Multiple Administrations of 64Cu-ATSM as a Novel Therapeutic Option for Glioblastoma: a Translational Study Using Mice with Xenografts

Glioblastoma is the most aggressive malignant brain tumor in humans and is difficult to cure using current treatment options. Hypoxic regions are frequently found in glioblastoma, and increased levels of hypoxia are associated with poor clinical outcomes of glioblastoma patients. Hypoxia plays important roles in the progression and recurrence of glioblastoma because of drug delivery deficiencies and induction of hypoxia-inducible factor-1α in tumor cells, which lead to poor prognosis. We focused on a promising hypoxia-targeted internal radiotherapy agent, ⁶⁴Cu-diacetyl-bis (N⁴-methylthiosemicarbazone) (⁶⁴Cu-ATSM), to address the need for additional treatment for glioblastoma. This compound can target the overreduced state under hypoxic conditions within tumors. Clinical positron emission tomography studies using radiolabeled Cu-ATSM have shown that Cu-ATSM accumulates in glioblastoma and its uptake is associated with high hypoxia-inducible factor-1α expression. To evaluate the therapeutic potential of this agent for glioblastoma, we examined the efficacy of ⁶⁴Cu-ATSM in mice bearing U87MG glioblastoma tumors. Administration of single dosage (18.5, 37, 74, 111, and 148 MBq) and multiple dosages (37 MBq × 4) of ⁶⁴Cu-ATSM was investigated. Single administration of ⁶⁴Cu-ATSM in high-dose groups dose-dependently inhibited tumor growth and prolonged survival, with slight and reverse signs of adverse events. Multiple dosages of ⁶⁴Cu-ATSM remarkably inhibited tumor growth and prolonged survival. By splitting the dose of ⁶⁴Cu-ATSM, no adverse effects were observed. Our findings indicate that multiple administrations of ⁶⁴Cu-ATSM have effective antitumor effects in glioblastoma without side effects, indicating its potential for treating this fatal disease.     

Isolation of an Arabidopsis thaliana mutant in which accumulation of cucumber mosaic virus coat protein is delayed

Cucumber mosaic virus (CMV) is known to systemically infect Arabidopsis thaliana ecotype Columbia plants. In order to identify the host factors involved in the multiplication of CMV, we isolated an A. thaliana mutant in which the accumulation of the coat protein (CP) of CMV in upper uninoculated leaves was delayed. Genetic analyses suggested that the phenotype of delayed accumulation of CMV CP in the mutant plants was caused by a single, nuclear and recessive mutation designated cum1-1, which was located on chromosome IV. The cum1-1 mutation did not affect the multiplication of tobacco mosaic virus, turnip crinkle virus or turnip yellow mosaic virus, which belong to different taxonomic groups from CMV. Accumulation of CMV CP in the inoculated leaves of cum1-1 plants was also delayed either when CMV virion or CMV virion RNA was inoculated. On the other hand, when cum1-1 and the wild-type Col-0 protoplasts were inoculated with CMV virion RNA by electroporation, the accumulations of CMV-related RNAs and the coat protein were similar. These results suggest that the cum1-1 mutation did not affect the uncoating of CMV virion and subsequent replication in an initially infected cell but affected the spreading of CMV within an infected leaf, possibly the cell-to-cell movement of CMV in a virus-specific manner.     

Effect of glycine betaine on the sucrose catabolism of an l-lysine producing mutant of Brevibacterium lactofermentum

Summary Effect of exogenous betaine on the growth of an l-lysine-producing mutant of Brevibacterium lactofermentum was examined in a medium containing different carbon sources such as glucose, fructose, or sucrose. The growth rate decreased significantly with a rise in temperature when sucrose was the carbon source. Both the specific sucrose consumption rate and the invertase activity of the mutant decreased with the culture period when the cultivation temperature was 35°C. The addition of betaine restored both growth and invertase activity on medium containing sucrose as the carbon source at 35°C. Betaine protected the invertase activity against the inactivating effects of high temperature in vitro. Furthermore, the addition of exogenous invertase into production medium at 35°C restored the growth rate to that at 32°C. These results indicated that growth decreased on medium containing sucrose at 35°C due to a decrease in invertase activity, and that addition of betaine was an effective way to enhance growth on this medium at a higher temperature. Offprint requests to: Y. Kawahara     

Biosynthesis of auxin-induced ethylene in mung bean hypocotyls

The pathway of ethylene biosynthesis in auxin-treated mung beanhypocotyls was investigated by comparing the specific radioactivitiesof ethylene produced and S-adenosylmethionine (SAM) in the tissuefollowing the administration of 3,4-¹⁴C-methionine, and by analyzingthe methionine metabolites. When the rate of auxin-induced ethyleneproduction was low due to a low concentration of auxin, thespecific radioactivity of ethylene released was always higherthan that of SAM in the tissue. When the tissue was treatedwith auxin, the tissue produced and accumulated a methioninemetabolite which was converted into ethylene more efficientlythan methionine. The metabolite was identified as 1-aminocyclopropane-l-carboxylicacid (ACC) by means of paper and thin-layer chromatography,high voltage paper electrophoresis and co-crystallization. ACCformation was neither inhibited by low oxygen nor by the inhibitoryprotein of ethylene synthesis, but inhibited by aminoethoxyvinylglycine(AVG). ACC application to the tissue greatly reduced incorporationof 3,4-¹⁴C-methionine into ethylene. The control tissue thatwas not treated with auxin also converted ACC into ethyleneindicating that the enzyme which converts ACC into ethyleneis already present in the tissue and that auxin induced productionof the enzymatic system responsible for the conversion of methionineinto ACC. Ethylene synthesis from ACC was not inhibited by AVG,abscisic acid, cycloheximide or actinomycin D, but inhibitedby low oxygen and the inhibitory protein. (Received November 21, 1979; )     

