Early enteral nutrition with arginine compensates for negative nitrogen balance in patients undergoing curative total gastrectomy.

The effects of early enteral arginine-rich nutrition (EAN) were analyzed among patients undergoing curative-intent total gastrectomy for gastric cancer. There were 19 patients in this prospective study, all randomly assigned to either a parenteral nutrition (PN) group or an EAN group for the first seven days after surgery. The EAN group received 1.8-fold greater arginine (10.1 g/day) compared with the PN group, which was administered through an enteral tube inserted into the jejunal loop. Both groups were provided almost identical amounts of total amino acids (54 g/day), and the total energy was set at 65% of the total requirement (25 kcal/kg/day). No significant differences were observed between the two groups in postoperative complications, length of hospital stay, oral intake, nutritional status, or body weight. The serum arginine profile was similar in the two groups, as it decreased significantly on postoperative day (POD) 1, and gradually returned to preoperative levels by POD 7. The nitrogen balance remained negative until POD 7 in the PN group, but turned neutral at POD 7 in the EAN group. While we could not confirm body weight loss improvement, these results suggested that early arginine-rich enteral nutrition could improve the nitrogen balance after total gastrectomy. J. Med. Invest. 70 : 325-333, August, 2023.

Effects of methylation of arginine residue 83 on the enzymatic activity of human arsenic (+3 oxidation state) methyltransferase.

Arsenic (+3 oxidation state) methyltransferase is an enzyme responsible for arsenic methylation, and it requires S-adenosyl-methionine (SAM) as a coenzyme. We here generated two mutants to clarify the role of the highly conserved 83rd arginine residue (Arg83) in Motif I, the SAM-binding domain, of human AS3MT. When the AS3MT activity was compared between the mutants and the wild type (WT) recombinant protein, little activity was detected in the glycine mutant (Arg83Gly) or lysine mutant (Arg83Lys). When we examined the ability of transfected HEK293 cells exposed to arsenite to methylate arsenic, the methylation ability was significantly reduced in Arg83Gly compared to the WT, but was not significantly different between Arg83Lys and WT. Western blot analysis of the recombinant WT and Arg83Gly with an antibody that recognizes methylated Arg showed that an Arg residue in the WT was mono- and di-methylated, but not in Arg83Gly. Furthermore, a peptide containing dimethylated Arg83 was detected by MALDI-TOF/MS of the WT digested with chymotrypsin. These results indicate that AS3MT maintains its enzymatic activity through the methyl modification of Arg83.

Sulfated vizantin causes detachment of biofilms composed mainly of the genus Streptococcus without affecting bacterial growth and viability.

BACKGROUND: Sulfated vizantin, a recently developed immunostimulant, has also been found to exert antibiofilm properties. It acts not as a bactericide, but as a detachment-promoting agent by reducing the biofilm structural stability. This study aimed to investigate the mechanism underlying this activity and its species specificity using two distinct ex vivo oral biofilm models derived from human saliva. RESULTS: The biofilm, composed mainly of the genus Streptococcus and containing 50 µM of sulfated vizantin, detached significantly from its basal surface with rotation at 500 rpm for only 15 s, even when 0.2% sucrose was supplied. Expression analyses for genes associated with biofilm formation and bacterial adhesion following identification of the Streptococcus species, revealed that a variety of Streptococcus species in a cariogenic biofilm showed downregulation of genes encoding glucosyltransferases involved in the biosynthesis of water-soluble glucan. The expression of some genes encoding surface proteins was also downregulated. Of the two quorum sensing systems involved in the genus Streptococcus, the expression of luxS in three species, Streptococcus oralis, Streptococcus gordonii, and Streptococcus mutans, was significantly downregulated in the presence of 50 µM sulfated vizantin. Biofilm detachment may be facilitated by the reduced structural stability due to these modulations. As a non-specific reaction, 50 µM sulfated vizantin decreased cell surface hydrophobicity by binding to the cell surface, resulting in reduced bacterial adherence. CONCLUSION: Sulfated vizantin may be a candidate for a new antibiofilm strategy targeting the biofilm matrix while preserving the resident microflora.

