Identification and surveillance of rare relapse-initiating stem cells during complete remission after transplantation.

ABSTRACT: Relapse after complete remission (CR) remains the main cause of mortality after allogeneic stem cell transplantation for hematological malignancies and, therefore, improved biomarkers for early prediction of relapse remains a critical goal toward development and assessment of preemptive relapse treatment. Because the significance of cancer stem cells as a source of relapses remains unclear, we investigated whether mutational screening for persistence of rare cancer stem cells would enhance measurable residual disease (MRD) and early relapse prediction after transplantation. In a retrospective study of patients who relapsed and patients who achieved continuous-CR with myelodysplastic syndromes and related myeloid malignancies, combined flow cytometric cell sorting and mutational screening for persistence of rare relapse-initiating stem cells was performed in the bone marrow at multiple CR time points after transplantation. In 25 CR samples from 15 patients that later relapsed, only 9 samples were MRD-positive in mononuclear cells (MNCs) whereas flowcytometric-sorted hematopoietic stem and progenitor cells (HSPCs) were MRD-positive in all samples, and always with a higher variant allele frequency than in MNCs (mean, 97-fold). MRD-positivity in HSPCs preceded MNCs in multiple sequential samples, in some cases preceding relapse by >2 years. In contrast, in 13 patients in long-term continuous-CR, HSPCs remained MRD-negative. Enhanced MRD sensitivity was also observed in total CD34+ cells, but HSPCs were always more clonally involved (mean, 8-fold). In conclusion, identification of relapse-initiating cancer stem cells and mutational MRD screening for their persistence consistently enhances MRD sensitivity and earlier prediction of relapse after allogeneic stem cell transplantation.

Hybrid approach for harmony transcatheter pulmonary valve replacement.

The Harmony™ Transcatheter Pulmonary Valve (Medtronic) was recently approved by the Food and Drug Administration for transcatheter pulmonary valve replacement in native right ventricular outflow tracts. Despite this milestone, some patients have main pulmonary arteries that are severely dilated and continue to require surgical pulmonary valve replacement. The hybrid approach combines surgical creation of a landing zone, transcatheter valve deployment, and suture stabilization of the implanted valve. In this case series, we report the first use of a hybrid approach for Harmony™ transcatheter pulmonary valve replacement. Two cases are reported with varying approaches for surgical creation of a landing zone followed by successful placement of a Harmony™ valve.

Carbon dioxide separation and capture by adsorption: a review.

Rising adverse impact of climate change caused by anthropogenic activities is calling for advanced methods to reduce carbon dioxide emissions. Here, we review adsorption technologies for carbon dioxide capture with focus on materials, techniques, and processes, additive manufacturing, direct air capture, machine learning, life cycle assessment, commercialization and scale-up.

Towards enhancement of gas-liquid mass transfer in bioelectrochemical systems: Validation of a robust CFD model.

Mass transfer has been identified as a major bottleneck in gas fermentation and microbial conversion of carbon dioxide to chemicals. We present a pragmatic and validated Computational Fluid Dynamics (CFD) model for mass transfer in bioelectrochemical systems. Experiments were conducted to measure mixing times and mass transfer in a Duran bottle and an H-cell. An Eulerian-Eulerian framework with a simplified model for the bubble size distribution (BSD) was developed that utilized only one additional equation for the bubble number density while including the breakup and coalescence. Validations of the CFD model for mixing times showed that the predictions were within the confidence intervals of the measurements, verifying the model's capability in simulating the hydrodynamics. Further validations were performed using constant and varying bubble diameters for the mass transfer. The results showed the benefits of a simplified BSD model, as it yielded improvements of seven and four times in accuracy when assessed against the experimental data for the Duran bottle and H-cell, respectively. Modeling of the H-cell predicted that a lower stirring rate improves mass transfer compared with higher stirring rates, which is of great importance when designing microbial cultivation processes. The model offers a feasible framework for advanced modeling of gas fermentation and microbial electrosynthesis.

appreci8: a pipeline for precise variant calling integrating 8 tools.

