bluuebunny/crossref_metadata_2025_split huggingface dataset:
Increased non‐viral gene transfer levels in mice by concentration of cationic lipid DNA complexes formed under optimized conditions
Authors: Nchinda Godwin , Zschörnig Olaf , Überla Klaus 
Published: July 2003

AbstractBackgroundA number of non‐viral gene transfer reagents including cationic lipid DNA complexes (LDC) have been developed and were improved by changing the ratio of their components. To determine the effect of other parameters during complex formation affecting the efficacy of LDC, the conditions during complexation were varied without changing the ratios of the components.MethodsLDC were formed at fixed ratios of an equimolar mixture of DOTAP and cholesterol to DNA to the condensing agent protamine sulfate according to different protocols at varying final concentrations. The influence of these parameters on transfection efficiency and physical properties of the complexes was determined.ResultsChanging the order of addition of compounds during complex formation affected the size distribution, the charge of the LDC, the interaction between the lipids and the accessibility of the DNA. At fixed ratios of the components, higher transfection efficiencies were observed with more condensed LDC. Complexation in higher volumes increased transduction efficiency of the complexes after intravenous inoculation. Due to restrictions on the injectable volume, the LDC were formed in the optimal volume and subsequently concentrated by ultrafiltration. The concentrated complexes maintained transduction efficiency and up to 60‐fold higher in vivo transduction levels were obtained.ConclusionsIn addition to the ratio of the components of cationic lipid DNA complexes, the final concentration and the order of addition of compounds during complex formation are critical for high transduction efficiency. Concentration of LDC formed under optimal conditions can be used to further increase in vivo gene transfer levels. Copyright © 2003 John Wiley & Sons, Ltd.
Proliferative activity and in vitro replication of HSV1716 in human metastatic brain tumours
Authors: Detta Allah , Harland June , Hanif Isma , Brown S. Moira , Cruickshank Garth 
Published: July 2003

AbstractBackgroundThe neurotropic herpes simplex virus mutant HSV1716 lacks the gene encoding the virulence factor ICP34.5 and cannot replicate in non‐dividing cells where proliferating cell nuclear antigen (PCNA) is not actively engaged in cellular DNA synthesis. In the brain, tumoral expression of PCNA therefore confers on it oncolytic specificity and may determine its efficacy. Three phase I trials in glioma patients and one in metastatic melanoma patients have established that HSV1716 is safe and replicates selectively in tumour tissue. Here we examine the in situ PCNA profiles of common human metastatic brain tumours and determine their in vitro permissivity for HSV1716 replication to ascertain their suitability for HSV1716 therapy.MethodsHistological sections of tumour biopsies obtained from patients undergoing craniotomies were stained for PCNA expression. The replicative ability of HSV wild‐type (17+) and mutant (1716) viruses was assessed in tissue cultures of the same tumour biopsies and in cancer cell lines by plaque assay.ResultsBiopsies of all 10 metastatic tumours (3 melanoma, 4 carcinoma and 3 adenocarcinoma) as well as 4 glioblastoma multiforme were positive for PCNA immunoexpression and supported the replication of HSV1716. The PCNA‐positive cells in the metastatic tumours were distributed comparatively in larger and more contiguous areas than in glioblastoma (1.69 ± 1.61 mm2 vs. 0.73 ± 0.77 mm2) and numbered 29.0 ± 12.4 and 12.6 ± 4.7%, respectively.ConclusionsThe results show that human cerebral metastatic tumours have generally larger and more contiguous proliferative areas, support efficient HSV1716 replication, and are thus potential candidates for such oncolytic viral therapy. Copyright © 2003 John Wiley & Sons, Ltd.
Phase I immunotherapy with a modified vaccinia virus (MVA) expressing human MUC1 as antigen‐specific immunotherapy in patients with MUC1‐positive advanced cancer
Authors: Rochlitz Christoph , Figlin Robert , Squiban Patrick , Salzberg Marc , Pless Miklos , Herrmann Richard , Tartour Eric , Zhao Yongxiang , Bizouarne Nadine , Baudin Martine , Acres Bruce 
Published: July 2003

