Adenoviral (Ad)-mediated in vivo gene transfer and expression are limited in part by cellular immune responses to viral-encoded proteins and͞or transgene immunogenicity. In an attempt to diminish the former responses, we have previously developed and described helper-dependent (HD) Ad vectors in which the viral protein coding sequences are completely eliminated. These HD vectors have up to 37 kb insert capacity, are easily propagated in a Cre recombinasebased system, and can be produced to high concentration and purity (>99.9% helper-free vector). In this study, we compared safety and efficacy of leptin gene delivery mediated by an HD vector (HD-leptin) and a first-generation E1-deleted Ad vector (Ad-leptin) in normal lean and ob͞ob (leptindeficient) mice. In contrast to evidence of liver toxicity, inf lammation, and cellular infiltration observed with Adleptin delivery in mice, HD-leptin delivery was associated with a significant improvement in associated safety͞toxicity and resulted in efficient gene delivery, prolonged elevation of serum leptin levels, and associated weight loss. The greater safety, efficient gene delivery, and increased insert capacity of HD vectors are significant improvements over current Ad vectors and represent favorable features especially for clinical gene therapy applications. Adenoviral (Ad) vectors are currently among the most efficient gene transfer vehicles for both in vitro and in vivo delivery, but the utilization of current Ad vectors for many gene therapy applications is limited by the transient nature of transgene expression obtained by these vectors (1-7). Several factors have been shown to contribute to and modulate the duration of Ad-mediated gene expression and the immunogenicity of these vectors, including ''leaky'' viral protein expression and the transgene that is delivered (8-15). The development of Ad vectors that are deleted in all viral protein-coding sequences offers the prospect of a potentially safer, less immunogenic vector with an insert capacity of up to 37 kb (16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26). This vector is supplied in trans with the structural proteins required for packaging and rescue and is thus helper-dependent (HD) (24).Leptin has been recently identified as a potent modulator of weight and food intake. Daily delivery of recombinant leptin protein was shown to induce weight reduction, suppress appetite, and decrease blood insulin and glucose levels in ob͞ob (leptindeficient) mice (27)(28)(29). It has been shown that delivery of the leptin cDNA by first-generation Ad vectors (Ad-leptin) may substitute for daily recombinant leptin protein treatment, although the effects were transient in both lean and ob͞ob treated mice (7,30). In the present study, we delivered the leptin cDNA using the HD virus (HD-leptin), testing the hypothesis that elimination of the viral protein coding sequences would diminish the vector's cellular immunogenicity and toxicity, and hence support its longevity in vivo. Because both the viral proteins and the trans...

BackgroundNotch receptors normally play a key role in guiding a variety of cell fate decisions during development and differentiation of metazoan organisms. On the other hand, dysregulation of Notch1 signaling is associated with many different types of cancer as well as tumor angiogenesis, making Notch1 a potential therapeutic target.Principal FindingsHere we report the in vitro activities of inhibitory Notch1 monoclonal antibodies derived from cell-based and solid-phase screening of a phage display library. Two classes of antibodies were found, one directed against the EGF-repeat region that encompasses the ligand-binding domain (LBD), and the second directed against the activation switch of the receptor, the Notch negative regulatory region (NRR). The antibodies are selective for Notch1, inhibiting Jag2-dependent signaling by Notch1 but not by Notch 2 and 3 in reporter gene assays, with EC50 values as low as 5±3 nM and 0.13±0.09 nM for the LBD and NRR antibodies, respectively, and fail to recognize Notch4. While more potent, NRR antibodies are incomplete antagonists of Notch1 signaling. The antagonistic activity of LBD, but not NRR, antibodies is strongly dependent on the activating ligand. Both LBD and NRR antibodies bind to Notch1 on human tumor cell lines and inhibit the expression of sentinel Notch target genes, including HES1, HES5, and DTX1. NRR antibodies also strongly inhibit ligand-independent signaling in heterologous cells transiently expressing Notch1 receptors with diverse NRR “class I” point mutations, the most common type of mutation found in human T-cell acute lymphoblastic leukemia (T-ALL). In contrast, NRR antibodies failed to antagonize Notch1 receptors bearing rare “class II” or “class III” mutations, in which amino acid insertions generate a duplicated or constitutively sensitive metalloprotease cleavage site. Signaling in T-ALL cell lines bearing class I mutations is partially refractory to inhibitory antibodies as compared to cell-penetrating gamma-secretase inhibitors.Conclusions/SignificanceAntibodies that compete with Notch1 ligand binding or that bind to the negative regulatory region can act as potent inhibitors of Notch1 signaling. These antibodies may have clinical utility for conditions in which inhibition of signaling by wild-type Notch1 is desired, but are likely to be of limited value for treatment of T-ALLs associated with aberrant Notch1 activation.

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We have compared the efficacy of daily injection of recomgiven as mean ± standard error of the mean.) Importantly, binant leptin protein (rh-leptin) with adenovirus-mediated rh-leptin and ex vivo-expressed Ad-leptin were equivalently delivery of the murine or human leptin gene (Ad-leptin) for active in a functional cell-based assay. The primary differtreatment of obesity in the obese (ob/ob) mouse model. ence in the two therapeutic approaches is the continuous We demonstrate an improved correction profile for obesity chronic secretion of leptin mediated by gene delivery, verand associated surrogate markers using the adenovirus sus the intermittent bolus delivery and rapid clearance of delivery method. Rate of weight loss and percentage satthe daily injection of rh-leptin protein. Thus, in vivo findings iety were significantly greater in the mice treated with Adsuggest that leptin effects are better achieved at lower leptin. These findings were associated with lower peak steady-state levels, a pharmacological feature attained serum leptin levels with Ad-leptin (22.9 ± 2.6 ng/ml for the here by gene therapy. These findings may have implihuman gene, and 48.9 ± 11.5 ng/ml for the murine gene) cations for the potential use of leptin in the treatment of compared to rh-leptin (385.2 ± 36.0 ng/ml). (Values are obesity.

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This paper gives an overview of research on the field of structure vibration-attitude coordinated control of spacecraft. First of all, the importance of the technology has been given an introduction, and then later the research progress of space structure dynamics modeling, research progress of structure vibration-attitude coordinated control of flexible spacecraft have been discussed respectively. Finally, future research on application of structure vibration-attitude coordinated control of spacecraft has been recommended.