Motivation: The application of next-generation sequencing in research and particularly in clinical routine requires valid variant calling results. However, evaluation of several commonly used tools has pointed out that not a single tool meets this requirement. False positive as well as false negative calls necessitate additional experiments and extensive manual work. Intelligent combination and output filtration of different tools could significantly improve the current situation. Results: We developed appreci8, an automatic variant calling pipeline for calling single nucleotide variants and short indels by combining and filtering the output of eight open-source variant calling tools, based on a novel artifact- and polymorphism score. Appreci8 was trained on two data sets from patients with myelodysplastic syndrome, covering 165 Illumina samples. Subsequently, appreci8's performance was tested on five independent data sets, covering 513 samples. Variation in sequencing platform, target region and disease entity was considered. All calls were validated by re-sequencing on the same platform, a different platform or expert-based review. Sensitivity of appreci8 ranged between 0.93 and 1.00, while positive predictive value ranged between 0.65 and 1.00. In all cases, appreci8 showed superior performance compared to any evaluated alternative approach. Availability and implementation: Appreci8 is freely available at https://hub.docker.com/r/wwuimi/appreci8/. Sequencing data (BAM files) of the 678 patients analyzed with appreci8 have been deposited into the NCBI Sequence Read Archive (BioProjectID: 388411; https://www.ncbi.nlm.nih.gov/bioproject/PRJNA388411). Supplementary information: Supplementary data are available at Bioinformatics online.

Gene expression and risk of leukemic transformation in myelodysplasia.

Myelodysplastic syndromes (MDSs) are a heterogeneous group of clonal hematopoietic disorders with a highly variable prognosis. To identify a gene expression-based classification of myelodysplasia with biological and clinical relevance, we performed a comprehensive transcriptomic analysis of myeloid neoplasms with dysplasia using transcriptome sequencing. Unsupervised clustering of gene expression data of bone marrow CD34+ cells from 100 patients identified 2 subgroups. The first subtype was characterized by increased expression of genes related to erythroid/megakaryocytic (EMK) lineages, whereas the second subtype showed upregulation of genes related to immature progenitor (IMP) cells. Compared with the first so-called EMK subtype, the IMP subtype showed upregulation of many signaling pathways and downregulation of several pathways related to metabolism and DNA repair. The IMP subgroup was associated with a significantly shorter survival in both univariate (hazard ratio [HR], 5.0; 95% confidence interval [CI], 1.8-14; P = .002) and multivariate analysis (HR, 4.9; 95% CI, 1.3-19; P = .02). Leukemic transformation was limited to the IMP subgroup. The prognostic significance of our classification was validated in an independent cohort of 183 patients. We also constructed a model to predict the subgroups using gene expression profiles of unfractionated bone marrow mononuclear cells (BMMNCs). The model successfully predicted clinical outcomes in a test set of 114 patients with BMMNC samples. The addition of our classification to the clinical model improved prediction of patient outcomes. These results indicated biological and clinical relevance of our gene expression-based classification, which will improve risk prediction and treatment stratification of MDS.

SF3B1-initiating mutations in MDS-RSs target lymphomyeloid hematopoietic stem cells.

Mutations in the RNA splicing gene SF3B1 are found in >80% of patients with myelodysplastic syndrome with ring sideroblasts (MDS-RS). We investigated the origin of SF3B1 mutations within the bone marrow hematopoietic stem and progenitor cell compartments in patients with MDS-RS. Screening for recurrently mutated genes in the mononuclear cell fraction revealed mutations in SF3B1 in 39 of 40 cases (97.5%), combined with TET2 and DNMT3A in 11 (28%) and 6 (15%) patients, respectively. All recurrent mutations identified in mononuclear cells could be tracked back to the phenotypically defined hematopoietic stem cell (HSC) compartment in all investigated patients and were also present in downstream myeloid and erythroid progenitor cells. While in agreement with previous studies, little or no evidence for clonal (SF3B1 mutation) involvement could be found in mature B cells, consistent involvement at the pro-B-cell progenitor stage was established, providing definitive evidence for SF3B1 mutations targeting lymphomyeloid HSCs and compatible with mutated SF3B1 negatively affecting lymphoid development. Assessment of stem cell function in vitro as well as in vivo established that only HSCs and not investigated progenitor populations could propagate the SF3B1 mutated clone. Upon transplantation into immune-deficient mice, SF3B1 mutated MDS-RS HSCs differentiated into characteristic ring sideroblasts, the hallmark of MDS-RS. Our findings provide evidence of a multipotent lymphomyeloid HSC origin of SF3B1 mutations in MDS-RS patients and provide a novel in vivo platform for mechanistically and therapeutically exploring SF3B1 mutated MDS-RS.

SF3B1 mutation identifies a distinct subset of myelodysplastic syndrome with ring sideroblasts.

