An Activated O N Acyl Transfer Auxiliary: Efficient Amide-Backbone Substitution of Hindered "Difficult" Peptides

Overcoming the phenomenon known as difficult synthetic sequences has been a major goal in solid-phase peptide synthesis for over 30 years. In this work the advantages of amide backbone-substitution in the solid-phase synthesis of difficult peptides are augmented by developing an activated N-alpha-acyl transfer auxiliary. Apart from disrupting troublesome intermolecular hydrogen-bonding networks, the primary function of the activated N-alpha-auxiliary was to facilitate clean and efficient acyl capture of large or beta-branched amino acids and improve acyl transfer yields to the secondary N-alpha-amine. We found o-hydroxyl-substituted nitrobenzyl (Hnb) groups were suitable N-alpha-auxiliaries for this purpose. The relative acyl transfer efficiency of the Hnb auxiliary was superior to the 2-hydroxy-4-methoxybenzyl (Hmb) auxiliary with protected amino acids of varying size. Significantly, this difference in efficiency was more pronounced between more sterically demanding amino acids. The Hnb auxiliary is readily incorporated at the N-alpha-amine during SPPS by reductive alkylation of its corresponding benzaldehyde derivative and conveniently removed by mild photolysis at 366 nm. The usefulness of the Hnb auxiliary for the improvement of coupling efficiencies in the chain-assembly of difficult peptides was demonstrated by the efficient Hnb-assisted Fmoc solid-phase synthesis of a known hindered difficult peptide sequence, STAT-91. This work suggests the Hnb auxiliary will significantly enhance our ability to synthesize difficult polypeptides and increases the applicability of amide-backbone substitution.

Polyinosinic acid and polycationic liposomes attenuate the hepatic clearance of circulating plasmid DNA

DNA that enters the circulation is rapidly cleared both by tissue uptake and by DNase-mediated degradation. In this study, we have examined the uptake of linear plasmid DNA in an isolated perfused liver model and following intra-arterial administration to rats. We found that the DNA was rapidly taken up by the isolated perfused liver without degradation. The single-pass extraction ratio was 0.76 +/- 0.05, the mean transit time was 15.3 +/- 3.6 s, and the volume of distribution was 0.29 +/- 0.07 ml/g. Hepatic uptake was saturable and was inhibited by polyinosinic acid or polycationic liposomes but not by condensation of the DNA with polylysine. When the linear plasmid DNA was administered in vivo, plasma half-life was 3.1 +/- 0.2 min, volume of distribution was 670 +/- 85 ml/kg, and clearance was 32 +/- 4 min. Coadministration of cationic liposomes decreased the volume of distribution to 180 +/- 28 ml/kg as well as the half-life (2.6 +/- 0.2 min). By contrast, polyinosinic acid significantly increased the circulating half-life (7.7 +/- 0.5 min), decreased the volume of distribution (95 +/- 17 ml/kg), and partially inhibited DNA degradation. When administered along with the liposomes and the polyinosinic acid, the distribution of plasmid-derived radioactivity decreased in the liver and increased in most other peripheral tissues. This study shows that pharmacological manipulation of the uptake and degradation of DNA can alter its distribution and clearance in vivo. These results may be useful in optimizing gene delivery procedures for in vivo gene therapy.

Transfer matrix eigenvalues of the anisotropic multiparametric U model

A multiparametric extension of the anisotropic U model is discussed which maintains integrability. The R-matrix solving the Yang-Baxter equation is obtained through a twisting construction applied to the underlying U-q(sl (2/1)) superalgebraic structure which introduces the additional free parameters that arise in the model. Three forms of Bethe ansatz solution for the transfer matrix eigenvalues are given which we show to be equivalent.

Modeling of heat transfer in fluidized-bed coating of cylinders

A numerical model of heat transfer in fluidized-bed coating of solid cylinders is presented. By defining suitable dimensionless parameters, the governing equations and its associated initial and boundary conditions are discretized using the method of orthogonal collocation and the resulting ordinary differential equations simultaneously solved for the dimensionless coating thickness and wall temperatures. Parametric Studies showed that the dimensionless coating thickness and wall temperature depend on the relative heat capacities of the polymer powder and object, the latent heat of fusion and the size of the cylinder. Model predictions for the coating thickness and wall temperature compare reasonably well with numerical predictions and experimental coating data in the literature and with our own coating experiments using copper cylinders immersed in nylon-11 and polyethylene powders. (C) 2001 Elsevier Science Ltd. All rights reserved.