Detection of bacterial DNA from central venous catheter removed from patients by next generation sequencing: a preliminary clinical study

Background

Catheter-related infection (CRI) is one of the serious challenges in clinical practice. This preliminary clinical study aimed to examine whether next-generation sequencing (NGS) targeting 16S rDNA, which was PCR-amplified directly from the tip of a central venous catheter (CVC), can be used to identify causative pathogens in CRI, compared to the culture method.

Methods

Hospitalized patients, from whom a CVC had just been removed, were prospectively enrolled and divided into the CRI-suspected and routine removal groups. DNA was extracted from the sonication fluid of CVC specimens derived from patients. For analysis of bacterial composition by NGS, the V3–V4 fragments of bacterial 16S rDNA were PCR-amplified, followed by index PCR and paired-end sequencing on an Illumina MiSeq device. Conventional culture methods were also performed in the CRI-suspected group.

Results

Of CVCs collected from the 156 enrolled patients (114 men; mean age 65.6 years), a total of 14 specimens [nine out of 31 patients suspected with CRI and five out of 125 patients without infection symptoms (routine removal group)] were PCR-positive. In five patients with definite CRI, Staphylococcus was the most frequently detected genus by NGS (4/5 specimens), although no pathogens were detected by NGS in the one remaining specimen. The genera identified by NGS were consistent with those from conventional culture tests. There was high agreement between NGS and the culture method in the CRI-suspected group, with sensitivity and specificity values of 80.0% and 76.9%, respectively; meanwhile, the false-positive rate of NGS was as low as 4.0% in the routine removal group. Moreover, several genera, besides the genus identified by culture test, were detected in each patient with definite CRI and surgical site infection (SSI). Additionally, in one patient with SSI, Enterococcaceae were detected not only by NGS but also by abdominal abscess drainage culture.

Conclusions

NGS targeting 16S rDNA was able to analyze the bacterial composition of CVC specimens and detect causative pathogens in patients with CRI and was therefore suggested as a promising diagnostic tool for CRI.

     

Stabilization of Photosystem II (O(2) Evolution) of Spinach Chloroplasts by Radiation-induced Immobilization

Spinach chloroplasts were immobilized with vinyl monomers by radiation-induced polymerization at low temperature and stored in buffer containing bovine serum albumin. The lifetime of O₂ evolution activity in photosystem II was prolonged remarkably in immobilized chloroplasts. Thermostability of immobilized chloroplasts stored in buffer containing bovine serum albumin was far better than that of immobilized chloroplasts in pure buffer and that of intact chloroplasts. When immobilized chloroplasts were stored in buffer including polyethylene glycol, the lifetime of O₂ evolution activity was longer than for those stored in buffer containing bovine serum albumin.     

Cattle cDNA clone encoding a new allele of the MHC class II DQA1 gene

The nucleotide sequence data reported in this paper have been submitted to the DDBJ, EMBL, and GenBank nucleotide sequence databases and have been assigned the accession number D50454     

PHOTOSYNTHETIC PIGMENT COMPOSITION IN THE PRIMITIVE GREEN ALGA MESOSTIGMA VIRIDE (PRASINOPHYCEAE): PHYLOGENETIC AND EVOLUTIONARY IMPLICATIONS1

The photosynthetic pigment composition of Mesostigma viride Lauterborn, a primitive green alga, was determined. This alga contained chl a and b, lycopene, lutein, siphonaxanthin, γ‐carotene, β‐carotene, antheraxanthin, violaxanthin, neoxanthin, and two novel carotenoid fatty acid esters, siphonaxanthin C12:0 ester and siphonaxanthin C14:0 ester. The esters were saturated, whereas all previously identified siphonaxanthin and loroxanthin esters have been mono‐unsaturated (trans‐Δ2). Neoxanthin was the all‐trans form. This is the first such case detected in the chloroplasts of green plants. The 9′‐cis form of neoxanthin is believed to be universally present in the chloroplasts of green plants (Streptophyta and Chlorophyta) and is a precursor of abscisic acid. However, the 9′‐cis form was not found in M. viride. Based on these results, we discuss the phylogenetic implications and early evolution of the antenna pigment system in green plants.     

CHARACTERIZATION OF TWO UNIQUE CAROTENOID FATTY ACID ESTERS FROM PTEROSPERMA CRISTATUM (PRASINOPHYCEAE,CHLOROPHYTA)1

Carotenoid composition and spectroscopic characteristics were analyzed for Pterosperma cristatum Schiller, one of the most primitive known members of green algae. This alga contained a substantial amount of carotenoid esters, siphonaxanthin C14:1 trans‐Δ2 ester and 6′‐OH siphonaxanthin C14:1 trans‐Δ2 ester, but lacked lutein. This is the first report of carotenoid C14:1 trans‐Δ2 esters from phototrophic organisms. In vivo absorption spectra and excitation spectra of the cells revealed that these carotenoids absorbed blue‐green light and could transfer energy to chl a. These carotenoids were concluded to function as antenna pigments in P. cristatum.