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.

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.

1-Norm waveform analysis for MR elastography-based quantification of inhomogeneity: Effects of the freeze-thaw cycle and Alzheimer's disease.

Background: Despite its success in the mechanical characterization of biological tissues, magnetic resonance elastography (MRE) uses ill-posed wave inversions to estimate tissue stiffness. 1-Norm has been recently introduced as a mathematical measure for the scattering of mechanical waves due to inhomogeneities based on an analysis of the delineated contours of wave displacement.

Purpose: To investigate 1-Norm as an MRE-based quantitative biomarker of mechanical inhomogeneities arising from microscopic structural tissue alterations caused by the freeze-thaw cycle (FTC) or Alzheimer's disease (AD).

Quantifying uniaxial prestress and waveguide effects on dynamic elastography estimates for a cylindrical rod.

Dynamic elastography attempts to reconstruct quantitative maps of the viscoelastic properties of materials by noninvasively measuring mechanical wave motion in them. The target motion is typically transversely-polarized relative to the wave propagation direction, such as bulk shear wave motion. In addition to neglecting waveguide effects caused by small lengths in one dimension or more, many reconstruction strategies also ignore nonzero, non-isotropic static preloads.

Decoupling Uniaxial Tensile Prestress and Waveguide Effects From Estimates of the Complex Shear Modulus in a Cylindrical Structure Using Transverse-Polarized Dynamic Elastography.

Dynamic elastography, whether based on magnetic resonance, ultrasound, or optical modalities, attempts to reconstruct quantitative maps of the viscoelastic properties of biological tissue, properties altered by disease and injury, by noninvasively measuring mechanical wave motion in the tissue. Most reconstruction strategies that have been developed neglect boundary conditions, including quasi-static tensile or compressive loading resulting in a nonzero prestress. Significant prestress is inherent to the functional role of some biological tissues currently being studied using elastography, such as skeletal and cardiac muscle, arterial walls, and the cornea.

Airway basal cells show a dedifferentiated KRT17Phenotype and promote fibrosis in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a fatal disease with limited treatment options. In this study, we focus on the properties of airway basal cells (ABC) obtained from patients with IPF (IPF-ABC). Single cell RNA sequencing (scRNAseq) of bronchial brushes revealed extensive reprogramming of IPF-ABC towards a KRT17 PTEN dedifferentiated cell type.

Tabletop MR elastography for investigating effects of the freeze-thaw cycle on the mechanical properties of biological tissues.

Purpose: We aimed at characterizing the effects of the freeze-thaw cycle (FTC) on ex vivo specimens of porcine muscle, liver, kidney, and brain using tabletop magnetic resonance elastography (MRE) combined with rheological modeling. While frozen tissue banks potentially facilitate access to large amounts of well-preserved biospecimens, the impact of the FTC on their viscoelastic properties remains elusive.

Methods: In this proof-of-concept study, fresh specimens from porcine lumbar muscle (n = 6), liver (n = 6), kidney (n = 6), and brain (n = 6) were examined before and after the FTC using 0.

Improved alpharetrovirus-based Gag.MS2 particles for efficient and transient delivery of CRISPR-Cas9 into target cells.

DNA-modifying technologies, such as the CRISPR-Cas9 system, are promising tools in the field of gene and cell therapies. However, high and prolonged expression of DNA-modifying enzymes may cause cytotoxic and genotoxic side effects and is therefore unwanted in therapeutic approaches. Consequently, development of new and potent short-term delivery methods is of utmost importance.

Optical coherence elastography for assessing the influence of intraocular pressure on elastic wave dispersion in the cornea.

The cornea is a highly specialized organ that relies on its mechanical stiffness to maintain its aspheric geometry and refractive power, and corneal diseases such as keratoconus have been linked to abnormal tissue stiffness and biomechanics. Dynamic optical coherence elastography (OCE) is a clinically promising non-contact and non-destructive imaging technique that can provide measurements of corneal tissue stiffness directly in vivo. The method relies on the concepts of elastography where shear waves are generated and imaged within a tissue to obtain mechanical properties such as tissue stiffness.

