S. I. Shram

Disaccharide pyrimidine nucleosides and their derivatives: a novel group of cell-penetrating inhibitors of poly(ADP-ribose) polymerase 1.

Nearly 30 synthetic nucleosides were tested with human recombinant poly(ADP-ribose) polymerase 1 as potential inhibitors of this enzyme. The most active compounds were some disaccharide analogues of thymidine: 3′-O-β-D-ribofuranosyl-5-iodo-dUrd (2d; IC50 = 45 μM), 3′-O-β-D-ribofuranosyl-2′-deoxythymidine (2e; IC50 = 38 μM), and 3′-O-β-D-ribofuranosyl-2′-deoxythymidine oxidized (4; IC50 = 25 μM). These compounds also reduced H2O2-induced synthesis of poly(ADP-ribose) in cultured human ovarian carcinoma (SKOV-3) cells in a dose-dependent manner. Furthermore, compounds 2d or 2e until a concentration of 1 mM did not affect growth of SKOV-3 cells, whereas dialdehyde compound 4, as well as thymidine, exhibited a significant cytotoxicity.

Structural-functional study of glycine-and-proline-containing peptides (Glyprolines) as potential neuroprotectors

A synthetic scheme for preparation of (Gly-Pro)n, (Pro-Gly)n (n = 2, 3), and (Pro-Gly-Pro)n (n = 1, 2) peptides was elaborated. The effect of the synthesized peptides and the Gly-Pro and Pro-Gly dipeptides on survival of cultured cells of PC12 rat pheochromocytoma was studied under the conditions of oxidative stress induced by brief incubation of the cells with hydrogen peroxide. Peptides of the general formula (Gly-Pro)n and the Pro-Gly-Pro peptide at a concentration of 0.2–100 μM were shown to decrease the number of damaged cells. The Gly-Pro peptide was the most active and decreased the number of damaged cells by 49% on average at a concentration of 100 μM.

Formation of spatial memory in rats with ischemic lesions to the prefrontal cortex; effects of a synthetic analog of ACTH(4-7).

Photochemically induced thrombosis of blood vessels in the prefrontal cortex in rats was shown to lead to ischemic infarcts in the lesion zone. Bilateral ischemic lesioning of the prefrontal cortex degraded measures of spatial memory when animals were tested in a Morris water maze with an invisible platform 20–24 days after surgery. Chronic intranasal administration of the peptide Met-Glu-His-Phe-Pro-Gly-Pro (Semax), a synthetic analog of ACTH(4-7), at a dose of 250 μg/kg/day during the first six days after photothrombosis, led to recovery of the animals’ learning ability. The long-term antiamnestic action of the peptide observed here may result from its neuroprotective activity and its ability to stimulate the synthesis of neurotrophic factors.

Isolation and Characterization of High Affinity Aptamers Against DNA Polymerase Iota

Human DNA-polymerase iota (Pol ι) is an extremely error-prone enzyme and the fidelity depends on the sequence context of the template. Using the in vitro systematic evolution of ligands by exponential enrichment (SELEX) procedure, we obtained an oligoribonucleotide with a high affinity to human Pol ι, named aptamer IKL5. We determined its dissociation constant with homogenous preparation of Pol ι and predicted its putative secondary structure. The aptamer IKL5 specifically inhibits DNA-polymerase activity of the purified enzyme Pol ι, but did not inhibit the DNA-polymerase activities of human DNA polymerases beta and kappa. IKL5 suppressed the error-prone DNA-polymerase activity of Pol ι also in cellular extracts of the tumor cell line SKOV-3. The aptamer IKL5 is useful for studies of the biological role of Pol ι and as a potential drug to suppress the increase of the activity of this enzyme in malignant cells.

Transcriptomic response of rat hippocampus and spleen cells to single and chronic administration of the peptide selank.

A new direction in designing new drugs able to effectively reduce anxiety without having side effects is the use of endogenous regulatory peptides. Research� ers of the Institute of Molecular Genetics, Russian Academy of Sciences, and the Zakusov Research Institute of Pharmacology, Russian Academy of Med� ical Sciences, have created the preparation selank, the effective substance of which is a synthetic peptide, an analogue of the short fragment Thr–Lys–Pro–Arg of the heavy chain of human immunoglobulin G, elon� gated at the C terminus with the tripeptide Pro–Gly– Pro. It was shown that selank has a stable nootropic and anxiolytic effects, facilitate brain cell survival in hypoxia, and exhibits an antiviral effect [3, 10].