Lentiviral vector-mediated tyro sinase-related protein 2 gene transfer to dendritic cells for the therapy of melanoma

Dendritic cells (DCs) are the most potent professional antigen-presenting cells (APCs), which play a vital role in primary immune responses. Introducing genes into DCs will allow constitutive expression of the encoded proteins and thus prolong the presentation of the antigens derived therefrom. In addition, multiple and unidentified epitopes encoded by the entire tumor-associated antigen (TAA) gene may enhance T cell activation. This study demonstrated that an HIV-1-based lentiviral vector conferred efficient gene transfer to DCs. The transgene, murine tyrosinase-related protein 2 (mTRP-2), encodes a clinically relevant melanoma-associated antigen (MAA), which has been found to be a tumor rejection antigen for B16 melanoma. The transfer and proper processing of mTRP-2 in DCs, in terms of RNA transcription activity and protein expression, were verified by RT-PCR and specific antibody, respectively. Administration of mTRP-2 gene-modified DCs (DC-HR'CmT2) to C57BL/6 mice evoked strong protection against tumor challenge, for which the presence of CD4(+) and CD8(+) cells during both the priming and challenge phase was essential. In a therapy model, our results showed that four of seven mice with preestablished tumor remained tumor free for 80 days after therapeutic vaccination. Given the results shown in this study, mTRP-2 gene transfer to DCs provides a potential therapeutic strategy for the management of melanoma, especially in the early stage of the disease.

Finite element modelling of heat transfer through permeable cracks in hydrothermal systems with upward throughflow

We use the finite element method to model the heat transfer phenomenon through permeable cracks in hydrothermal systems with upward throughflow. Since the finite element method is an approximate numerical method, the method must be validated before it is used to soh,e any new, kind of problem. However, the analytical solution, which can be used to validate the finite element method and other numerical methods, is rather limited in the literature, especially, for the problem considered here. Keeping this in mind, we have derived analytical solutions for the temperature distribution along the vertical axis of a crack in a fluid-saturated porous layer. After the finite element method is validated by comparing the numerical solution with the analytical solution for the same benchmark problem, it is used to investigate the pore-fluid flow and heat transfer in layered hydrothermal systems with vertical permeable cracks. The related analytical and numerical results have demonstrated that vertical cracks are effective and efficient members to transfer heat energy from the bottom section to the top section in hydrothermal systems with upward throughflow.

Magnetization transfer imaging for polymer gel dosimetry

Off-resonance RF pre-saturation was used to obtain contrast in MRI images of polymer gel dosimeters irradiated to doses up to 50 Gy. Two different polymer gel dosimeters composed of 2-hydroxyethyl-acryl ate or methacrylic acid monomers mixed with N, N'-methylene-bisacrylamide (BIS), dispersed in an aqueous gelatin matrix were evaluated. Radiation-induced polymerization of the co-monomers generates a fast-relaxing insoluble polymer. Saturation of the polymer using off-resonance Gaussian RF pulses prior to a spin-echo read-out with a short echo time leads to contrast that is dependent on the absorbed dose. This contrast is attributed to magnetization transfer (MT) between free water and the polymer, and direct saturation of water was found to be negligible under the prevailing experimental conditions. The usefulness of MT imaging was assessed by computing the dose resolution obtained with this technique. We found a low value of dose resolution over a wide range of doses could be obtained with a single experiment. This is an advantage over multiple spin echo (MSE) experiments using a single echo spacing where an optimal dose resolution is achieved over only very limited ranges of doses. The results suggest MT imaging protocols may be developed into a useful tool for polymer gel dosimetry.

