiesis in SDS, we investigate somatic mutations acquired by patients with SDS followed longitudinally. Here we report that multiple independent somatic hematopoietic clones arise early in life, most commonly harboring heterozygous mutations in EIF6 or TP53. We show that germline SBDS deficiency establishes a fitness constraint that drives selection of somatic clones via two distinct mechanisms with different clinical consequences. EIF6 inactivation mediates a compensatory pathway with limited leukemic potential by ameliorating the underlying SDS ribosome defect and enhancing clone fitness. TP53 mutations define a maladaptive pathway with enhanced leukemic potential by inactivating tumor suppressor checkpoints without correcting the ribosome defect. Subsequent development of leukemia was associated with acquisition of biallelic TP53 alterations. These results mechanistically link leukemia predisposition to germline genetic constraints on cellular fitness, and provide a rational framework for clinical surveillance strategies.

Targeted mutagenesis was used to produce two mutations in the murine hemochromatosis gene (Hfe) locus. The first mutation deletes a large portion of the coding sequence, generating a null allele. The second mutation introduces a missense mutation (C282Y) into theHfe locus, but otherwise leaves the gene intact. This mutation is identical to the disease-causing mutation in patients with hereditary hemochromatosis. Mice carrying each of the two mutations were bred and analyzed. Homozygosity for either mutation results in postnatal iron loading. The effects of the null mutation are more severe than the effects of the C282Y mutation. Mice heterozygous for either mutation accumulate more iron than normal controls. Interestingly, although liver iron stores are greatly increased, splenic iron is decreased. We conclude that the C282Y mutation does not result in a null allele.

Microcytic anemia (mk) mice and Belgrade (b) rats have severe iron deficiency anemia due to defects in intestinal iron transport and erythroid iron utilization. Both animal mutants carry the same missense mutation in Nramp2, the first mammalian iron transporter to be identified. This mutation, in which glycine 185 is changed to arginine (G185R), occurs within predicted transmembrane domain 4 of the protein. We have performed site-directed mutagenesis of murine Nramp2, focusing on amino acids of transmembrane domain 4 that are highly conserved among Nramp-like proteins. We have expressed each mutant form in transfected cells and examined iron transport function, subcellular localization, and protein amounts. All tested forms of Nramp2 localize to the plasma membrane and to transferrin-containing endosomes. Most transmembrane domain 4 mutations affect the amount of protein detected and consequently show diminished iron transport. The G185R mutation, however, causes near total loss of Nramp2 function that cannot be fully explained by a decreased amount of protein, indicating that G185R disrupts iron transport through an alteration in the function of Nramp2, rather than degradation of the protein. © 1998 by The American Society of Hematology.

SUMMARY SF3B1 is recurrently mutated in chronic lymphocytic leukemia (CLL), but its role in the pathogenesis of CLL remain elusive. Here, we show that conditional expression of Sf3b1-K700E mutation in mouse B cells disrupts pre-mRNA splicing, alters cell development, and induces a state of cellular senescence. Combination with Atm deletion leads to the overcoming of cellular senescence and the development of CLL-like disease in elderly mice. These CLL-like cells show genome instability and dysregulation of multiple CLL-associated cellular processes, including deregulated B cell receptor (BCR) signaling, which we also identified in human CLL cases. Notably, human CLLs harboring SF3B1 mutations exhibit altered response to BTK inhibition. Our murine model of CLL thus provides insights into human CLL disease mechanisms and treatment.

Summary Overlapping clinical phenotypes and an expanding breadth and complexity of genomic associations are a growing challenge in the diagnosis and clinical management of Mendelian disorders. The functional consequences and clinical impacts of genomic variation may involve unique, disorder-specific, genomic DNA methylation episignatures. In this study, we describe 19 novel episignature disorders and compare the findings alongside 38 previously established episignatures for a total of 57 episignatures associated with 65 genetic syndromes. We demonstrate increasing resolution and specificity ranging from protein complex, gene, sub-gene, protein domain, and even single nucleotide-level Mendelian episignatures. We show the power of multiclass modeling to develop highly accurate and disease-specific diagnostic classifiers. This study significantly expands the number and spectrum of disorders with detectable DNA methylation episignatures, improves the clinical diagnostic capabilities through the resolution of unsolved cases and the reclassification of variants of unknown clinical significance, and provides further insight into the molecular etiology of Mendelian conditions.

3158 MicroRNA miR-125b has been shown to be involved in different kind of leukemia. Indeed, the chromosomal translocation t(2;11)(p21;q23) found in patients with myelodysplasia and acute myeloid leukemia leads to an overexpression of miR-125b up to 90 fold. Moreover, miR-125b is also upregulated in patients with B-cell acute lymphoblastic leukemia carrying the t(11;14)(q24;q32) translocation. To decipher the presumed oncogenic mechanism of miR-125b, we used transplantation experiments in mice. All of the mice transplanted with fetal liver cells ectopically expressing miR-125b showed an increase in white blood cell count, in particular in neutrophils and monocytes, associated with a macrocytic anemia. Among these mice, half of them died of B-cell acute lymphoblastic leukemia, T-cell acute lymphoblastic leukemia, or a myeloproliferative disorder, suggesting an important role of miR-125b in myeloid and lymphoid lineages. Co-expression of miR-125b and the BCR-ABL fusion gene in transplanted cells accelerated the development of leukemia in mice, compared to control mice expressing only BCR-ABL, suggesting that miR-125b confers a proliferative advantage to the leukemic cells. Thus we showed that the overexpression of miR-125b is sufficient to induce leukemia in vivo and decrease the latency of BCR-ABL -induced leukemia. Disclosures: No relevant conflicts of interest to declare.

Our study failed to prove the superiority of ER over OR. If anatomy is suitable, ER of PAA in the elderly and high-risk patients is justified. For emergent PAA repairs, MAEs are frequent after both ER and OR; ER has not changed the severe prognosis of acute limb ischemia from PAA. A multicenter randomized controlled trial of PAA patients with acute presentation is warranted.