Arsenite suppresses the transcriptional activity of EVI1 through the binding to CCHC-type Zn finger domain.

Transcription factor EVI1 is essential for normal hematopoiesis in embryos but is aberrantly elevated in bone marrow cells of myelodysplastic syndrome (MDS) patients. EVI1 and its downstream GATA-2 appear to be a possible therapeutic target of MDS. Here we found that treatment of EVI1-expressing K562 cells with arsenite (As(III)) reduced the mRNA and protein levels of EVI1 and GATA-2. A gel shift assay using the nuclear extract of K562 cells showed that As(III) suppressed the DNA-binding activity of EVI1. The DNA-binding activity of the recombinant EVI1 protein was also suppressed by As(III) but was recovered by excess amounts of dithiothreitol, suggesting the involvement of cysteine residues of EVI1. Since the 7th Zn finger domain of EVI1, having a motif of CCHC, is known to be involved in DNA-binding, the synthetic peptide of 7th Zn finger domain was reacted with As(III) and subjected to MALDI-TOF-MS analysis. The results showed that As(III) binds to this peptide via three cysteine residues. As(III)-induced reduction of the DNA-binding activity of the recombinant EVI1 was abolished by the mutations of each of three cysteine residues to alanine in the 7th Zn finger domain. These results demonstrate that As(III) causes the down-regulation of EVI1 and GATA-2 by inhibiting the transcriptional activity of EVI1 through the binding to the cysteine residues of CCHC-type Zn finger domain.

Dinorcassane Diterpenoid from Boesenbergia rotunda Rhizomes Collected in Lower Myanmar.

A new dinorcassane diterpenoid, seikphoochinal A (1), and four known compounds, pinostrobin (2), 4',7-dimethylkaempferol (3), and galanals A (4) and B (5), were isolated from the chloroform-soluble crude extract of wild type Boesenbergia rotunda rhizomes collected in Lower Myanmar. The chemical structures of these compounds were identified, using a combination of spectroscopic methods. The presence of the diterpenoids 1, 4, and 5 demonstrated the structural diversity of wild type B. rotunda. Among the isolates, compounds 4 and 5 exhibited significant antiproliferative activities against a small panel of human cancer cell lines, including lung (LK-2, A549), stomach (ECC4), breast (MCF7), cervix (HeLa), and prostate (DU145).

Tetrahydrofuran lignans: Melanogenesis inhibitors from Premna integrifolia wood collected in Myanmar.

Two new tetrahydrofuran lignans, taungtangyiols A (1) and B (2), and eight known furofuran lignans (3-10), were isolated from the chloroform extract of Premna integrifolia wood collected in Myanmar. Their structures were elucidated by extensive spectroscopic techniques. The X-ray crystal structure of 1 clearly indicated its relative configuration. Taungtangyiols A (1) and B (2) inhibited the deposition of melanin in B16F10 mouse melanoma cells, with IC50 values of 50.7 and 40.9 µM, respectively, without notable cytotoxicity. An SAR study demonstrated that the furofuran and dioxymethylene moieties of the lignans play a vital role in inhibiting melanogenesis.

Labdane diterpenoids from Curcuma amada rhizomes collected in Myanmar and their antiproliferative activities.

