Supramolecular Assembly of Aminoethylene‐Lipopeptide PMO Conjugates into RNA Splice‐Switching Nanomicelles

Phosphorodiamidate morpholino oligomers (PMOs) are oligonucleotide analogs that can be used for therapeutic modulation of pre‐mRNA splicing. Similar to other classes of nucleic acid‐based therapeutics, PMOs require delivery systems for efficient transport to the intracellular target sites. Here, artificial peptides based on the oligo(ethylenamino) acid succinyl‐tetraethylenpentamine (Stp), hydrophobic modifications, and an azide group are presented, which are used for strain‐promoted azide‐alkyne cycloaddition conjugation with splice‐switching PMOs. By systematically varying the lead structure and formulation, it is determined that the type of contained fatty acid and supramolecular assembly have a critical impact on the delivery efficacy. A compound containing linolenic acid with three cis double bonds exhibits the highest splice‐switching activity and significantly increases functional protein expression in pLuc/705 reporter cells in vitro and after local administration in vivo. Structural and mechanistic studies reveal that the lipopeptide PMO conjugates form nanoparticles, which accelerate cellular uptake and that the content of unsaturated fatty acids enhances endosomal escape. In an in vitro Duchenne muscular dystrophy exon skipping model using H2K‐mdx52 dystrophic skeletal myotubes, the highly potent PMO conjugates mediate significant splice‐switching at very low nanomolar concentrations. The presented aminoethylene‐lipopeptides are thus a promising platform for the generation of PMO‐therapeutics with a favorable activity/toxicity profile.     

哈得逊隆起下石炭统基准面旋回与储集层预测

依据基准面旋回原理和方法,综合利用哈得逊地区的钻井、测井、地震以及地球化学资料,在研究区下石炭统识别出1个Ⅰ型和3个Ⅱ型层序界面,将哈得逊地区下石炭统划分为3个中期基准面旋回：SC1、SC2、SC3,对应于早石炭世海侵的3个海进-海退旋回.SC1旋回以滨岸砂坝沉积为主,而SC2、SC3旋回发育潮坪沉积体系.研究认为,SC1旋回滨岸砂岩及SC2下降半旋回的潮间带薄层砂岩为有利储集砂体,是地层-岩性型油气藏的重点勘探目标.     

Driving Mechanisms of Delamination Related Reliability Problems in Exposed Pad Packages

Exposed pad packages were introduced in the late 1980s and early 1990s because of their excellent thermal and electrical performance. Despite these advantages, the exposed pad packages experience a lot of thermo-hygro-mechanical related reliability problems during qualification and testing. Examples are die lift, which occurs predominantly after moisture sensitivity level conditions, and die-attach to leadframe delamination leading to downbond stitch breaks during temperature cycling. In this chapter, nonlinear finite element (FE) models using fracture mechanics based J-integral calculations are used to assess the reliability problems of the exposed pad package family. Using the parametric FE models any geometrical and material effects can be explored to their impact on the occurrence diepad delamination, and dielift. For instance the impact of diepad size is found to be of much less importance as the impact of die thickness is. Using the fracture mechanics approach, the starting location for the delamination from thermo-hygro-mechanical point of view is deducted. The results indicate that when diepad delamination is present, cracks are likely to grow beneath the die and dielift will occur. The interaction between dielift and other failure modes, such as lifted ball bonds, are not found to be very significant. The FE models are combined with simulation-based optimization methods to deduct design guidelines for optimal reliability of the exposed pad family.     

High-resolution HLA-DRB typing using denaturing gradient gel electrophoresis and direct sequencing

Skeletal development of transgenic mice with a type II collagen mutation was analyzed and compared with wild-type littermates. The single base substitution in Col2a1 resulted in a glycine to serine mutation within the helical domain and corresponded to one previously identified in a patient with the lethal human chondrodysplasia, hypochondrogenesis (Horton et al. [1992] Proc. Natl. Acad. Sci. U.S.A. 89:4583-4587). Skeletal staining of embryos from 14.5 through 18.5 days of gestation demonstrated a dwarf phenotype in the transgenic embryos, most notably short limb bones and vertebral column that was first detected at 15.5 days post-coitus. In addition to the reduced length, the extent of ossification was less in the transgenic mice. The architecture of the long bone growth plate was abnormal in the transgenic tissue, in particular there was no discernible proliferative zone. There were few stacks of characteristically flattened cells and the overall length of the growth plate in the mutant embryos was reduced. At the ultrastructural level, there were fewer collagen fibrils present in the transgenic mouse cartilage compared to that of wild-type littermates. Ultrastructural localization of collagen types II, IX and XI revealed a similar pattern between the transgenic and wild-type pups, suggesting that the collagen fibrils present in the matrix of littermates with both phenotypes had a similar composition. Skeletal analysis and cartilage histochemistry indicated that effect of the type II collagen mutation was to reduce the density of the collagen fibrils within the cartilage matrix which was associated with delayed bone formation and resulted in a short-limbed phenotype.     

Classification of the effectiveness of combination treatments

According to FDA regulations, a combination drug is not efficacious unless each component contributes to the claimed effects. For a univariate endpoint, this implies that the combination at specific doses must be superior to each of its components at the same doses. More demanding is the property of synergy, in which the effect of the combination must be superior to the effect expected based on those of its components. If it is equal to those effects, it is additive, and if it is inferior, it is antagonistic. We give regions in the combination dose plane where these concepts are well defined. If the effect of the combination is greater than the greatest effect achievable by any of its components it is therapeutically synergistic. A combination can be antagonistic, yet its components can still contribute to the claimed effects. If it is additive, synergistic or therapeutically synergistic, its components must contribute to the claimed effects. We relate these concepts and provide designs and sequential procedures for determining whether a combination is therapeutically synergistic, synergistic, additive, antagonistic and contributing or antagonistic and non-contributing.