Refractory anemia with ring sideroblasts (RARS) is a myelodysplastic syndrome (MDS) characterized by isolated erythroid dysplasia and 15% or more bone marrow ring sideroblasts. Ring sideroblasts are found also in other MDS subtypes, such as refractory cytopenia with multilineage dysplasia and ring sideroblasts (RCMD-RS). A high prevalence of somatic mutations of SF3B1 was reported in these conditions. To identify mutation patterns that affect disease phenotype and clinical outcome, we performed a comprehensive mutation analysis in 293 patients with myeloid neoplasm and 1% or more ring sideroblasts. SF3B1 mutations were detected in 129 of 159 cases (81%) of RARS or RCMD-RS. Among other patients with ring sideroblasts, lower prevalence of SF3B1 mutations and higher prevalence of mutations in other splicing factor genes were observed (P < .001). In multivariable analyses, patients with SF3B1 mutations showed significantly better overall survival (hazard ratio [HR], .37; P = .003) and lower cumulative incidence of disease progression (HR = 0.31; P = .018) compared with SF3B1-unmutated cases. The independent prognostic value of SF3B1 mutation was retained in MDS without excess blasts, as well as in sideroblastic categories (RARS and RCMD-RS). Among SF3B1-mutated patients, coexisting mutations in DNA methylation genes were associated with multilineage dysplasia (P = .015) but had no effect on clinical outcome. TP53 mutations were frequently detected in patients without SF3B1 mutation, and were associated with poor outcome. Thus, SF3B1 mutation identifies a distinct MDS subtype that is unlikely to develop detrimental subclonal mutations and is characterized by indolent clinical course and favorable outcome.

Progression in patients with low- and intermediate-1-risk del(5q) myelodysplastic syndromes is predicted by a limited subset of mutations.

A high proportion of patients with lower-risk del(5q) myelodysplastic syndromes will respond to treatment with lenalidomide. The median duration of transfusion-independence is 2 years with some long-lasting responses, but almost 40% of patients progress to acute leukemia by 5 years after starting treatment. The mechanisms underlying disease progression other than the well-established finding of small TP53-mutated subclones at diagnosis remain unclear. We studied a longitudinal cohort of 35 low- and intermediate-1-risk del(5q) patients treated with lenalidomide (n=22) or not (n=13) by flow cytometric surveillance of hematopoietic stem and progenitor cell subsets, targeted sequencing of mutational patterns, and changes in the bone marrow microenvironment. All 13 patients with disease progression were identified by a limited number of mutations in TP53, RUNX1, and TET2, respectively, with PTPN11 and SF3B1 occurring in one patient each. TP53 mutations were found in seven of nine patients who developed acute leukemia, and were documented to be present in the earliest sample (n=1) and acquired during lenalidomide treatment (n=6). By contrast, analysis of the microenvironment, and of hematopoietic stem and progenitor cells by flow cytometry was of limited prognostic value. Based on our data, we advocate conducting a prospective study aimed at investigating, in a larger number of cases of del(5q) myelodysplastic syndromes, whether the detection of such mutations before and after lenalidomide treatment can guide clinical decision-making.

Analysis of the DNA methylome and transcriptome in granulopoiesis reveals timed changes and dynamic enhancer methylation.

In development, epigenetic mechanisms such as DNA methylation have been suggested to provide a cellular memory to maintain multipotency but also stabilize cell fate decisions and direct lineage restriction. In this study, we set out to characterize changes in DNA methylation and gene expression during granulopoiesis using 4 distinct cell populations ranging from the oligopotent common myeloid progenitor stage to terminally differentiated neutrophils. We observed that differentially methylated sites (DMSs) generally show decreased methylation during granulopoiesis. Methylation appears to change at specific differentiation stages and overlap with changes in transcription and activity of key hematopoietic transcription factors. DMSs were preferentially located in areas distal to CpG islands and shores. Also, DMSs were overrepresented in enhancer elements and enriched in enhancers that become active during differentiation. Overall, this study depicts in detail the epigenetic and transcriptional changes that occur during granulopoiesis and supports the role of DNA methylation as a regulatory mechanism in blood cell differentiation.

Aberrant splicing of genes involved in haemoglobin synthesis and impaired terminal erythroid maturation in SF3B1 mutated refractory anaemia with ring sideroblasts.

