Engineered packaging cell line for the enhanced production of baboon-enveloped retroviral vectors.

The baboon endogenous retrovirus (BaEV) glycoprotein is superior to the commonly used vesicular stomatitis virus glycoprotein (VSVg) for retroviral gene transfer into resting hematopoietic stem cells and lymphocyte populations. The derivative BaEVRLess (lacking the R domain) produces higher viral titers compared with wild-type BaEV, but vector production is impaired by syncytia formation and cell death of the HEK293T cells due to the high fusogenic activity of the glycoprotein. This lowers viral titers, leads to increased batch-to-batch variability, and impedes the establishment of stable packaging cell lines essential for the economical production of viral supernatants.

Gene editing without ex vivo culture evades genotoxicity in human hematopoietic stem cells.

Gene editing the BCL11A erythroid enhancer is a validated approach to fetal hemoglobin (HbF) induction for β-hemoglobinopathy therapy, though heterogeneity in edit allele distribution and HbF response may impact its safety and efficacy. Here, we compare combined CRISPR-Cas9 editing of the BCL11A +58 and +55 enhancers with leading gene modification approaches under clinical investigation. Dual targeting of the BCL11A +58 and +55 enhancers with 3xNLS-SpCas9 and two single guide RNAs (sgRNAs) resulted in superior HbF induction, including in sickle cell disease (SCD) patient xenografts, attributable to simultaneous disruption of core half E-box/GATA motifs at both enhancers.

UM171 enhances fitness and engraftment of gene-modified hematopoietic stem cells from patients with sickle cell disease.

Hematopoietic stem cell (HSC) transplantation with lentiviral vector (LVV)-transduced autologous cells has proven an effective therapeutic strategy for sickle cell disease (SCD). However, ex vivo culture or proliferative stress associated with in vivo reconstitution may amplify any underlying genetic risk of leukemia. We aimed to minimize culture-induced stress and reduce genomic damage during ex vivo culture and enhance stem cell fitness and reconstitution of SCD CD34+ cells transduced with BCL11A shmiR-encoding LVV.

Scalable assessment of genome editing off-targets associated with genetic variants.

Genome editing with RNA-guided DNA binding factors carries risk of off-target editing at homologous sequences. Genetic variants may introduce sequence changes that increase homology to a genome editing target, thereby increasing risk of off-target editing. Conventional methods to verify candidate off-targets rely on access to cells with genomic DNA carrying these sequences.

Engineering an inducible leukemia-associated fusion protein enables large-scale ex vivo production of functional human phagocytes.

Ex vivo expansion of human CD34+ hematopoietic stem and progenitor cells remains a challenge due to rapid differentiation after detachment from the bone marrow niche. In this study, we assessed the capacity of an inducible fusion protein to enable sustained ex vivo proliferation of hematopoietic precursors and their capacity to differentiate into functional phagocytes. We fused the coding sequences of an FK506-Binding Protein 12 (FKBP12)-derived destabilization domain (DD) to the myeloid/lymphoid lineage leukemia/eleven nineteen leukemia (MLL-ENL) fusion gene to generate the fusion protein DD-MLL-ENL and retrovirally expressed the protein switch in human CD34+ progenitors.

A novel high-titer, bifunctional lentiviral vector for autologous hematopoietic stem cell gene therapy of sickle cell disease.

A major limitation of gene therapy for sickle cell disease (SCD) is the availability and access to a potentially curative one-time treatment, due to high treatment costs. We have developed a high-titer bifunctional lentiviral vector (LVV) in a vector backbone that has reduced size, high vector yields, and efficient gene transfer to human CD34 hematopoietic stem and progenitor cells (HSPCs). This LVV contains locus control region cores expressing an anti-sickling β-globin gene and two microRNA-adapted short hairpin RNA simultaneously targeting and transcripts to maximally induce fetal hemoglobin (HbF) expression.

Development of an explainable AI system using routine clinical parameters for rapid differentiation of inflammatory conditions.

Introduction: Inflammatory conditions in patients have various causes and require different treatments. Bacterial infections are treated with antibiotics, while these medications are ineffective against viral infections. Autoimmune diseases and graft-versus-host disease (GVHD) after allogeneic stem cell transplantation, require immunosuppressive therapies such as glucocorticoids, which may be contraindicated in other inflammatory states.

Genetic reversal of the globin switch concurrently modulates both fetal and sickle hemoglobin and reduces red cell sickling.

We previously reported initial clinical results of post-transcriptional gene silencing of BCL11A expression (NCT03282656) reversing the fetal to adult hemoglobin switch. A goal of this approach is to increase fetal hemoglobin (HbF) expression while coordinately reducing sickle hemoglobin (HbS) expression. The resulting combinatorial effect should prove effective in inhibiting HbS polymerization at lower physiologic oxygen values thereby mitigating disease complications.

Prediction of Clinical Outcomes with Explainable Artificial Intelligence in Patients with Chronic Lymphocytic Leukemia.

Background: The International Prognostic Index (IPI) is applied to predict the outcome of chronic lymphocytic leukemia (CLL) with five prognostic factors, including genetic analysis. We investigated whether multiparameter flow cytometry (MPFC) data of CLL samples could predict the outcome by methods of explainable artificial intelligence (XAI). Further, XAI should explain the results based on distinctive cell populations in MPFC dot plots.

A Bioinformatics View on Acute Myeloid Leukemia Surface Molecules by Combined Bayesian and ABC Analysis.

"Big omics data" provoke the challenge of extracting meaningful information with clinical benefit. Here, we propose a two-step approach, an initial unsupervised inspection of the structure of the high dimensional data followed by supervised analysis of gene expression levels, to reconstruct the surface patterns on different subtypes of acute myeloid leukemia (AML). First, Bayesian methodology was used, focusing on surface molecules encoded by cluster of differentiation (CD) genes to assess whether AML is a homogeneous group or segregates into clusters.

