Preclinical studies for pharmacokinetics and biodistribution of Ad-stTRAIL, an adenovirus delivering secretable trimeric TRAIL for gene therapy

Malignant glioma is the most frequent type in brain tumors. The prognosis of this tumor has not been significantly improved for the past decades and the average survival of patients is less than one year. Thus, an effective novel therapy is urgently needed. TNF-related apoptosis inducing ligand (TRAIL), known to have tumor cell-specific killing activity, has been investigated as a novel therapeutic for cancers. We have developed Ad-stTRAIL, an adenovirus delivering secretable trimeric TRAIL for gene therapy and demonstrated the potential to treat malignant gliomas. Currently, this Ad-stTRAIL gene therapy is under phase I clinical trial for malignant gliomas. Here, we report preclinical studies for Ad-stTRAIL carried out using rats. We delivered Ad-stTRAIL intracranially and determined its pharmacokinetics and biodistribution. Most Ad-stTRAIL remained in the delivered site and the relatively low number of viral genomes was detected in the opposite site of brain and cerebrospinal fluid. Similarly, only small portion of the viral particles injected was found in the blood plasma and major organs and tissues, probably due to the brain-blood barrier. Multiple administrations did not lead to accumulation of Ad-stTRAIL at the injection site and organs. Repeated delivery of Ad-stTRAIL did not show any serious side effects. Our data indicate that intracranially delivered Ad-stTRAIL is a safe approach, demonstrating the potential as a novel therapy for treating gliomas.     

Analysis of immunoarrays using a gold grating-based dual mode surface plasmon-coupled emission (SPCE) sensor chip

We have developed a novel dual mode immunoassay platform that combines the advantages of real-time, label free measurement of surface plasmon resonance (SPR) and the highly directional surface plasmon-coupled emission (SPCE) using a gold grating-based sensor chip. Since only fluorophore-labeled analyte molecules that are close to the metal surface of the sensor chip will couple to the surface plasmon, SPCE detection is highly surface-specific leading to background suppression and increased sensitivity. Theoretical calculations were done to find SPR and SPCE angles for a sensor chip optimized for Alexa Fluor 647. We have confirmed the SPR and SPCE responses on the dual mode sensor chip using Alexa Fluor 647 labeled anti-mouse IgG. Signal fluctuation of the dual mode sensor chip reader was below 1.2% and 0.8% for SPR and SPCE, respectively. The SPR response in this configuration showed a minimum detection level of 1 μg ml(-1), and the SPCE response showed a minimum detection level of 1 ng ml(-1) for the same sample. A range of human IgG concentrations in human serum was also analyzed with the dual mode sensor chip. The SPCE measurement is more sensitive than the SPR real-time measurement, and substantially extends the dynamic range of the assay platform, as well as enabling independent measurements of co-localized analytes on the same sensor chip region of interest. Since this assay platform is capable of measuring more than 1000 spatially encoded regions of interest on a 1 cm(2) sensor chip, it has the potential for high-content analyses of biological samples with both research and clinical applications.     

Synthesis of nanoporous silica aerogel by ambient pressure drying

A crack-free silica aerogel monolith was fabricated from a cheap water glass derived silicic acid solution by adding glycerol, which served as a drying control chemical additive (DCCA). The OH surfaces of the wet gel with glycerol were modified using a TMCS/n-hexane mixture followed by solvent exchange from water to n-hexane. The obtained surface modified wet gel was dried at 75 °C under ambient pressure. The addition of glycerol appears to give the wet gel a more homogeneous microstructure (larger pore size and uniform size distribution) as well as enhanced stiffness. However, glycerol also retards surface modification and solvent exchange. The aerogel synthesized with glycerol added to the silica sol maintained a relatively low bulk density compared with the aerogels aged in a mixed ethanol (EtOH)/TEOS solution. The reproducibility of aerogel production was further improved in the aerogel synthesized with glycerol added to the silica sol and aged in a 70%EtOH/30%TEOS solution.     

The Bis(pentafluoroethyl)phosphinous Acid (C2F5)2POH

Phosphinous acid or phosphane oxide? Both tautomers of (C₂F₅)₂POH (see picture) are found in the neat liquid, whereas only phosphinous acid is present in the gas phase or in solution. The synthesis starting from (C₂F₅)₃PF₂, the thermodynamics of the tautomerization, and the detection and thermodynamics of the cis‐/trans‐P? OH rotamers of the acid are described.