AbstractBackgroundThe MUC1 protein is a highly glycosylated mucin normally found at the apical surface of mucin‐secreting epithelial cells in many types of tissues. MUC1 is expressed, but heavily underglycosylated, in different human tumors. TG4010 is a viral suspension of a recombinant vaccinia vector (MVA) containing DNA sequences coding for the human MUC1 antigen and interleukin‐2 (IL‐2). This product was developed for use as a vaccine in cancer patients whose tumors express the MUC1 antigen. The objective of the present study was to determine the safety of the product and to define the dose of TG4010 to be used in further clinical trials.Materials and methodsThirteen patients with different solid tumors were treated by repeated intramuscular injection with increasing doses of TG4010 in two separate phase I studies, one in Europe (Basel—CR) and one in the United States (UCLA—RF): a total of 6 patients were treated at a dose of 5 × 106 pfu, 3 patients at 5 × 107 pfu, and 4 patients at 108 pfu. Safety, efficacy, and different immunological tests were the endpoints of the study.ResultsTolerance of TG4010 was excellent, and side effects mainly consisted of injection site pain and influenza‐like symptoms. There was no apparent detrimental effect of repeated injections of the vaccinia virus. Four of thirteen evaluable patients showed stabilization of their disease for 6 to 9 months. One lung cancer patient who was initially progressing after the first injections later showed a marked decrease in the size of his metastases that lasted for 14 months. Some T cell proliferative immune responses were seen in five patients.ConclusionsThe administration of TG4010 was generally well tolerated in patients with metastatic tumors, and transient disease stabilization was observed in several patients, warranting further clinical studies with the product. Copyright © 2003 John Wiley & Sons, Ltd.
Lentivirally transduced dendritic cells as a tool for cancer immunotherapy
Authors: Breckpot Karine , Dullaers Melissa , Bonehill Aude , Van Meirvenne Sonja , Heirman Carlo , De Greef Catherine , van der Bruggen Pierre , Thielemans Kris 
Published: July 2003

AbstractBackgroundDendritic cells (DC) are the professional antigen‐presenting cells of the immune system, fully equipped to prime naive T cells and thus essential components for cancer immunotherapy.MethodsWe tested the influence of several elements (cPPT, trip, WPRE, SIN) on the transduction efficiency of human DC. Human and murine DC were transduced with tNGFR‐encoding lentiviruses to assess the effect of transduction on phenotype and function. Human DC were transduced with lentiviruses encoding huIi80MAGE‐A3 and murine DC with huIi80tOVA to test antigen presentation.ResultsA self‐inactivating (SIN) lentiviral vector containing the trip element was most efficient in transducing human DC. The transduction of DC with trip/SIN tNGFR encoding lentiviral vectors at MOI 15 resulted in stable gene expression in up to 94.6% (murine) and 88.2% (human) of the mature DC, without perturbing viability, phenotype and function. Human huIi80MAGE‐A3‐transduced DC were able to stimulate MAGE‐A3‐specific CD4+ and CD8+ T cell clones and could prime both MAGE‐A3‐specific CD4+ and CD8+ T cells in vitro. Murine huIi80tOVA‐transduced DC were able to present OVA peptides in the context of MHC class I and class II in vitro and induced a strong OVA‐specific cytotoxic T lymphocyte response in vivo, that was protective against subsequent challenge with OVA‐expressing tumor cells.ConclusionsWe show that, using lentiviral vectors, efficient gene transfer in human and murine DC can be obtained and that these DC can elicit antigen‐specific immune responses in vitro and in vivo. The composition of the transfer vector has a major impact on the transduction efficiency. Copyright © 2003 John Wiley & Sons, Ltd.
Low molecular weight protamine as an efficient and nontoxic gene carrier: in vitro study
Authors: Park Yoon Jeong , Liang Jun Feng , Ko Kyung Soo , Kim Sung Wan , Yang Victor C. 
Published: July 2003