The Low Expression of Fc-Gamma Receptor III (CD16) and High Expression of Fc-Gamma Receptor I (CD64) on Neutrophil Granulocytes Mark Severe COVID-19 Pneumonia.

Hyperinflammation through neutrophil granulocytes contributes to disease severity in COVID-19 pneumonia and promotes acute lung failure. Understanding the mechanisms of the dysregulations within the myeloid cell compartment may help to improve therapies for severe COVID-19 infection. Here, we investigated the immunopathological characteristics of circulating neutrophil granulocytes and monocytes in 16 patients with COVID-19 pneumonia by multiparameter flow cytometry in comparison to 9 patients with pulmonary infiltrates but without COVID-19.

Flow cytometry datasets consisting of peripheral blood and bone marrow samples for the evaluation of explainable artificial intelligence methods.

Three different Flow Cytometry datasets consisting of diagnostic samples of either peripheral blood (pB) or bone marrow (BM) from patients without any sign of bone marrow disease at two different health care centers are provided. In Flow Cytometry, each cell rapidly passes through a laser beam one by one, and two light scatter, and eight surface parameters of more than 100.000 cells are measured per sample of each patient.

Topological phonon transport in an optomechanical system.

Light is a powerful tool for controlling mechanical motion, as shown by numerous applications in the field of cavity optomechanics. Recently, small scale optomechanical circuits, connecting a few optical and mechanical modes, have been demonstrated in an ongoing push towards multi-mode on-chip optomechanical systems. An ambitious goal driving this trend is to produce topologically protected phonon transport.

Economic Impact of COVID-19 Pandemic on Dental Practices in Germany: A Cross-Sectional Survey.

An observational cross-sectional survey was planned and carried out to evaluate the economic impact of the SARS-CoV-2/COVID-19 pandemic on dental practices in Germany. An online-questionnaire was developed and previously calibrated by a group consisting of experts from dentists, lawyers, and business economists (n = 21; Intra-Class-Coefficient > 0.8).

Development of a double shmiR lentivirus effectively targeting both BCL11A and ZNF410 for enhanced induction of fetal hemoglobin to treat β-hemoglobinopathies.

A promising treatment for β-hemoglobinopathies is the de-repression of γ-globin expression leading to increased fetal hemoglobin (HbF) by targeting BCL11A. Here, we aim to improve a lentivirus vector (LV) containing a single BCL11A shmiR (SS) to further increase γ-globin induction. We engineered a novel LV to express two shmiRs simultaneously targeting BCL11A and the γ-globin repressor ZNF410.

Monitoring multiple myeloma in the peripheral blood based on cell-free DNA and circulating plasma cells.

With the advent of novel, highly effective therapies for multiple myeloma (MM), classical serologic monitoring appears insufficient for response assessment and prediction of relapse. Moreover, serologic studies in MM are hampered by interference of therapeutic antibodies. The detection of malignant plasma cell clones by next generation sequencing (NGS) or multiparameter flow cytometry (MFC) circumvents these difficulties and can be performed in the peripheral blood (pB) by targeting circulating cell-free DNA (cfDNA) or circulating plasma cells (CPCs), thus also avoiding an invasive sampling procedure.

Foliar ẟN patterns in legumes and non-N fixers across a climate gradient, Hawai'i Island, USA.

Recent studies from the Hawaiian Islands showed that pedogenic thresholds demarcate domains in which rock-derived nutrient dynamics remain similar across wide variations in rainfall. These thresholds appear related to certain aspects of N cycling, but the degree to which they correspond to patterns of biological N fixation (BNF)-the dominant input of N into less-managed ecosystems-remains unclear. We measured aboveground plant biomass, foliar nutrient concentrations, and foliar δN along a climate gradient on ~ 150,000-year-old basaltic substrate to characterize foliar N sources and spatially relate them to soil nutrients.

Deletion of murine leads to de-repression of via decreased KAISO levels and accelerates a malignancy phenotype in a murine model of lymphoma.

RHOH/TFF, a member of the RAS GTPase super family, has important functions in lymphopoiesis and proximal T cell receptor signalling and has been implicated in a variety of leukaemias and lymphomas. RHOH was initially identified as a translocation partner with BCL-6 in non-Hodgkin lymphoma (NHL), and aberrant somatic hypermutation (SHM) in the 5' untranslated region of the RHOH gene has also been detected in Diffuse Large B-Cell Lymphoma (DLBCL). Recent data suggest a correlation between RhoH expression and disease progression in Acute Myeloid Leukaemia (AML).

Stacked conservation practices reduce nitrogen loss: A paired watershed study.

Combinations of best management practices (BMPs) are needed to achieve nutrient reduction goals in the Mississippi/Atchafalaya River Basin (MARB), but field results are crucial to encourage stacked adoption of BMPs. A paired catchment-scale study (2015-18) was done to assess the impact of (i) BMPs, (ii) precipitation patterns, and (iii) seasonality on nitrogen (N) export. Flow-weighted samples were collected and analyzed for total ammonia nitrogen (TAN), nitrate (NO-N), and total nitrogen (TN).

Superior Overall Survival in Patients with Colorectal Cancer, Regular Aspirin Use, and Combined Wild-Type PIK3CA and KRAS-Mutated Tumors.

The impact of aspirin use after the diagnosis of colorectal cancer is unknown. Among others, (phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha) mutational status was proposed as a molecular biomarker for the response to adjuvant aspirin therapy. However, prognostic data on aspirin use after a colorectal cancer diagnosis in relation to mutational status is limited.