Four new labdane diterpenoids, 12ß-hydroxy-15-norlabda-8(17),13(14)-dien-16-oic acid (1), (E)-15-ethoxy-15-methoxylabda-8(17),12-dien-16-al (2), (E)-15α-ethoxy-14α-hydroxylabda-8(17),12-dien-16-olide (3), and 15-ethoxy-12ß-hydroxylabda-8(17),13(14)-dien-16,15-olide (4) were isolated from the methanol extract of Curcuma amada rhizomes collected in Myanmar, together with 13 known analogs. Their structures were elucidated by extensive spectroscopic techniques. All of the isolates were evaluated for their antiproliferative activities against a small panel of five different human cancer cell lines (A549, human lung cancer; HeLa, human cervical cancer; MCF7, human breast cancer; PANC-1 and PSN-1, human pancreatic cancer). Among the isolates, compounds 2-4, 7, 8, 12, and 17 showed mild antiproliferative activities with IC50 values ranging from 19.7 to 96.1µM. (E)-14-Hydroxy-15-norlabda-8(17),12-dien-16-al (11) exhibited strong antiproliferative activities selectively against HeLa, PANC-1, and PSN-1 cells, with IC50 values of 5.88, 1.00, and 3.98µM, respectively. These potencies were comparable to those of the positive control, 5-fluorouracil.

Synthesis and evaluation of c-di-4'-thioAMP as an artificial ligand for c-di-AMP riboswitch.

Cyclic-di-adenosine monophosphate (c-di-AMP) is a bacterial second messenger that binds to an RNA receptor called riboswitch and regulates its downstream genes involving cell wall metabolism, ion transport, and spore germination. Therefore, the c-di-AMP riboswitch can be a novel target of antibiotics. In this study, we synthesized c-di-4'-thioAMP (1), which possesses a sulfur atom instead of an oxygen atom in the furanose ring, as a candidate of a bioisoster for natural c-di-AMP. The resulting 1 bound to the c-di-AMP riboswitch with a micromolar affinity (34.8µM), and the phosphodiesterase resistance of 1 was >12-times higher than that of c-di-AMP. Thus, 1 can be considered to be a stable ligand against a c-di-AMP riboswitch.

Two new cyclopentenones and a new furanone from Baeckea frutescens and their cytotoxicities.

Two new cyclopentenones, frutescencenones A (1) and B (2), and a new furanone derivative, frutescencenone C (3), together with two known cyclopentenones (4 and 5), were isolated from the leaves of Baeckea frutescens. Their structures were deduced by comprehensive spectroscopic analyses, including 1D and 2D NMR, and HREIMS data. Frutescencenone A (1) showed moderate growth inhibitory activity against human lung A549, pancreatic PSN-1, and breast MDA-MB-231 cancer cell lines, with IC50 values of 36.3µM, 38.2µM, and 29.3µM, respectively. In contrast, frutescencenone C (3) showed selective cytotoxic activity against PSN-1, with an IC50 value of 20.1µM.

Two New Diterpenoids from Salvia przewarskii.

One abietane-type and one dinoricetexane-type diterpenoid, salviskin A (1) and salviskin B (2), respectively, together with fourteen known diterpenoids, were isolated from Salvia przewarskii. Structural elucidation of these compounds was performed by spectroscopic methods including 2D NMR. The compounds isolated were evaluated for their cytotoxicity against HeLa and HL-60 cells.

New cytotoxic phloroglucinols, baeckenones D-F, from the leaves of Indonesian Baeckea frutescens.

Three new phloroglucinols, baeckenones D-F (1-3), were obtained from the leaves of Indonesian Baeckea frutescens, along with the known unusual endoperoxide, phloroglucinol (4). The structures of the isolated compounds were elucidated by 1D and 2D NMR and HREIMS spectra. Furthermore, the stereochemistry of baeckenone D (1) was established by an X-ray diffraction analysis. Among the isolated compounds 1-4, baeckenone F (3) showed moderate cytotoxic activities against human pancreatic (PSN-1), lung (A549), and breast (MDA-MB-231) cancer cell lines, with IC50 values of 33.3 µM, 34 µM, and 39.3 µM, respectively.

Design of a bioreductively-activated fluorescent pH probe for tumor hypoxia imaging.

