We evaluated the frequency, genetic architecture, clinico-pathologic features and prognostic impact of RUNX1 mutations in 2439 adult patients with newly-diagnosed acute myeloid leukemia (AML). RUNX1 mutations were found in 245 of 2439 (10%) patients; were almost mutually exclusive of AML with recurrent genetic abnormalities; and they co-occurred with a complex pattern of gene mutations, frequently involving mutations in epigenetic modifiers (ASXL1, IDH2, KMT2A, EZH2), components of the spliceosome complex (SRSF2, SF3B1) and STAG2, PHF6, BCOR. RUNX1 mutations were associated with older age (16-59 years: 8.5%; ⩾60 years: 15.1%), male gender, more immature morphology and secondary AML evolving from myelodysplastic syndrome. In univariable analyses, RUNX1 mutations were associated with inferior event-free (EFS, P<0.0001), relapse-free (RFS, P=0.0007) and overall survival (OS, P<0.0001) in all patients, remaining significant when age was considered. In multivariable analysis, RUNX1 mutations predicted for inferior EFS (P=0.01). The effect of co-mutation varied by partner gene, where patients with the secondary genotypes RUNX1/ASXL1 (OS, P=0.004), RUNX1/SRSF2 (OS, P=0.007) and RUNX1/PHF6 (OS, P=0.03) did significantly worse, whereas patients with the genotype RUNX1/IDH2 (OS, P=0.04) had a better outcome. In conclusion, RUNX1-mutated AML is associated with a complex mutation cluster and is correlated with distinct clinico-pathologic features and inferior prognosis.

Single nucleotide polymorphisms (SNPs) of the NOD2/CARD15 gene resulting in a diminished nuclear factor-B (NF-B) response to bacterial cell wall products have been associated with an increased incidence of Crohn disease. To assess a possible contribution of NOD2/CARD15 mutations to graft-versus-host disease (GvHD) and complications following allogeneic stem cell transplantation, we retrospectively typed DNA from donor/recipient pairs in 169 consecutive patients receiving transplants from related or unrelated donors. Mutated alleles were observed in 21% of patients and in 14% of donors. Cumulative incidence of 1-year, transplant-related mortality rose from 20% in donor/recipient pairs without mutated SNPs to 49% in pairs with recipient mutations (P ‫؍‬ .03) and 59% in pairs with donor mutations (P < .005), and was highest in 12 pairs with mutated alleles in both donor and recipients (83%; P < .001). Similar associations were observed for severe overall and severe gastrointestinal GvHD. The impact of NOD2/CARD15 mutations was more prominent for HLAidentical sibling transplantations but was also observed in unrelated donor transplantation. Mutations proved to be independent risk factors for transplant-related mortality. Our findings indicate a major role of monocyte/macrophage dysfunction in the pathophysiology of GvHD and strongly suggest a future risk assessment or even donor selection through NOD2/CARD15 typing. (Blood. 2004;104: 889-894)

Key Points• Urinary 3-IS levels predict outcome after ASCT and are associated with antibiotics and NOD2/CARD15 variants.Indole, which is produced from L-tryptophan by commensal bacteria expressing tryptophanase, not only is an important intercellular signal in microbial communities, but also modulates mucosal barrier function and expression of pro-and anti-inflammatory genes by intestinal epithelial cells. Here, we hypothesized that decreased urinary excretion of 3-indoxyl sulfate (3-IS), the major conjugate of indole found in humans, may be a marker of gut microbiota disruption and increased risk of developing gastrointestinal (GI) graftversus-host-disease. Using liquid chromatography/tandem mass spectrometry, 3-IS was determined in urine specimens collected weekly within the first 28 days after allogeneic stem cell transplantation (ASCT) in 131 patients. Low 3-IS levels within the first 10 days after ASCT were associated with significantly higher transplant-related mortality (P 5 .017) and worse overall survival (P 5 .05) 1 year after ASCT. Least absolute shrinkage and selection operator regression models trained on lognormalized counts of 763 operational taxonomic units derived from next-generation sequencing of the hypervariable V3 region of the 16S ribosomal RNA gene showed members of the families of Lachnospiraceae and Ruminococcaceae of the class of Clostridia to be associated with high urinary 3-IS levels, whereas members of the class of Bacilli were associated with low 3-IS levels. Risk factors of early suppression of 3-IS levels were the type of GI decontamination (P 5 .01), early onset of antibiotic treatment (P 5 .001), and recipient NOD2/ CARD15 genotype (P 5 .04). In conclusion, our findings underscore the relevance of microbiota-derived indole and metabolites thereof in mucosal integrity and protection from inflammation. (Blood. 2015;126(14):1723-1728) IntroductionAllogeneic stem cell transplantation (ASCT) constitutes a potential curative therapy for various hematologic malignancies, bone marrow failure, and immune deficiency syndromes. However, this treatment is still associated with a high risk of mortality because of infectious complications and acute graft-versus-host disease (GVHD). A significant part of these severe complications originates from the gastrointestinal (GI) tract. 1The introduction of 16S ribosomal RNA (rRNA) sequencing has provided novel insights into the diversity and complexity of the gut ecosystem.2 Loss of a diverse composition of the microbiome has been associated with a variety of diseases including inflammatory bowel and autoimmune diseases. At least in part this may be because of the increasingly recognized role that commensal bacteria play in maintaining immunologic homeostasis and epithelial integrity and in exerting anti-inflammatory effects and intestinal tolerance by inducing regulatory T cells. 3,4 Recent studies have demonstrated an association between intestinal bacterial diversity in both mouse models and humans and outcome of ASCT.5-7 A significantly higher ...