Investigating the heterogeneity of viscoelastic properties in prostate cancer using MR elastography at 9.4T in fresh prostatectomy specimens.

Purpose: To quantify the heterogeneity of viscoelastic tissue properties in prostatectomy specimens from men with prostate cancer (PC) using MR elastography (MRE) with histopathology as reference.

Methods: Twelve fresh prostatectomy specimens were examined in a preclinical 9.4T MRI scanner.

A Multiplex CRISPR-Screen Identifies PLA2G4A as Prognostic Marker and Druggable Target for HOXA9 and MEIS1 Dependent AML.

and are frequently upregulated in acute myeloid leukemia (AML), including those with MLL-rearrangement. Because of their pivotal role in hemostasis, HOXA9 and MEIS1 appear non-druggable. We, thus, interrogated gene expression data of pre-leukemic (overexpressing ) and leukemogenic (overexpressing and ; H9M) murine cell lines to identify cancer vulnerabilities.

Concurrent OCT and OCT angiography of retinal neurovascular degeneration in the 5XFAD Alzheimer's disease mice.

: As one part of the central nervous system, the retina manifests neurovascular defects in Alzheimer's disease (AD). Quantitative imaging of retinal neurovascular abnormalities may promise a new method for early diagnosis and treatment assessment of AD. Previous imaging studies of transgenic AD mouse models have been limited to the central part of the retina.

Efficient Genetic Safety Switches for Future Application of iPSC-Derived Cell Transplants.

Induced pluripotent stem cell (iPSC)-derived cell products hold great promise as a potential cell source in personalized medicine. As concerns about the potential risk of graft-related severe adverse events, such as tumor formation from residual pluripotent cells, currently restrict their applicability, we established an optimized tool for therapeutic intervention that allows drug-controlled, specific and selective ablation of either iPSCs or the whole graft through genetic safety switches. To identify the best working system, different tools for genetic iPSC modification, promoters to express safety switches and different safety switches were combined.

Measuring viscoelastic parameters in Magnetic Resonance Elastography: a comparison at high and low magnetic field intensity.

Magnetic Resonance Elastography (MRE) is a non-invasive imaging technique which involves motion-encoding MRI for the estimation of the shear viscoelastic properties of soft tissues through the study of shear wave propagation. The technique has been found informative for disease diagnosis, as well as for monitoring of the effects of therapies. The development of MRE and its validation have been supported by the use of tissue-mimicking phantoms.

Spatial heterogeneity of hepatic fibrosis in primary sclerosing cholangitis vs. viral hepatitis assessed by MR elastography.

Spatial heterogeneity of hepatic fibrosis in primary sclerosing cholangitis (PSC) in comparison to viral hepatitis was assessed as a potential new biomarker using MR elastography (MRE). In this proof-of-concept study, we hypothesized a rather increased heterogeneity in PSC and a rather homogeneous distribution in viral hepatitis. Forty-six consecutive subjects (PSC: n = 20, viral hepatitis: n = 26) were prospectively enrolled between July 2014 and April 2017.

Axially- and torsionally-polarized radially converging shear wave MRE in an anisotropic phantom made via Embedded Direct Ink Writing.

Magnetic Resonance Elastography (MRE) is a non-invasive imaging method to quantitatively map the shear viscoelastic properties of soft tissues. In this study, Embedded Direct Ink Writing is used to fabricate a muscle mimicking anisotropic phantom that may serve as a standard for imaging studies of anisotropic materials. The technique allowed us to obtain a long shelf life silicone-based phantom expressing transverse isotropic mechanical properties.

Longitudinal study of sub-regional cerebral viscoelastic properties of 5XFAD Alzheimer's disease mice using multifrequency MR elastography.

Purpose: To study sub-regional, longitudinal changes occurring inside brains of 5XFAD mice, an Alzheimer's disease (AD) model, based on viscoelastic parameters derived using MR elastography and their spatial variation.

