Correlation of abrin-mediated inhibition of protein synthesis and apoptosis.

The plant toxin, abrin, a type-II ribosome inactivating protein, is extremely lethal, the human fatal dose being ~1 µg/kg body weight. Abrin has been classified as an agent for bioterrorism, which is of concern. Conversely, the high toxic property of abrin has been employed in generating immunotoxins, whereas its toxin moiety is conjugated to cell surface marker-specific antibodies for cell-targeted killing. Different cell types exhibit variable levels of sensitivity to abrin toxicity; therefore, adequate knowledge of the molecular mechanism that governs the activity of the protein would be a safeguard. To gain insights into this, two cell lines requiring strikingly different concentrations of abrin for inactivating ribosomes were studied. Employing conjugates of the wild-type and active site mutant of abrin A chain with the ricin B chain, it was found that abrin-induced apoptosis was dependent on inhibition of protein synthesis (PSI) leading to ER-stress in Ovcar-3 cells, but not in KB cells. Abrin was also observed to cause direct DNA damage in KB cells, while in Ovcar-3 cells abrin-induced DNA damage was found to be dependent on caspases. Overall, the study demonstrates that the correlation of abrin-mediated PSI and apoptosis is cell-specific and abrin can induce more than one pathway to cause cell death. © 2018 IUBMB Life, 71(3):357-363, 2019.

Total synthesis and biological evaluation of the cytotoxic resin glycosides ipomoeassin A-F and analogues.

A multitasking C-silylation strategy using the readily available compound 26 as a surrogate for cinnamic acid represents the key design element of a total synthesis of all known members of the ipomoeassin family of resin glyosides. This protecting group maneuver allows the unsaturated acids decorating the glucose subunit of the targets to be attached at an early phase of the synthesis, prevents their participation in the ruthenium-catalyzed ring-closing metathesis (RCM) used to form the macrocyclic ring, and protects them against reduction during the hydrogenation of the resulting cycloalkene over Wilkinson's catalyst. As the C-silyl group can be concomitantly removed with the O-TBS substituent using tris(dimethylamino)sulfonium difluorotrimethylsilicate (TASF) in acetonitrile, no separate protecting group manipulations were necessary in the final stages, thus contributing to a favorable overall "economy of steps". In addition to the naturally occurring ipomoeassins, a small set of synthetic analogues has also been prepared by "diverted total synthesis". The cytotoxicity of these compounds was assayed with two different cancer cell lines. The recorded data confirm previous findings that the acylation- and oxygenation pattern of these amphiphilic glycoconjugates is highly correlated with their biological activity profile. Ipomoeassin F turned out to be the most promising member of the series, showing IC(50) values in the low nanomolar range.

Astasin, a novel cytotoxic carbohydrate-conjugated ergosterol from the colorless euglenoid, Astasia longa.

A novel cytotoxic carbohydrate-conjugated ergosterol (astasin) was found in cells of the colorless euglenoid, Astasia longa. Astasin accounted for about 2.4% of the total lipid of the cells. FAB-MS spectra of astasin showed MH+, 583.3387 (M+, C35H50O7). Astasin was composed of ergosterol (1 eq.), alpha-D-xylopyranose (1 eq.), and oxalic acid (1 eq.). By the acetylation using acetic anhydride and pyridine, oxalic acid was removed from astasin, and three hydroxyl groups of the xylopyranose moiety were acetylated. The two dimensional 13C- and 1H-NMR spectra suggest the oxalic acid was esterified with hydroxyl groups at C-2 and C-3 of the xylopyranose moiety and the hydroxyl group at C-1 of the xylopyranose was glycosidically linked to the hydroxyl group at C-3' of the ergosterol moiety. From the results, the structure of astasin was identified as 2,3-oxalyl-alpha-D-xylopyranosyl (1 --> 3')ergosterol. When cells of HL 60, a human lymphoma, were cultured with astasin, 50% of the cell growth was inhibited at 5.0 micrograms astasin/ml medium, and the cell growth was inhibited completely and 50% of the initial cells was killed at 10.0 micrograms astasin/ml medium.

Effect of a Staphylococcus epidermidis-extracted slime factor on human natural killer cell activity.

A slime factor produced by Staphylococcus epidermidis was a complex glycoconjugate extracted by the phenol extraction method. The potential stimulatory or inhibitory capacity of the phenol-extracted slime (PES) was tested on human natural killer cell cytotoxic activity. Various concentrations of the PES preparation were incubated with the effector cells 30 min before and during the assay period. The PES factor inhibited natural killer cell cytotoxic activity at a concentration of 250 micrograms/ml and at higher concentrations (p < 0.05). The inhibition of natural killer cell cytotoxic activity may probably be related to the complex composition of the slime substance.

[New biologically active substances from protozoa and an evaluation of their action]. / Novye biologicheski aktivnye veshchestva prosteishikh i otsenka ikh deistviia.

Novel biologically active substances were obtained from nonpathogenic for humans and free-living protozoa: Crithidia oncopelti, Trypanosoma lewisi, and Astasia longa. There were studied conditions of biosynthesis, composition and biological activity of the total lipid fraction from the protozoa species, sucrose ethers and fatty acids, the latter were isolated from A. longa--(astazilide preparation), reserve beta-1,3 glucan from A. longa (astazian preparation), and fraction of surface glycophospholipids and peptides from Crithidia oncopelti (GLP preparation). Two of three preparations studied (astazilide and GLP) are the complexes of natural substances with certain composition. Conditions of the protozoa cultivation were developed to provide standardization of the complex composition. Division of the complexes into separate components decreased or abolished the biological activity. It was established that the substances studied modify biological reactions, that is exert a systemic effect. They may be used for inhibiting the growth of experimental tumors and preventing the metastatic spreading in tumor-carrier animals. The substances obtained belong to the group of nonspecific stimulators for cellular immunity which can be used in medicine, veterinary medicine, food industry, pharmacologic industry and cosmetology.

A glycoconjugate from corms of saffron plant (Crocus sativus L.) inhibits root growth and affects in vitro cell viability.

A glycoconjugate has been characterized from saffron corms (Crocus sativus L.) that inhibits the growth of roots of Nicotiana tabacum and Arabidopsis thaliana, at concentrations ranging from 1-100 micrograms m-3. Roots of seedlings grown in the presence 0.1 microgram m-3 glycoconjugate showed bulging of epidermal cells, whereas at 10 micrograms m-3, roots were completely devoid of hairs. At 100 micrograms m-3 glycoconjugate the cell walls of the root vascular tissues were thicker and, overall, the vascular tissue was enlarged. In addition, this glycan is cytotoxic to isolated tobacco cells and protoplasts, with 50% cell death induced by 0.5 and 2 micrograms m-3 glycoconjugate, respectively. Morphological and biochemical changes induced by the exposure to the glycoconjugate included cell size decrease, loss of regular cell shape, cytoplasm collapse, and release of intracellular proteins. This molecule at low concentrations (0.1 microgram m-3) mimics the effects of Yariv phenylglycosides and of mutant Arabidopsis which present defective or missing arabinogalactan-proteins (AGPs) in roots, indicating the glycoconjugate might interact with cell surface AGPs.