In allogeneic stem cell transplantation (ASCT), systemic broad-spectrum antibiotics are frequently used for treatment of infectious complications, but their effect on microbiota composition is still poorly understood. Here, in a retrospective analysis of 621 patients, who underwent ASCT at the University Medical Center of Regensburg and Memorial Sloan Kettering Cancer Center in New York, we assessed the impact of timing of peri-transplant antibiotic treatment on intestinal microbiota composition as well as transplant-related mortality (TRM) and overall survival. Early exposure to antibiotics was associated with lower urinary 3-indoxyl sulfate levels (p<0.001) and a decrease in fecal abundance of commensal Clostridiales (p=0.03) compared to late antibiotic treatment, which was particularly significant (p=0.005) for Clostridium cluster XIVa in the Regensburg group. Earlier antibiotic treatment prior to ASCT was further associated with a higher TRM (34%, n=79/236) compared to post-ASCT (21% n=62/297, p=0.001) or no antibiotics (7% n=6/88, p<0.001). Timing of antibiotic treatment was the dominant independent risk factor for TRM (HR 2.0, p≤0.001) in multivariate analysis beside increase age (HR 2.15, p=0.004), reduced Karnofsky performance status (HR 1.47, p=0.03) and female donor/ male recipient sex combination (HR 1.56, p=0.02) A competing-risk analysis revealed the independent effect of early initiation of antibiotics on GvHD-related TRM (p=0.004) in contrast to infection-related TRM and relapse (p=ns). The poor outcome associated with early administration of antibiotic therapy that is active against commensal organisms, and specifically the possibly protective Clostridiales calls for the use of Clostridiales-sparing antibiotics and rapid restoration of microbiota diversity after cessation of antibiotic treatment.

The JAK2-V617F mutation occurs in about 50% of patients with myelofibrosis and might be a reliable marker to monitor residual disease after allogeneic stem cell transplantation. We describe a new, highly sensitive (> 0.01%) real-time polymerase chain reaction (PCR) to monitor and quantify V617F-JAK2-positive cells after dose-reduced allogeneic stem cell transplantation. After 22 allogeneic stem cell transplantation procedures in 21 JAK2-positive patients with myelofibrosis, 78% became PCR negative. In 15 of 17 patients (88%), JAK2 remained negative after a median follow-up of 20 months. JAK2 negativity was achieved after a median of 89 days after allograft (range, 19-750 days). A significant inverse correlation was seen for JAK2 positivity and donor-cell chimerism (r : ؊0.91, P < .001). IntroductionMyelofibrosis with myeloid metaplasia (MMM) is a myeloproliferative disease of the multipotent hematopoietic progenitor cells, leading to an inappropriate release of fibrogenic cytokines or growth factors in the bone marrow, thus inducing a fibrosis of extracellular bone marrow texture and extramedullary hematopoiesis. [1][2][3][4][5] Allogeneic stem cell transplantation is the only curative approach in patients with myelofibrosis, since no other therapeutic strategy resulted in a complete resolution of fibrosis content. [6][7][8] After standard conditioning, 5-year survival rates of 47% and 58% have been reported. However, the treatment-related mortality rates in those studies were rather high, with 27% 8 and 33%. 7 To reduce the high treatment-related mortality after stem cell transplantation and to perform this treatment approach in elderly patients, so-called nonmyeloablative (ie, dose-reduced) conditioning regimens were introduced and have shown encouraging preliminary results in patients with myelofibrosis. 9-10 The establishment of valid complete remission criteria for myelofibrosis, especially after allogeneic stem cell transplantation, remains a major issue. The criteria for complete remission recently proposed by the International Working Group for Myelofibrosis Research and Treatment (IWG-MRT) include the disappearance of disease-related syndromes, peripheral blood levels of hemoglobin greater than or equal to 110 g/L (11 g/dL) and platelet counts greater than or equal to 100 ϫ 10 9 /L. 11 After allogeneic stem cell transplantation, these parameters are often influenced by graft-versus-host disease, infections, or poor graft function, and they cannot be used as valid remission criteria. On the other hand, normal blood counts and disappearance of disease-related syndromes do not exclude residual disease. For other diseases such as chronic myeloid leukemia (CML), myeloma, and chronic lymphocytic leukemia (CLL), the monitoring of molecular remission after allogeneic stem cell transplantation is based on quantifying specific genetic markers such as bcr/abl transcripts or patient-specific rearrangements. The available data suggest that achievement of molecular remission results in a long-term freedom from disea...