Effect of Mn(II) on error-prone DNA polymerase iota activity in extracts from human normal and tumor cells

DNA polymerase iota (Pol ι) which has some peculiar features and is characterized by an extremely error prone DNA synthesis, belongs to the group of enzymes preferentially activated by Mn2+ instead of Mg2+. In this work, using misGvA method (misincorporation of “G” versus “A”, method of Gening) we studied the effect of Mn2+ on DNA synthesis in cell extracts from a—normal human and murine tissues, b—human tumor (uveal melanoma), and c—cultured human tumor cell lines SKOV-3 and HL-60. Each group displayed characteristic features of Mn-dependent DNA synthesis. Changes in Mn-dependent DNA synthesis caused by malignant transformation of normal tissues are described. It was also shown that the error prone DNA synthesis catalyzed by Pol ι in extracts of all cell types was efficiently suppressed by an RNA aptamer (IKL5) against Pol ι obtained by us earlier. The results obtained suggest that IKL5 might be used to suppress enhanced activity of Pol ι in tumor cells.

Expression changes caused by the peptide semax in the intracellular signal pathway genes in rat hippocamp.

334 ACTH (4–7) is the shortest N-terminal fragment of adrenocorticotropic hormone able to influence animal behavior [1]. A number of its synthetic analogues have been developed and tested [2] for designing nootropic drugs derived from the ACTH (4–7) fragment. The teams from the Institute of Molecular Genetics and Moscow State University created the peptide Met–Glu– His–Phe–Pro–Gly–Pro, which was named semax. It has been shown that semax enhances the survival of brain cells during hypoxia and increases the selective attention and memory trace consolidation [3]. Semaxbased drugs are used for treating craniocerebral injuries, strokes, and optic nerve diseases, as well as during post-reanimation treatment [4, 5].

Enzyme self-inactivation is a main limitation of the preparation of eicosanoids. Enzymatic synthesis of thromboxane and 12(S)-hydroxytetraenoic acid.

Synthesis of prostanoids is accompanied by various processes reducing the product yield. These processes are also known to affect syntheses of thromboxane (TX) and 12(S)-hydroxy-5(Z),8(Z),10(E),14(Z)-eicosatetraenoic acid (12-HETE). Partially purified preparations of TX synthase and prostaglandin (PG) synthase were used to optimize TX synthesis with respect to concentrations of the enzymes and eicosapolyenoic acid (EPA). Conditions for the maximum product yield and the minimum consumption of enzymes were determined. Consumption of the TX synthase was large owing to its inactivation during the reaction and the nonenzymatic destruction of the intermediate product PG-endoperoxide. Separate addition of PG and TX synthases increased the product yield by preventing EPA sorption on ballast proteins. Microsomal 12-lipoxygenase (12-LO) was also shown to be inactivated during the reaction, and this process was the major limitation of 12-HETE synthesis. Lipoxygenase reaction in the presence of some reducing agents led to a considerable increase of the 12-HETE yield, supposedly by preventing further oxidation of the 12-LO reaction product 12-hydroperoxy derivative of eicosatetraenoic acid. The possibility of using human blood platelet microsomes for preparation of some derivatives of EPAs is discussed.

Enzymatic synthesis of tritium labelled eicosanoids

Methods for the synthesis of tritium labelled prostaglandins, thromboxanes, and hydroxy-derivatives of eicosapolyenoic acids using mono-, two-, and three-enzyme systems have been developed. Optimization of the eicosanoid syntheses with respect to the concentrations of enzymes and eicosapolyenoic acid, time, and temperature of incubation are reported. It is demonstrated that a number of compounds can be produced by enzymatic modifications of prostaglandins and thromboxane Procedures for the preparation of twenty tritium labelled compounds are described.

The penetration of 5-oxo-Pro-Arg-Pro into the brain and the major metabolic pathways of this peptide in the rat brain and blood at the intranasal and intravenous administration

It was shown that the neuroactive peptide 5-oxo-Pro-Arg-Pro (5-oxo-PRP) is detected in the brain in the time interval of 5–120 min after it was intravenously or intranasally administered to rats; the maximum concentration of labeled tripeptide in these modes of administration was observed after 30 and 10 min, respectively. A significant difference in the concentrations of 5-oxo-PRP in the blood and brain (the latter was 50 times lower) during intravenous administration indicates a relatively low permeability of the peptide across the blood–brain barrier. Pharmacokinetic data analysis showed that, when administered intranasally, approximately 45% of the total number of 5-oxo-PRP detectable in the brain in the entire period of study enters via transport from the nasal cavity, and the rest of the peptide enters through the blood–brain barrier from the blood stream. It was found that 5-oxo-PRP in rats is rapidly metabolized forming proteolytic products, mainly amino acids, and degradation products, presumably oxidized these amino acids.