AbstractBackgroundThe structural similarity between low molecular weight protamine (LMWP), prepared by enzymatic digestion of protamine, and HIV‐TAT protein transduction peptide suggested the feasibility of LMWP as an efficient carrier for delivering therapeutic genes while alleviating the cytotoxicity of currently employed gene carriers.MethodsLMWP was prepared by enzymatic digestion of protamine with thermolysine. The prepared LMWP peptide and TAT peptide, as well as their complexes with plasmid DNA (pDNA), were examined for cellular uptake behaviors by using confocal microscopy and flow cytometry. The complexation of pDNA and LMWP was monitored by gel retardation test as well as size and zeta potential measurements, and was then further assessed by DNase I protection assay. The transfection efficiency of pDNA/LMWP was examined by varying the pDNA content and charge ratio in the complex, and then compared with that of pDNA/PEI. Cytotoxicity induced by pDNA/LMWP and pDNA/PEI was also examined.ResultsPrepared LMWP showed similar transcellular localization behavior and kinetics to those of TAT, and efficiently transferred the pDNA into nucleus and cytoplasm in a short time period. The size and zeta potential of the pDNA/LMWP complex were 120 nm and 30 mV, respectively, which were adequately suitable for cellular uptake. After forming the complex, LMWP appeared to effectively protect pDNA against DNase I attack. The pDNA/LMWP complex showed significantly enhanced gene transfer than both naked pDNA and the pDNA/PEI complex, while exhibiting a markedly reduced cytotoxicity than that of the pDNA/PEI complex.ConclusionsThe present study suggested that LMWP could be a useful and safe tool for enhancing delivery of bioactive molecules and therapeutic DNA products into cells when applied in gene therapy. Copyright © 2003 John Wiley & Sons, Ltd.
Gene therapy for HIV/AIDS: the potential for a new therapeutic regimen
Authors: Fanning Greg , Amado Rafael , Symonds Geoff 
Published: July 2003

AbstractHuman Immunodeficiency Virus (HIV) is the etiologic agent of Acquired Immunodeficiency Syndrome (AIDS). HIV/AIDS is a disease that, compared with the not so distant past, is now better held in check by current antiretroviral drugs. However, it remains a disease not solved. Highly active antiretroviral therapy (HAART) generally uses two non‐nucleoside and one nucleoside reverse transcriptase (RT) inhibitor or two non‐nucleoside RT and one protease inhibitor. HAART is far more effective than the mono‐ or duo‐therapy of the past, which used compounds like the nucleoside reverse transcriptase inhibitor AZT or two nucleoside reverse transcriptase inhibitors. However, even with the relatively potent drug cocktails that comprise HAART, there are the issues of (i) HIV escape mutants, (ii) an apparent need to take the drugs in an ongoing manner, and (iii) the drugs' side effects that are often severe. This review speaks to the potential addition to these potent regimens of another regimen, namely the genetic modification of target hematopoietic cells. Such a new treatment paradigm is conceptually attractive as it may yield the constant intracellular expression of an anti‐HIV gene that acts to inhibit HIV replication and pathogenicity. A body of preclinical work exists showing the inhibition of HIV replication and decreased HIV pathogenicity by anti‐HIV genetic agents. This preclinical work used hematopoietic cell lines and primary cells as the target tissue. More recently, several clinical trials have sought to test this concept in vivo. Copyright © 2003 John Wiley & Sons, Ltd.
Molecular electrostatic potential as a factor of drug–receptor recognition
Authors: Dukhovich Felix S. , Darkhovskii Michael B. 
Published: July 2003

AbstractWhen a drug molecule approaches a non‐specific acceptor both molecules are in electrostatic fields of equal sign which prevents drug–acceptor complex formation. At the same time, the drug–acceptor system does not achieve the thermodynamic global minimum. Otherwise, when a certain drug interacts with its specific receptor, mutual compensation of their molecular electrostatic potentials takes place. Then separate atoms and groups of the drug molecule can bind to the receptor. We show that the fundamental role of molecular electrostatic potential in the process of drug–receptor recognition consists in fast correction of errors. Copyright © 2003 John Wiley & Sons, Ltd.
Long‐range and short‐range mechanisms of hydrophobic attraction and hydrophilic repulsion in specific and aspecific interactions
Authors: van Oss Carel Jan 
Published: July 2003

