Quantization of the electromagnetic field outside static black holes and its application to low-energy phenomena

We discuss the Gupta-Bleuler quantization of the free electromagnetic field outside static black holes in the Boulware vacuum. We use a gauge which reduces to the Feynman gauge in Minkowski spacetime. We also discuss its relation with gauges used previously. Then we apply the low-energy sector of this field theory to investigate some low-energy phenomena. First, we discuss the response rate of a static charge outside the Schwarzschild black hole in four dimensions. Next, motivated by string physics, we compute the absorption cross sections of low-energy plane waves for the Schwarzschild and extreme Reissner-Nordstr\"om black holes in arbitrary dimensions higher than three.Comment: 26 pages (revtex), no figures, misprints in some conditions correcte

Managing the sequence-specificity of antisense oligonucleotides in drug discovery

All drugs perturb the expression of many genes in the cells that are exposed to them. These gene expression changes can be divided into effects resulting from engaging the intended target and effects resulting from engaging unintended targets. For antisense oligonucleotides, developments in bioinformatics algorithms, and the quality of sequence databases, allow oligonucleotide sequences to be analyzed computationally, in terms of the predictability of their interactions with intended and unintended RNA targets. Applying these tools enables selection of sequence-specific oligonucleotides where no- or only few unintended RNA targets are expected. To evaluate oligonucleotide sequence-specificity experimentally, we recommend a transcriptomics protocol where two or more oligonucleotides targeting the same RNA molecule, but with entirely different sequences, are evaluated together. This helps to clarify which changes in cellular RNA levels result from downstream processes of engaging the intended target, and which are likely to be related to engaging unintended targets. As required for all classes of drugs, the toxic potential of oligonucleotides must be evaluated in cell- and animal models before clinical testing. Since potential adverse effects related to unintended targeting are sequence-dependent and therefore species-specific, in vitro toxicology assays in human cells are especially relevant in oligonucleotide drug discovery

Quantum Mechanical Studies of DNA and LNA

Quantum mechanical (QM) methodology has been employed to study the structure activity relations of DNA and locked nucleic acid (LNA). The QM calculations provide the basis for construction of molecular structure and electrostatic surface potentials from molecular orbitals. The topologies of the electrostatic potentials were compared among model oligonucleotides, and it was observed that small structural modifications induce global changes in the molecular structure and surface potentials. Since ligand structure and electrostatic potential complementarity with a receptor is a determinant for the bonding pattern between molecules, minor chemical modifications may have profound changes in the interaction profiles of oligonucleotides, possibly leading to changes in pharmacological properties. The QM modeling data can be used to understand earlier observations of antisense oligonucleotide properties, that is, the observation that small structural changes in oligonucleotide composition may lead to dramatic shifts in phenotypes. These observations should be taken into account in future oligonucleotide drug discovery, and by focusing more on non RNA target interactions it should be possible to utilize the exhibited property diversity of oligonucleotides to produce improved antisense drugs

Electronic Structures of LNA Phosphorothioate Oligonucleotides

Important oligonucleotides in anti-sense research have been investigated in silico and experimentally. This involves quantum mechanical (QM) calculations and chromatography experiments on locked nucleic acid (LNA) phosphorothioate (PS) oligonucleotides. iso-potential electrostatic surfaces are essential in this study and have been calculated from the wave functions derived from the QM calculations that provide binding information and other properties of these molecules. The QM calculations give details of the electronic structures in terms of e.g., energy and bonding, which make them distinguish or differentiate between the individual PS diastereoisomers determined by the position of sulfur atoms. Rules are derived from the electronic calculations of these molecules and include the effects of the phosphorothioate chirality and formation of electrostatic potential surfaces. Physical and electrochemical descriptors of the PS oligonucleotides are compared to the experiments in which chiral states on these molecules can be distinguished. The calculations demonstrate that electronic structure, electrostatic potential, and topology are highly sensitive to single PS configuration changes and can give a lead to understanding the activity of the molecules. Keywords: LNA phosphorothioate, DNA/LNA oligonucleotide, diastereoisomers, Hartree-Fock calculations, iso-potential surface, anion chromatogram

Locked nucleic acid (LNA) mediated improvements in siRNA stability and functionality

Therapeutic application of the recently discovered small interfering RNA (siRNA) gene silencing phenomenon will be dependent on improvements in molecule bio-stability, specificity and delivery. To address these issues, we have systematically modified siRNA with the synthetic RNA-like high affinity nucleotide analogue, Locked Nucleic Acid (LNA). Here, we show that incorporation of LNA substantially enhances serum half-life of siRNA's, which is a key requirement for therapeutic use. Moreover, we provide evidence that LNA is compatible with the intracellular siRNA machinery and can be used to reduce undesired, sequence-related off-target effects. LNA-modified siRNAs targeting the emerging disease SARS, show improved efficiency over unmodified siRNA on certain RNA motifs. The results from this study emphasize LNA's promise in converting siRNA from a functional genomics technology to a therapeutic platform

Erythema nodosum and the risk of tuberculosis in a high incidence setting

Objective: This study estimates the erythema nodosum (EN) incidence in a tuberculosis (TB) endemic setting and evaluates the likelihood of a subsequent TB diagnosis among individuals with Mycobacterium tuberculosis infection (MTI) with or without EN. Design: We estimated EN incidence rates (IRs) in East Greenland in 2010–2011 and conducted a cohort study following all individuals who tested positive for MTI from 1 January 2010 until 31 December 2012. A personal identifier allowed individual follow-up in the mandatory TB register. MTI was defined by a positive interferon-gamma release assay. TB incidence rate ratios (IRRs) among participants with or without EN were estimated with the Cox proportional hazard model. Results: We identified 38 EN cases corresponding to an IR of 500/100,000 inhabitants/year. All cases were among individuals with MTI. The EN IR was 11.79 (95% CI 5.73–24.27) times higher for BCG-unvaccinated compared with BCG-vaccinated individuals. The TB IRR was 25 (95% CI 11–60) within 1 month of EN compared to individuals without EN. Conclusion: This study documents a high EN incidence in a TB endemic region. EN occurred only in individuals with MTI, and predominantly among BCG-unvaccinated individuals. EN was significantly associated with a TB diagnosis within 1 month of diagnosis

Short locked nucleic acid antisense oligonucleotides potently reduce apolipoprotein B mRNA and serum cholesterol in mice and non-human primates

The potency and specificity of locked nucleic acid (LNA) antisense oligonucleotides was investigated as a function of length and affinity. The oligonucleotides were designed to target apolipoprotein B (apoB) and were investigated both in vitro and in vivo. The high affinity of LNA enabled the design of short antisense oligonucleotides (12- to 13-mers) that possessed high affinity and increased potency both in vitro and in vivo compared to longer oligonucleotides. The short LNA oligonucleotides were more target specific, and they exhibited the same biodistribution and tissue half-life as longer oligonucleotides. Pharmacology studies in both mice and non-human primates were conducted with a 13-mer LNA oligonucleotide against apoB, and the data showed that repeated dosing of the 13-mer at 1–2 mg/kg/week was sufficient to provide a significant and long lasting lowering of non-high-density lipoprotein (non-HDL) cholesterol without increasing serum liver toxicity markers. The data presented here show that oligonucleotide length as a parameter needs to be considered in the design of antisense oligonucleotide and that potent short oligonucleotides with sufficient target affinity can be generated using the LNA chemistry. Conclusively, we present a 13-mer LNA oligonucleotide with therapeutic potential that produce beneficial cholesterol lowering effect in non-human primates