Refractory anaemia with ring sideroblasts (RARS) is distinguished by hyperplastic inefficient erythropoiesis, aberrant mitochondrial ferritin accumulation and anaemia. Heterozygous mutations in the spliceosome gene SF3B1 are found in a majority of RARS cases. To explore the link between SF3B1 mutations and anaemia, we studied mutated RARS CD34(+) marrow cells with regard to transcriptome sequencing, splice patterns and mutational allele burden during erythroid differentiation. Transcriptome profiling during early erythroid differentiation revealed a marked up-regulation of genes involved in haemoglobin synthesis and in the oxidative phosphorylation process, and down-regulation of mitochondrial ABC transporters compared to normal bone marrow. Moreover, mis-splicing of genes involved in transcription regulation, particularly haemoglobin synthesis, was confirmed, indicating a compromised haemoglobinization during RARS erythropoiesis. In order to define the phase during which erythroid maturation of SF3B1 mutated cells is most affected, we assessed allele burden during erythroid differentiation in vitro and in vivo and found that SF3B1 mutated erythroblasts showed stable expansion until late erythroblast stage but that terminal maturation to reticulocytes was significantly reduced. In conclusion, SF3B1 mutated RARS progenitors display impaired splicing with potential downstream consequences for genes of key importance for haemoglobin synthesis and terminal erythroid differentiation.

Global changes in DNA methylation in Alzheimer's disease peripheral blood mononuclear cells.

Changes in epigenetic marks may help explain the late onset of Alzheimer's disease (AD). In this study we measured genome-wide DNA methylation by luminometric methylation assay, a quantitative measurement of genome-wide DNA methylation, on DNA isolated from peripheral blood mononuclear cells of 37 subjects with late-onset AD (LOAD) and 44 healthy controls (CT). We found an increase in global DNA methylation in LOAD subjects compared to CT (p=0.0122), associated with worse cognitive performances (p=0.0002). DNA hypermethylation in LOAD group was paralleled by higher DNA methyltransferase 1 (DNMT1) gene expression and protein levels. When data were stratified on the basis of the APOE polymorphisms, higher DNA methylation levels were associated with the presence of APOE ε4 allele (p=0.0043) in the global population. Among the APOE ε3 carriers, a significant increase of DNA methylation was still observed in LOAD patients compared to healthy controls (p=0.05). Our data suggest global DNA methylation in peripheral samples as a useful marker for screening individuals at risk of developing AD.

p53 protein expression independently predicts outcome in patients with lower-risk myelodysplastic syndromes with del(5q).

Del(5q) myelodysplastic syndromes defined by the International Prognostic Scoring System as low- or intermediate-1-risk (lower-risk) are considered to have an indolent course; however, recent data have identified a subgroup of these patients with more aggressive disease and poorer outcomes. Using deep sequencing technology, we previously demonstrated that 18% of patients with lower-risk del(5q) myelodysplastic syndromes carry TP53 mutated subclones rendering them at higher risk of progression. In this study, bone marrow biopsies from 85 patients treated with lenalidomide in the MDS-004 clinical trial were retrospectively assessed for p53 expression by immunohistochemistry in association with outcome. Strong p53 expression in ≥ 1% of bone marrow progenitor cells, observed in 35% (30 of 85) of patients, was significantly associated with higher acute myeloid leukemia risk (P=0.0006), shorter overall survival (P=0.0175), and a lower cytogenetic response rate (P=0.009), but not with achievement or duration of 26-week transfusion independence response. In a multivariate analysis, p53-positive immunohistochemistry was the strongest independent predictor of transformation to acute myeloid leukemia (P=0.0035). Pyrosequencing analysis of laser-microdissected cells with strong p53 expression confirmed the TP53 mutation, whereas cells with moderate expression predominantly had wild-type p53. This study validates p53 immunohistochemistry as a strong and clinically useful predictive tool in patients with lower-risk del(5q) myelodysplastic syndromes. This study was based on data from the MDS 004 trial (clinicaltrials.gov identifier: NCT00179621).

Hypothalamic mitochondrial dysfunction associated with anorexia in the anx/anx mouse.

The anorectic anx/anx mouse exhibits disturbed feeding behavior and aberrances, including neurodegeneration, in peptidergic neurons in the appetite regulating hypothalamic arcuate nucleus. Poor feeding in infants, as well as neurodegeneration, are common phenotypes in human disorders caused by dysfunction of the mitochondrial oxidative phosphorylation system (OXPHOS). We therefore hypothesized that the anorexia and degenerative phenotypes in the anx/anx mouse could be related to defects in the OXPHOS. In this study, we found reduced efficiency of hypothalamic OXPHOS complex I assembly and activity in the anx/anx mouse. We also recorded signs of increased oxidative stress in anx/anx hypothalamus, possibly as an effect of the decreased hypothalamic levels of fully assembled complex I, that were demonstrated by native Western blots. Furthermore, the Ndufaf1 gene, encoding a complex I assembly factor, was genetically mapped to the anx interval and found to be down-regulated in anx/anx mice. These results suggest that the anorexia and hypothalamic neurodegeneration of the anx/anx mouse are associated with dysfunction of mitochondrial complex I.