Analysis of steady-state heat transfer through mid-crustal vertical cracks with upward throughflow in hydrothermal systems

We conduct a theoretical analysis of steady-state heat transfer problems through mid-crustal vertical cracks with upward throughflow in hydrothermal systems. In particular, we derive analytical solutions for both the far field and near field of the system. In order to investigate the contribution of the forced advection to the total temperature of the system, two concepts, namely the critical Peclet number and the critical permeability of the system, have been presented and discussed in this paper. The analytical solution for the far field of the system indicates that if the pore-fluid pressure gradient in the crust is lithostatic, the critical permeability of the system can be used to determine whether or not the contribution of the forced advection to the total temperature of the system is negligible. Otherwise, the critical Peclet number should be used. For a crust of moderate thickness, the critical permeability is of the order of magnitude of 10(-20) m(2), under which heat conduction is the overwhelming mechanism to transfer heat energy, even though the pore-fluid pressure gradient in the crust is lithostatic. Furthermore, the lower bound analytical solution for the near field of the system demonstrates that the permeable vertical cracks in the middle crust can efficiently transfer heat energy from the lower crust to the upper crust of the Earth. Copyright (C) 2002 John Wiley Sons, Ltd.

Effects of hot intrusions on pore-fluid flow and heat transfer in fluid-saturated rocks

We use the finite element method to solve coupled problems between pore-fluid flow and heat transfer in fluid-saturated porous rocks. In particular, we investigate the effects of both the hot pluton intrusion and topographically driven horizontal flow on the distributions of the pore-flow velocity and temperature in large-scale hydrothermal systems. Since general mineralization patterns are strongly dependent on distributions of both the pore-fluid velocity and temperature fields, the modern mineralization theory has been used to predict the general mineralization patterns in several realistic hydrothermal systems. The related numerical results have demonstrated that: (1) The existence of a hot intrusion can cause an increase in the maximum value of the pore-fluid velocity in the hydrothermal system. (2) The permeability of an intruded pluton is one of the sensitive parameters to control the pore-fluid flow, heat transfer and ore body formation in hydrothermal systems. (3) The maximum value of the pore-fluid velocity increases when the bottom temperature of the hydrothermal system is increased. (4) The topographically driven flow has significant effects on the pore-fluid flow, temperature distribution and precipitation pattern of minerals in hydrothermal systems. (5) The size of the computational domain may have some effects on the pore-fluid flow and heat transfer, indicating that the size of a hydrothermal system may affect the pore-fluid flow and heat transfer within the system. (C) 2003 Elsevier Science B.V. All rights reserved.

Transfer and Reverse Transfer of Knowledge of International Acquisitions: the case of the acquisition of the Perez Companc by Petrobras in Argentina

The objective of this article was to analyze the processes of transfer and reverse trans fer of knowledge following. international acquisitions made by Brazilian multinational companies. Reverse transfer is understood,as the process of transferring knowledge from the acquired company to the acquirer. Therefore, a case study was conducted on the acquisition of the Perez Companc group by Petrobras in Argentina. The study is qualitative. Primary data were obtained and eight members of the international managing board of Petrobras were interviewed. After the first moment of integration, reported as conflictive, there was a better integration of the companies, mainly in the technical areas of, the oil and gas exploration activities. The size of Perez Companc, its aim (a company of energy, not only oil and gas company) and the length of time were critical factors for the transfer of best practices between the companies. The expatriation of the employees is seen as a key-tool, as well as the technical visits, for the transfer of knowledge.. An. additional contribution of the study was to present the results of the research on the process of transfer and reverse transfer of knowledge in Brazilian multinational companies, since most studies on the theme focus on the motivators and challenges concerning these processes.

Efficient Phosphodiester Hydrolysis by Luminescent Terbium(III) and Europium(III) Complexes

The synthesis and structures of two new isostructural mononuclear [Ln(L)(NO(3))(H(2)O)(3)](NO(3))(2) complexes, with Ln = Tb (complex 1) and Eu (complex 2), which display high activity in the hydrolysis of the substrate 2,4-bis(dinitrophenyl)phosphate, are reported. These complexes displayed catalytic behavior similar to the mononuclear gadolinium complex [Gd(L)(NO(3))(H(2)O)(3)](NO(3))(2) previously reported by us (lnorg. Chem. 2008, 47, 2919-2921); one hydrolysis reaction in two stages where the diesterase and monoesterase activities could be monitored separately, with the first stage dependent on and the second independent of the complex concentration. Through potentiometric studies, electrospray ionization mass spectrometry (ESI-MS) analysis, and determination of the kinetic behaviors of 1 and 2 in acetonitrile/water solution, the species present in solution could be identified and suggested a dinuclear species, with one hydroxo group, as the most prominent catalyst under mild conditions. The complexes show high activity (k(1)= 7 and 18 s(-1) for 1 and 2, respectively) and catalytic efficiency. Complexes 1 and 2 were found to be active toward the cleavage of plasmid DNA, and complete kinetic studies were carried out. Studies with a radical scavenger (dimethylsulfoxide) confirmed the hydrolytic action of 1 and 2 in the cleavage of DNA. Studies on the incubation of distamycin with plasmid DNA suggested that 1 and 2 are regio-specific, interacting with the minor groove of DNA. These complexes displayed luminescent properties. Complex 1 showed higher emission intensity than 2 due to a more efficient energy transfer between triplet and emission levels of terbium (T -> (5)D(4)), along with nonradiative deactivation mechanisms of the excited states of europium via multiphonon decays and the ligand-to-metal charge transfer state. Lifetime measurements of the (5)D(4) and (5)D(0) excited levels for 1 and 2, respectively, indicated the numbers of coordinated water molecules for the complexes.