AbstractAmong the three different non‐covalent forces acting in aqueous media, i.e. Lifshitz–van der Waals (LW), Lewis acid–base (AB) and electrical double layer (EL) forces, the AB forces or electron–acceptor/electron–donor interactions are quantitatively by far the predominant ones. A subset of the AB forces acting in water causes the hydrophobic effect, which is the attraction caused by the hydrogen‐bonding (AB) free energy of cohesion between the water molecules which surround all apolar as well as polar molecules and particles when they are immersed in water. As the polar energy of cohesion among water molecules is an innate property of water, the hydrophobic attraction (due to the hydrophobic effect) is unavoidably always present in aqueous media and has a value of ΔGhydrophobic = −102 mJ/m2, at 20 °C, being equal to the AB free energy of cohesion between the water molecules at that temperature. The strong underlying hydrophobic attraction due to this effect can, however, be surmounted by very hydrophilic molecules and particles that attract water molecules more strongly than the free energy of attraction of these molecules or particles for one another, plus the hydrogen‐bonding free energy of cohesion between the water molecules, thus resulting in a net non‐electrical double layer repulsion. Each of the three non‐covalent forces, LW, AB or EL, any of which can be independently attractive or repulsive, decays, dependent on the circumstances, as a function of distance according to different rules. These rules, following an extended DLVO (XDLVO) approach, are given, as well as the measurement methods for the LW, AB and EL surface thermodynamic properties, determined at ‘contact’. The implications of the resulting hydrophobic attractive and hydrophilic repulsive free energies, as a function of distance, are discussed with respect to specific and aspecific interactions in biological systems. The discussion furnishes a description of the manner by which shorter‐range specific attractions can surmount the usually much stronger long‐range aspecific repulsion, and ends with examples of in vitro and in vivo effects of hydrophilization of biopolymers, particles or surfaces by linkage with polyethylene oxide (PEO; also called polyethylene glycol, PEG). Copyright © 2003 John Wiley & Sons, Ltd.
Analysis of the pH‐dependencies of the association and dissociation kinetics of HIV‐1 protease inhibitors
Authors: Gossas Thomas , Danielson U. Helena 
Published: July 2003

AbstractThe kinetic constants for the interactions between HIV‐1 protease and a selection of inhibitors were determined at different pH‐values using a biosensor based interaction assay. Since this technique does not involve a substrate, it was possible to determine the pH‐dependencies of the association and dissociation rates of an inhibitor, without the complication of a pH‐dependent enzyme‐substrate/product equilibrium. The importance of these interactions was evaluated by correlating the free energy changes upon association and dissociation of inhibitors with the predicted change in electrostatic properties of the interacting groups as a result of altered pH. It was found that the kinetic parameters varied with pH in a unique manner for all inhibitors, demonstrating that the kinetic features were associated with the specific structure of each inhibitor. Association and dissociation had different pH‐profiles, indicating that the two processes proceeded by different pathways/mechanisms. The energy barrier for dissociation of the enzyme–indinavir complex increased with pH from 4.1 to 7.4, while it was generally reduced for the other inhibitors as the pH was increased from 5.1 to 7.4. The pH‐dependent interactions involved in the recognition/binding of inhibitors and in the stabilization of the complex were identified by analysing three‐dimensional structures of enzyme–inhibitor complexes. The interaction between the pyridine nitrogen of indinavir with Arg‐8 was hypothesized to be responsible for the unique pH‐dependency of indinavir. The analysis revealed features of interactions that are significant for understanding enzyme function and for optimization of new drug leads. It also highlighted the importance of environmental conditions on interactions. Copyright © 2003 John Wiley & Sons, Ltd.
Directional shape complementarity at the protein–DNA interface
Authors: Yeh Ching‐Sheng , Chen Fang‐Ming , Wang Jaw‐Yuan , Cheng Tian‐Lu , Hwang Ming‐Jing , Tzou Wen‐Shyong 
Published: July 2003

AbstractNature utilizes various styles of architecture for DNA‐binding proteins to recognize diverse DNA sequences, a process facilitated by a complementary surface between protein and DNA. However, the extent and ways this ‘shape complementarity’ occurs at the protein–DNA interface have yet to be characterized. Here, by analyzing a set of diverse protein–DNA complexes of known three‐dimensional structures, we investigated whether the normal vectors of a protein surface at the interface exhibited any relationship with DNA conformation. Generally, the normal vectors of a DNA‐contacting protein surface distinctly preferred certain angles, enabling them to align with certain axes characterizing the conformation of DNA. Thus, a new geometric property of DNA‐binding protein is demonstrated, i.e. the ‘shape complementarity’ of protein–DNA recognition clearly bears the property of ‘directionality’. Copyright © 2003 John Wiley & Sons, Ltd.
Solid supports for microarray immunoassays
Authors: Kusnezow Wlad , Hoheisel Jörg D. 
Published: July 2003