Congestive Heart Failure in an Adolescent With a Ruptured Sinus of Valsalva Aneurysm.

Sinus of Valsalva aneurysm (SVA) is an abnormal dilatation of the aortic root located between the aortic valve annulus and the sinotubular junction and is rare in the pediatric population. This case report describes a unique case of a 16-year-old adolescent patient admitted with progressive heart failure symptoms and diagnosed with a ruptured noncoronary SVA. He underwent surgical repair of the SVA with autologous pericardial patches and had an uncomplicated postoperative course. A genetic workup revealed an underlying 22q11.2 deletion that is infrequently associated with SVA.

Prognostic DNA methylation patterns in cytogenetically normal acute myeloid leukemia are predefined by stem cell chromatin marks.

Cytogenetically normal acute myeloid leukemia (CN-AML) compose between 40% and 50% of all adult acute myeloid leukemia (AML) cases. In this clinically diverse group, molecular aberrations, such as FLT3-ITD, NPM1, and CEBPA mutations, recently have added to the prognostic accuracy. Aberrant DNA methylation is a hallmark of cancer, including AML. We investigated in total 118 CN-AML samples in a test and a validation cohort for genome-wide promoter DNA methylation with Illumina Methylation Bead arrays and compared them with normal myeloid precursors and global gene expression. IDH and NPM1 mutations were associated with different methylation patterns (P = .0004 and .04, respectively). Genome-wide methylation levels were elevated in IDH-mutated samples (P = .006). We observed a negative impact of DNA methylation on transcription. Genes targeted by Polycomb group (PcG) proteins and genes associated with bivalent histone marks in stem cells showed increased aberrant methylation in AML (P < .0001). Furthermore, high methylation levels of PcG target genes were independently associated with better progression-free survival (odds ratio = 0.47, P = .01) and overall survival (odds ratio = 0.36, P = .001). In summary, genome-wide methylation patterns show preferential methylation of PcG targets with prognostic impact in CN-AML.

Damus-Kaye-Stansel procedure 5 years after Fontan operation with ligated main pulmonary artery.

We report a case of tricuspid atresia with transposed great arteries and rudimentary right ventricle owing to which the patient developed severe subaortic stenosis and restrictive bulboventricular foramen 5 years after her extracardiac Fontan operation. She underwent a successful modified Damus-Kaye-Stansel operation using her native pulmonary valve. Spiral cardiac computed tomography with three-dimensional reconstructions was instrumental in pre-operative surgical planning.

Detection of Lung Tumors in CT Scan Images using Convolutional Neural Networks.

Changing the human being's lifestyle, has caused, or exacerbated many diseases. One of these diseases is cancer, and among all kind of cancers like, brain and pulmonary; lungs cancer is fatal. The cancers could be detected early to save lives using Computer Aided Diagnosis (CAD) systems. CT scans medical images are one the best images in detecting these tumors in lung that are especially accepted among doctors. However, location and random shape of tumors, and the poor quality of CT scans images are one the biggest challenges for physicians in identifying these tumors. Therefore, deep learning algorithms have been highly regarded by researchers. This paper presents a new method for identifying tumors and pulmonary nodules in CT scans images based on convolution neural network algorithm with which tumor is accurately identified. The active counter algorithm will show the detected tumor. The proposed method is qualitatively measured by the sensitivity assessment criteria and dice similarity criteria. The obtained results with 98.33% accuracy 99.25% validity and 98.18% dice similarity criterion show the superiority of the proposed method.

Pouch Transfer for Single Coronary Artery With Nodal Artery Variant in Arterial Switch Operation.

The arterial switch operation with single coronary artery variance is an independent risk factor for increased operative mortality. There are reports of technical modifications, such as the double-barreled sinus pouch configuration, to improve geometric reimplantation of the single coronary into the neoaortic sinus. We describe the novel application of this technique for transferring a single coronary artery with a separate nodal artery emanating from the opposite sinus during an arterial switch operation.

Intraoperative Sapien S3 Valve Placement in 2 Patients With Multi-Valvar Disease Operation: A Unique Hybrid Procedure.

There has been a paradigm shift in the management of patients with congenital heart disease with a move away from conventional surgical treatment in favor of a percutaneous catheter-based approach across the spectrum of valvular heart diseases. The Sapien S3 valve implantation in the pulmonary position has been previously reported using a conventional transcatheter approach in patients with pulmonary insufficiency due to an enlarged right ventricular outflow tract. In this report, we present 2 unique cases of intraoperative hybrid implantation of Sapien S3 valves in patients with complex pulmonic and tricuspid valvular disease.