Superior lentiviral vectors designed for BSL-0 environment abolish vector mobilization.

Lentiviral vector mobilization following HIV-1 infection of vector-transduced cells poses biosafety risks to vector-treated patients and their communities. The self-inactivating (SIN) vector design has reduced, however, not abolished mobilization of integrated vector genomes. Furthermore, an earlier study demonstrated the ability of the major product of reverse transcription, a circular SIN HIV-1 vector comprising a single- long terminal repeat (LTR) to support production of high vector titers. Here, we demonstrate that configuring the internal vector expression cassette in opposite orientation to the LTRs abolishes mobilization of SIN vectors. This additional SIN mechanism is in part premised on induction of host PKR response to double-stranded RNAs comprised of mRNAs transcribed from cryptic transcription initiation sites around 3'SIN-LTR's and the vector internal promoter. As anticipated, PKR response following transfection of opposite orientation vectors, negatively affects their titers. Importantly, shRNA-mediated knockdown of PKR rendered titers of SIN HIV-1 vectors comprising opposite orientation expression cassettes comparable to titers of conventional SIN vectors. High-titer vectors carrying an expression cassette in opposite orientation to the LTRs efficiently delivered and maintained high levels of transgene expression in mouse livers. This study establishes opposite orientation expression cassettes as an additional PKR-dependent SIN mechanism that abolishes vector mobilization from integrated and episomal SIN lentiviral vectors

Expression of human adenosine deaminase in nonhuman primates after retrovirus-mediated gene transfer.

Primate bone marrow cells were infected with a retroviral vector carrying the genes for human adenosine deaminase (h-ADA) and bacterial neomycin resistance (neor). The infected cells were infused back into the lethally irradiated donor animals. Several monkeys fully reconstituted and were shown to express the h-ADA and neor genes at low levels in their recirculating hematopoietic cells for short periods of time

Creating New β-Globin-Expressing Lentiviral Vectors by High-Resolution Mapping of Locus Control Region Enhancer Sequences.

Hematopoietic stem cell gene therapy is a promising approach for treating disorders of the hematopoietic system. Identifying combinations of cis-regulatory elements that do not impede packaging or transduction efficiency when included in lentiviral vectors has proven challenging. In this study, we deploy LV-MPRA (lentiviral vector-based, massively parallel reporter assay), an approach that simultaneously analyzes thousands of synthetic DNA fragments in parallel to identify sequence-intrinsic and lineage-specific enhancer function at near-base-pair resolution. We demonstrate the power of LV-MPRA in elucidating the boundaries of previously unknown intrinsic enhancer sequences of the human β-globin locus control region. Our approach facilitated the rapid assembly of novel therapeutic βAS3-globin lentiviral vectors harboring strong lineage-specific recombinant control elements capable of correcting a mouse model of sickle cell disease. LV-MPRA can be used to map any genomic locus for enhancer activity and facilitates the rapid development of therapeutic vectors for treating disorders of the hematopoietic system or other specific tissues and cell types

Relativistic nuclear model with point-couplings constrained by QCD and chiral symmetry

We derive a microscopic relativistic point-coupling model of nuclear many-body dynamics constrained by in-medium QCD sum rules and chiral symmetry. The effective Lagrangian is characterized by density dependent coupling strengths, determined by chiral one- and two-pion exchange and by QCD sum rule constraints for the large isoscalar nucleon self-energies that arise through changes of the quark condensate and the quark density at finite baryon density. This approach is tested in the analysis of the equations of state for symmetric and asymmetric nuclear matter, and of bulk and single-nucleon properties of finite nuclei. In comparison with purely phenomenological mean-field approaches, the built-in QCD constraints and the explicit treatment of pion exchange restrict the freedom in adjusting parameters and functional forms of density dependent couplings. It is shown that chiral (two-pion exchange) fluctuations play a prominent role for nuclear binding and saturation, whereas strong scalar and vector fields of about equal magnitude and opposite sign, induced by changes of the QCD vacuum in the presence of baryonic matter, generate the large effective spin-orbit potential in finite nuclei.Comment: 46 pages, 12 figures, uses elsart.cls, revised version, to appear in Nucl.Phys. A735 (2004) 449-48

Hormander class of pseudo-differential operators on compact Lie groups and global hypoellipticity

In this paper we give several global characterisations of the Hormander class of pseudo-differential operators on compact Lie groups. The result is applied to give criteria for the ellipticity and the global hypoellipticity of pseudo-differential operators in terms of their matrix-valued full symbols. Several examples of the first and second order globally hypoelliptic differential operators are given. Where the global hypoelliptiticy fails, one can construct explicit examples based on the analysis of the global symbols.Comment: 20 page