     

Lactic Acid Production from a Whole Slurry of Acid-Pretreated Spent Coffee Grounds by Engineered Saccharomyces cerevisiae

Spent coffee grounds (SCG) generated after coffee extraction are the main byproduct of the coffee industry. Valorization of the SCG has been increasingly focused following considerable attention in coffee consumption. Lactic acid bacteria fermentation is the primary source of generation of lactic acid, a monomer of polylactic acid that has various industrial applications; however, because of the low tolerance of lactic acid bacteria to toxic compounds, it is necessary to apply Saccharomyces cerevisiae to produce lactic acid whose tolerance to toxic compounds is higher. In this study, we evaluated the feasibility of using SCG as substrate for the production of lactic acid by S. cerevisiae strain expressing heterologous lactate dehydrogenase. The fermentation profiles of the engineered yeast showed that lactic acid production was promoted by xylose addition. From simultaneous saccharification and fermentation (SSF) using a whole slurry of acid-pretreated SCG, containing high amounts of hemicellulose fractions, lactic acid (0.11 g) and ethanol (0.10 g) per g SCG were obtained after 24 h of SSF, of which yields were 413% and 221% higher, respectively, than those of washed pretreated SCG. Thus, fermentation of whole slurry SCG by engineered S. cerevisiae is a suitable way of lactic acid production, selectively.

     

Ring opening of 2,5-didehydrothiophene: structures and rearrangements of C4H2S isomers

Electronic structures and rearrangement pathways of several C4H2S isomers are computationally investigated by methods based on coupled cluster theory and density functional theory. Six singlet C4H2S isomers lie within ca. 30 kcal/mol above butatrienethione (6), the apparent global minimum. Ethynylthioketene (7) lies only 2 kcal/mol higher in energy than cumulene 6. Two open-chain isomers, butadiynylthiol (8) and diethynyl sulfide (9), reside ca. 9 and 24 kcal/mol above 6, respectively. Lying 30 kcal/mol above 6, two cyclic singlet isomers, ethynylthiirene (10) and cyclopropenylidenemethanthione (11), are nearly degenerate in energy. Thiophene-2,5-diyl (12) lies substantially higher in energy than 6 (ca. 45 kcal/mol) and is predicted to rearrange preferentially by C-S bond cleavage, leading to thioketene 7, rather than by C-C bond cleavage, leading to diethynyl sulfide (9; retro-Bergman cyclization). Accurate spectroscopic properties of these C4H2S isomers, as well as an understanding of their rearrangement pathways, should facilitate the detection and characterization of these isomers in the laboratory and the interstellar medium.     

Preferentially linear connection of gold nanoparticles in derivatization with phosphorothioate oligonucleotides

A gold nanosphere in water is considered to attain special stability in derivatization like an artificial atom when the octet rule is satisfied by forming four covalent bonds with two 5'-phosphorothioate-modified oligonucleotide molecules. Owing to this, the hybridization of two mutually complementary gold-bound oligonucleotides makes gold nanospheres preferentially connected linearly by duplexes to produce strands like linear artificial molecules. We have then fixated the linear strands of DNA-linked gold nanospheres by reducing Ag(+) ion clusters immobilized around duplexes to show the absorption spectrum of silver-coated artificial-molecular nanorods.     

Density functional study of intradimer proton transfers in hydrated adenine dimer ions, A2(+)(H2O)n (n = 0-2)

Structures of mono- and dihydrated adenine dimers and their cations were calculated using B3LYP density functional theory with the 6-31+G(d,p) basis set, in order to help understand photofragmentation experiments of hydrated adenine dimers from the energetics point of view. Several important pathways leading to the major fragmentation product, protonated adenine ion (AH(+)), thermodynamically at minimum costs were investigated at the ground-state electronic potential surface of hydrated adenine dimer cations. Our calculations suggest that the proton transfer from one adenine moiety to the other in hydrated dimer ions readily occurs with negligible barriers in normal hydration conditions. In asymmetrically hydrated ions, however, the proton transfer to more hydrated adenine moieties is kinetically hindered due to heightened transition-state barriers, while the other way is still barrierless. Such directional preference in proton transfer may be characterized as a unique dimer ion property, stemming from the difference in basicity of the two nitrogen atoms involved in the double hydrogen bond that would be equivalent without hydration. We also found that dimer cleavage requires about 4 times larger energy than evaporation of individual water molecules, so it is likely that most solvent molecules evaporate before the eventual dimer cleavage when available internal energy is limited.