Adoptive transfer of gene-engineered CD4+ helper T cells induces potent primary and secondary tumor rejection

Because CD4(+) T cells play a key role in aiding cellular immune responses, we wanted to assess whether increasing numbers of gene-engineered antigen-restricted CD4(+) T cells could enhance an antitumor response mediated by similarly gene-engineered CD8(+) T cells. In this study, we have used retroviral transduction to generate erbB2-reactive mouse T-cell populations composed of various proportions of CD4(+) and CD8(+) cells and then determined the antitumor reactivity of these mixtures. Gene-modified CD4(+) and CD8(+) T cells were shown to specifically secrete Tc1 (T cytotoxic-1) or Tc2 cytokines, proliferate, and lyse erbB2(+) tumor targets following antigen ligation in vitro. In adoptive transfer experiments using severe combined immunodeficient (scid) mice, we demonstrated that injection of equivalent numbers of antigen-specific engineered CD8(+) and CD4(+) T cells led to significant improvement in survival of mice bearing established lung metastases compared with transfer of unfractionated (largely CD8(+)) engineered T cells. Transferred CD4(+) T cells had to be antigen-specific (not just activated) and secrete interferon gamma (IFN-gamma) to potentiate the antitumor effect. Importantly, antitumor responses in these mice correlated with localization and persistence of gene-engineered T cells at the tumor site. Strikingly, mice that survived primary tumor challenge could reject a subsequent re-challenge. Overall, this study has highlighted the therapeutic potential of using combined transfer of antigen-specific gene-modified CD8(+) and CD4(+) T cells to significantly enhance T-cell adoptive transfer strategies for cancer therapy.

Wavefront analysis and modulation transfer function of three multifocal intraocular lenses

Purpose: To evaluate wavefront performance and modulation transfer function (MTF) in the human eye aft er the implantation of diffractive or refractive multifocal intraocular lenses (IOLs). Materials and Methods: This was a prospective, interventional, comparative, nonrandomized clinical study. Uncorrected distance and near visual acuity, and wavefront analysis including MTF curves (iTrace aberrometer, Tracey Technologies, Houston, TX, USA) were measured in 60 patients aft er bilateral IOL implantation with 6 months of follow-up. Forty eyes received the diffractive ReSTOR (Alcon), 40 eyes received the refractive ReZoom (Advanced Medical Optics) and 40 eyes, the Tecnis ZM900 (Advanced Medical Optics). The comparison of MTF and aberration between the intraocular lenses was performed using analysis of variance (ANOVA), followed by the Dunn test when necessary. Results: The mean uncorrected distance visual acuity was similar in all three groups of multifocal IOLs. The ReSTOR group provided better uncorrected near visual acuity than the ReZoom group (P < 0.001), but similar to the Tecnis group. Spherical aberration was significantly higher in the ReZoom group (P = 0.007). Similar MTF curves were found for the aspheric multifocal IOL Tecnis and the spheric multifocal IOL ReSTOR, and both performed better than the multifocal IOL ReZoom in a 5 mm pupil (P < 0.001 at all spatial frequencies). Conclusions: Diffractive IOLs studied presented similar MTF curves for a 5 mm pupil diameter. Both diffractive IOLs showed similar spherical aberration, which was significantly better with the full-diffractive IOL Tecnis than with the refractive IOL ReZoom.