AbstractStimulated by the achievements of the first phase in genomics and the resulting need of assigning functions to the acquired sequence information, novel formats of immunoassays are being developed for high‐throughput multi‐analyte studies. In principle, they are similar in nature to the microarray assays already established at the level of nucleic acids. However, the biochemical diversity and the sheer number of proteins are such that an equivalent analysis is much more complex and thus difficult to accomplish. The wide range of protein concentration complicates matters further. Performing microarray immunoassays already represents a challenge at the level of preparing a working chip surface. Arrays have been produced on filter supports, in microtiter plate wells and on glass slides, the last two usually coated with one‐, two‐ or three‐dimensionally structured surface modifications. The usefulness and suitability of all these support media for the construction and application of antibody microarrays are reviewed in this manuscript in terms of the different kinds of immunoassay and the various detection procedures. Additionally, the employment of microarrays containing alternative sensor molecules is discussed in this context. The sensitivity of microspot immunoassays predicted by the current analyte theory is not yet a reality, indicating the extent of both the technology's potential and the size of the task still ahead. Copyright © 2003 John Wiley & Sons, Ltd.
Derivatization reaction of the mycotoxin moniliformin with 1,2‐diamino‐4,5‐dichlorobenzene: structure elucidation of an unexpected reaction product by liquid chromatography/tandem mass spectrometry and liquid chromatography/nuclear magnetic resonance spectroscopy
Authors: Zöllner Peter , Lienau Annette , Albert Klaus , Lindner Wolfgang 
Published: July 2003

AbstractThe derivatization reaction of the mycotoxin moniliformin with 1,2‐diamino‐4,5‐dichlorobenzene was previously introduced to improve distinctly the sensitivity of an assay applying high‐performance liquid chromatography prior to fluorescence detection. In the course of the implementation of this derivatization approach into a liquid chromatographic/mass spectrometric method, an unexpected derivatization product has now been discovered by mass spectrometry. In order to elucidate its structure, detailed investigations with liquid chromatography/tandem mass spectrometry and liquid chromatography coupled on‐line with NMR spectroscopy were performed. These studies give evidence for a heterocyclic structure that has been formed by the loss of one water and one carbon monoxide molecule. A reasonable mechanism for this derivatization reaction is proposed. Copyright © 2003 John Wiley & Sons, Ltd.
Internal residue loss produced by rearrangement of a novel cationic glycosphingolipid, glyceroplasmalopsychosine, in collision‐induced dissociation
Authors: Tadano‐Aritomi Keiko , Hikita Toshiyuki , Kubota Masayuki , Kasama Takeshi , Toma Kazunori , Hakomori Sen‐itiroh , Ishizuka Ineo 
Published: July 2003

AbstractA novel plasmal conjugate of galactosylsphingosine (psychosine), Gro1(3)‐O‐plasmal‐O‐6Galβ‐sphingosine (glyceroplasmalopsychosine), was analyzed by electrospray ionization and liquid secondary ion mass spectrometry with low‐ or high‐energy collision‐induced dissociation (CID). In the product ion spectra of the [M + H]+ ions, [M + H − glycerol]+ ions arising from the loss of a glycerol were predominant. Unexpectedly, CID of the [M + H − glycerol]+ ion produced an outstanding ion, [(M + H − glycerol) − Hex]+, which required the loss of the galactose from inside the molecule. This ion was greatly reduced in the spectra of N,N‐dimethyl derivatives, indicating that the [(M + H − glycerol) − Hex]+ ion is formed from an intramolecular rearrangement with migration of the plasmal residue to the free amino group of sphingosine. It would be expected that the rearrangement occurs simultaneously with the elimination of glycerol or a rearranged [M + H]+ ion leads to the elimination of glycerol, to form a Schiff base‐type [M + H − glycerol]+ ion, from which the terminal galactose could be removed by the normal mechanism of glycosidic cleavage. On the other hand, the [M + Na − glycerol]+ ion derived from the sodiated molecule did not produce an ion corresponding to the rearrangement reaction, possibly owing to a higher stability of the sodiated ions against conformational changes. Copyright © 2003 John Wiley & Sons, Ltd.