We have designed and evaluated UTX-12 as a novel fluorescent pH probe for tumor hypoxia imaging. UTX-12 consists of a p-nitro benzyl moiety, which is a latent hypoxia-selective leaving group activated by nitro reduction, directly linked to SNARF. Although UTX-12 itself is colorless and non-fluorescent in aqueous solution, nitro reduction triggers the release of SNARF which has well-characterized long wavelength absorption and fluorescence that is sensitive to pH. The resultant SNARF, released intracellularly by enzymatic reduction of UTX-12, allows measurement of pH by pH-dependent dual emission shifts. UTX-12 showed clear differences in fluorescence behavior between hypoxic and aerobic conditions in liver microsomes and inside V79 cells. These data are confirmation that UTX-12 is biologically reduced inside tumor cells and the released SNARF should monitor intracellular pH of tumor cells selectively with reduced background signal.

Synthesis and biological activity of 1-methyl-tryptophan-tirapazamine hybrids as hypoxia-targeting indoleamine 2,3-dioxygenase inhibitors.

We have designed and synthesized new hypoxic-neoplastic cells-targeted indoleamine 2,3-dioxygenase (IDO) inhibitors. 1-Methyl-tryptophan (1MT)-tirapazamine (TPZ, 3-amino-1,2,4-benzotriazine 1,4-dioxide) hybrid inhibitors including 1 (TX-2236), 2 (TX-2235), 3 (TX-2228), and 4 (TX-2234) were prepared. All of these compounds were uncompetitive IDO inhibitors. TPZ-monoxide hybrids 1 and 3 showed higher IDO inhibitory activities than TPZ hybrids 2 and 4. Among these hybrids, hybrid 1 was the most potent IDO inhibitor. TPZ hybrids 2 and 4 showed stronger hypoxia-selective cytotoxicity than TPZ to EMT6/KU cells. These data suggest that TPZ hybrids 2 and 4 may act through their dual biological functions: first, they function as hypoxic cytotoxins in hypoxic cells, and then are metabolized to their TPZ-monoxide (3-amino-1,2,4-benzotriazine 1-oxide) hybrids, which function as IDO inhibitors.

TX-2152: a conformationally rigid and electron-rich diyne analogue of FTY720 with in vivo antiangiogenic activity.

We designed FTY720 analogues with conformationally rigid and electron-rich acetylenic chains as antiangiogenic agents (the monoyne 1: TX-2148, the diyne 2: TX-2152, the triyne 3: TX-2256). Molecular orbital (MO) calculations of our designed acetylenic analogues and FTY720 showed that the localization of the lowest unoccupied MO and the highest occupied MO increased from phenyl ring to acetylenic chain compared with that of FTY720. These acetylenic analogues were synthesized from p-hydroxyphenylethanol as a starting material. The construction of the acetylenic chain was carried out by an iterative strategy using a Sonogashira cross-coupling reaction and desilylative bromination in two steps. The corresponding overall yields of the monoyne 1, the diyne 2, and the triyne 3 were 27% (11 steps), 13% (13 steps), and 10% (15 steps). The in vivo antiangiogenic activities of these acetylenic analogues and FTY720 were evaluated by the chick embryo chorioallantoic membrane (CAM) assay and compared to the activities of the known antiangiogenic agent TNP-470. The diyne 2 showed more potent antiangiogenic activity (90% inhibition) than FTY720 (77% inhibition) and other acetylenic analogues (the monoyne 1: 42% inhibition, the triyne 3: 60% inhibition), and TNP-470 (82% inhibition) at a dose of 10 microg/CAM, without showing toxicity. The diyne 2 also had potent inhibitory activity at a dose of 5 and 2.5 microg/CAM. These results indicate that the flexibility of C8 alkyl chain of FTY720 is not required for its antiangiogenic activity. We suggest that the diyne 2 (TX-2152) may be a promising candidate as an antiangiogenic agent for antineoplastic drug discovery.