Essential self-adjointness, generalized eigenforms, and spectra for the ∂ˉ\bar\partial-Neumann problem on GG-manifolds

Let $M$ be a strongly pseudoconvex complex manifold which is also the total space of a principal $G$-bundle with $G$ a Lie group and compact orbit space $\bar M/G$. Here we investigate the $\bar\partial$-Neumann Laplacian on $M$. We show that it is essentially self-adjoint on its restriction to compactly supported smooth forms. Moreover we relate its spectrum to the existence of generalized eigenforms: an energy belongs to $\sigma(\square)$ if there is a subexponentially bounded generalized eigenform for this energy. Vice versa, there is an expansion in terms of these well-behaved eigenforms so that, spectrally, almost every energy comes with such a generalized eigenform.Comment: 25 page

Synthetic Lethality of Chk1 Inhibition Combined with p53 and/or p21 Loss During a DNA Damage Response in Normal and Tumor Cells

Cell cycle checkpoints ensure genome integrity and are frequently compromised in human cancers. A therapeutic strategy being explored takes advantage of checkpoint defects in p53-deficient tumors in order to sensitize them to DNA-damaging agents by eliminating Chk1-mediated checkpoint responses. Using mouse models, we demonstrated that p21 is a key determinant of how cells respond to the combination of DNA damage and Chk1 inhibition (combination therapy) in normal cells as well as in tumors. Loss of p21 sensitized normal cells to the combination therapy much more than did p53 loss and the enhanced lethality was partially blocked by CDK inhibition. In addition, basal pools of p21 (p53 independent) provided p53 null cells with protection from the combination therapy. Our results uncover a novel p53-independent function for p21 in protecting cells from the lethal effects of DNA damage followed by Chk1 inhibition. As p21 levels are low in a significant fraction of colorectal tumors, they are predicted to be particularly sensitive to the combination therapy. Results reported in this study support this prediction

The axial ratio of hcp iron at the conditions of the Earth's inner core

We present ab initio calculations of the high-temperature axial c/a ratio of hexagonal-close-packed (hcp) iron at Earth's core pressures, in order to help interpret the observed seismic anisotropy of the inner core. The calculations are based on density functional theory, which is known to predict the properties of high-pressure iron with good accuracy. The temperature dependence of c/a is determined by minimising the Helmholtz free energy at fixed volume and temperature, with thermal contributions due to lattice vibrations calculated using harmonic theory. Anharmonic corrections to the harmonic predictions are estimated from calculations of the thermal average stress obtained from ab initio molecular dynamics simulations of hcp iron at the conditions of the inner core. We find a very gradual increase of axial ratio with temperature. This increase is much smaller than found in earlier calculations, but is in reasonable agreement with recent high-pressure, high-temperature diffraction measurements. This result casts doubt on an earlier interpretation of the seismic anisotropy of the inner core

On the well-posedness of the stochastic Allen-Cahn equation in two dimensions

White noise-driven nonlinear stochastic partial differential equations (SPDEs) of parabolic type are frequently used to model physical and biological systems in space dimensions d = 1,2,3. Whereas existence and uniqueness of weak solutions to these equations are well established in one dimension, the situation is different for d \geq 2. Despite their popularity in the applied sciences, higher dimensional versions of these SPDE models are generally assumed to be ill-posed by the mathematics community. We study this discrepancy on the specific example of the two dimensional Allen-Cahn equation driven by additive white noise. Since it is unclear how to define the notion of a weak solution to this equation, we regularize the noise and introduce a family of approximations. Based on heuristic arguments and numerical experiments, we conjecture that these approximations exhibit divergent behavior in the continuum limit. The results strongly suggest that a series of published numerical studies are problematic: shrinking the mesh size in these simulations does not lead to the recovery of a physically meaningful limit.Comment: 21 pages, 4 figures; accepted by Journal of Computational Physics (Dec 2011

Temperature dependence in interatomic potentials and an improved potential for Ti

The process of deriving an interatomic potentials represents an attempt to integrate out the electronic degrees of freedom from the full quantum description of a condensed matter system. In practice it is the derivatives of the interatomic potentials which are used in molecular dynamics, as a model for the forces on a system. These forces should be the derivative of the free energy of the electronic system, which includes contributions from the entropy of the electronic states. This free energy is weakly temperature dependent, and although this can be safely neglected in many cases there are some systems where the electronic entropy plays a significant role. Here a method is proposed to incorporate electronic entropy in the Sommerfeld approximation into empirical potentials. The method is applied as a correction to an existing potential for titanium. Thermal properties of the new model are calculated, and a simple method for fixing the melting point and solid-solid phase transition temperature for existing models fitted to zero temperature data is presented.Comment: CCP 201