Methods: Female 5XFAD and non-transgenic B6SJLF1/J mice as controls (n = 9 for both groups) were used for the study. Scans were performed inside a 9.

Competitive sgRNA Screen Identifies p38 MAPK as a Druggable Target to Improve HSPC Engraftment.

Previous gene therapy trials for X-linked chronic granulomatous disease (X-CGD) lacked long-term engraftment of corrected hematopoietic stem and progenitor cells (HSPCs). Chronic inflammation and high levels of interleukin-1 beta (IL1B) might have caused aberrant cell cycling in X-CGD HSPCs with a concurrent loss of their long-term repopulating potential. Thus, we performed a targeted CRISPR-Cas9-based sgRNA screen to identify candidate genes that counteract the decreased repopulating capacity of HSPCs during gene therapy.

Hematopoietic stem-cell senescence and myocardial repair - Coronary artery disease genotype/phenotype analysis of post-MI myocardial regeneration response induced by CABG/CD133+ bone marrow hematopoietic stem cell treatment in RCT PERFECT Phase 3.

Background: Bone marrow stem cell clonal dysfunction by somatic mutation is suspected to affect post-infarction myocardial regeneration after coronary bypass surgery (CABG).

Methods: Transcriptome and variant expression analysis was studied in the phase 3 PERFECT trial post myocardial infarction CABG and CD133 bone marrow derived hematopoetic stem cells showing difference in left ventricular ejection fraction (∆LVEF) myocardial regeneration Responders (n=14; ∆LVEF +16% day 180/0) and Non-responders (n=9; ∆LVEF -1.1% day 180/0).

CXCR4/MIF axis amplifies tumor growth and epithelial-mesenchymal interaction in non-small cell lung cancer.

Overexpression of C-X-C chemokine receptor type 4 (CXCR4) has been shown in several cancers, including non-small cell lung cancer (NSCLC) and is linked to early metastasis and worse prognosis. The crosstalk between cancer cells and tumor stroma promotes the growth and metastasis and CXCR4 signaling is a key element of this crosstalk. To test the effects of CXCR4 overexpression (CXCR4-OE), we transduced the human NSCLC cell line A549 by using a lentiviral vector.

Preclinical Evaluation of a Novel Lentiviral Vector Driving Lineage-Specific BCL11A Knockdown for Sickle Cell Gene Therapy.

In this work we provide preclinical data to support initiation of a first-in-human trial for sickle cell disease (SCD) using an approach that relies on reversal of the developmental fetal-to-adult hemoglobin switch. Erythroid-specific knockdown of BCL11A via a lentiviral-encoded microRNA-adapted short hairpin RNA (shRNA) leads to reactivation of the gamma-globin gene while simultaneously reducing expression of the pathogenic adult sickle β-globin. We generated a refined lentiviral vector (LVV) BCH-BB694 that was developed to overcome poor vector titers observed in the manufacturing scale-up of the original research-grade LVV.

Prostate cancer assessment using MR elastography of fresh prostatectomy specimens at 9.4 T.

Purpose: Despite its success in the assessment of prostate cancer (PCa), in vivo multiparametric MRI has limitations such as interobserver variability and low specificity. Several MRI methods, among them MR elastography, are currently being discussed as candidates for supplementing conventional multiparametric MRI. This study aims to investigate the detection of PCa in fresh ex vivo human prostatectomy specimens using MR elastography.

Differential Transgene Silencing of Myeloid-Specific Promoters in the Safe Harbor Locus of Induced Pluripotent Stem Cell-Derived Myeloid Cells.

Targeted integration into a genomic safe harbor, such as the locus on chromosome 19, promises predictable transgene expression and reduces the risk of insertional mutagenesis in the host genome. The application of gamma-retroviral long terminal repeat (LTR)-driven vectors, which semirandomly integrate into the genome, has previously caused severe adverse events in some clinical studies due to transactivation of neighboring proto-oncogenes. Consequently, the site-specific integration of a therapeutic transgene into a genomic safe harbor locus would allow stable genetic correction with a reduced risk of insertional mutagenesis.