— Peptides are low-molecular-weight substances involved in numerous physiological processes, such as growth and development of the organism, stress, regulation of the emotional state, sexual behavior, and immune responses. Most peptides act as ligands for proteins, the interaction with which leads to highly selective effects. Such properties, as well as a low toxicity, allow us to consider peptides as potential effective drugs. The preparation of peptide drugs became possible at the beginning of the 20th century after the development of a method of their selective synthesis. Despite the success of the synthesis of the first peptide drugs, many issues associated with an increase in the stability, bioavailability, half-life, and the ability to move through the cell membranes remain unresolved. The review considers the historical path of development of the synthesis and production of peptides, and discusses modern approaches to the creation of peptide drugs and their application in biopharmaceutics, including in the development of original peptide drugs in Russia.

Прионный белок является одной из мембранных мишеней бета-амилоида, основного нейротоксического агента при болезни Альцгеймера. Нами был осуществлен выбор и синтез фрагментов прионного белка 17–33, 23–33, 95–110 и 101–115, ответственных за связывание с бета-амилоидом. Было изучено влияние иммунизации пептидами на развитие признаков болезни у животных с экспериментально индуцированной формой болезни Альцгеймера. Показано, что иммунизация свободным пептидом 17–33 и белковыми конъюгатами пептидов 23–33 и 101–115 приводит к восстановлению состояния пространственной памяти животных. Также показано, что иммунизация пептидом 17–33 приводит к снижению уровня мозгового бета-амилоида и восстановлению морфофункциональных показателей мозга.

In the work we investigated the possibility of reducing the harmful effects of ionizing radiation on the hematopoietic system of mice with linear and cyclic peptides of low molecular weight. The results showed that after double intraperitoneal injections of the peptides to irradiated mice (4 Gy) practically complete regeneration of cellularity of bone marrow and spleen observed on the eighth day. Signs of accelerated hematopoietic regeneration in the peripheral blood (reticulocytosis) were also observed. The positive effect of the peptides by criteria of cellularity of the bone marrow and spleen was seen in animals got higher dose (6 Gy), however, it was poorer, as compared with the effect observed in mice with dose of 4 Gy.

Our work was aimed at researching into the influence of dipeptide (gamma-dGlu-dTrp) "Timodepressin" and this dipeptide-based tripeptides on the colony-forming ability of the irradiated in vitro bone marrow and hemopoietic stem cells of the normal organism. Also studied was the effect of various doses (1-1000 microg/kg) of one oftripeptides (dAla-gammadGlu-dTrp) on the output of exogenous splenic colonies in the case of its introduction 48 hours before irradiation. It is shown that the mode of influence of the preparations produced on the basis ofdipeptides dGlu-dTrp and gamma-dGlu-dTrp on the initial stages ofa hemopoiesis in the normal and irradiated organism depends on the nature of the additional amino-acid residue and its optical orientation.

We have performed structural and functional studies of the hemotropic activity for a number of novel 2,5-diketopiperazine peptidomimetic derivatives. We employed a mouse model of hemopoietic stem cells cloning in the spleen of lethally irradiated animals. Biologic activity of synthetic products was studied in two experimental models: 1) in vitro irradiated bone marrow SFU-S was used for studying the radio modifying activity; 2) the biological effect of peptidomimetics on the intact non-irradiated bone marrow was evaluated in vivo. Various ways of administration of the peptidomimetics studied were used in the in vivo experiments: intravenous, intraperitoneal or subcutaneous injections and oral administration in the dose range of 10-10000 microg/kg. As a result of our work, we have discovered 2,5-diketopiperazine peptidomimetic derivatives with the dual activity: stimulation of intact committed SFU-S and radiomodifying activity.

The hydrolytic stability of therapeutic peptides such as dalargin, stemokin and some others, including cyclic tripeptides modified by ibuprofen and aspirin, was studied. Two experimental systems were used, one containing purified enzymes pepsin, trypsin and chymotrypsin and other based on fragments of rat stomach and ileum. It was found that linear peptides without D-aminoacids are hydrolyzed by fragments of stomach and ileum but resistant to hydrolysis with purified enzymes. The peptides with D-aminoacids and cyclic peptides are stable in all experimental conditions used, however, peptides modified with aspirin lost acetyl moiety of aspirin residue in acidic medium, the process is accelerated in presence of pepsin.

The hydrolytic stability of a range of cyclic tripeptides, including the therapeutically important dalargin and stemokin, as well as peptides modified by ibuprofen and aspirin, has been studied. The first two experimental systems used utilized purified enzymes (pepsin, trypsin, and chymotrypsin), while the second one utilized fragments of the stomach and small intestine of rats. The linear peptides containing only L -amino acid residues were shown to be hydrolyzed by stomach and intestine fragments, although some of these peptides were resistant to hydrolysis by individual enzymes. The peptides containing D -amino acid residues and cyclic peptides were stable under all of the conditions used, but the peptides modified by aspirin lost the acetyl group of the aspirin moiety in acidic media, this process being accelerated in the presence of pepsin.

Administration of indomethacin (250 mg/kg) to mice was followed by the formation of severe ulcerative and erosive injury to the gastric mucosa, inhibition of DNA synthesis, and development of oxidative stress. Fivefold pretreatment with sedatin (100 μg/kg) decreased the area of indomethacin-induced ulcers and erosions, stimulated DNA synthesis, and reduced the severity of oxidative stress. Non-arginine dermorphin analogue did not stimulate DNA synthesis and had no effect on the degree of oxidative stress. After pretreatment with L-NAME, sedatin did not modulate the synthesis of DNA under conditions of indomethacin-induced injury to the gastric mucosa.

We studied the effect of dipeptide γ-d-Glu-d-Trp (thymodepressin) on migration of CD34+ hemopoietic precursors and their direct adhesion to fibronectin in tumor-bearing mice on days 8, 11, 15, and 17 of tumor growth and on expression of CXCR-4 (CD184+) to SDF-1 and integrin β1 (CD29+) by bone marrow cells. In tumor-bearing mice treated with γ-d-Glu-d-Trp, the percent of CD34+ hemopoietic precursors in the peripheral blood considerably decreased throughout the observation period; the content of CD34+ hemopoietic precursors in the tumor tissue was 2–3-fold below the control against the background of increased content of CD34+ cells in the bone marrow. In animals treated with the peptide, the content of cells expressing CXCR-4 in the peripheral blood, bone marrow, and tumor tissue significantly decreased, while the percent of cells expressing integrin β1 receptor (CD29+) in the bone marrow increased 2-fold, which was paralleled by an almost 2-fold increase in the percent of cells binding to fibronectin. We hypothesized that dipeptide γ-d-Glu-d-Trp suppressed mobilization/migration of CD34+ hemopoietic precursor cells from the bone marrow to the peripheral blood of tumor-bearing mice.

We studied delayed effects of sedatin (arginine-containing μ/δ-opioid receptor agonist) and its arginine-free analog on the ontogenetic development of fish. Fertilized eggs of the Amur sturgeon (Acipenser schrenckii) were treated with the peptides. The results indicate that sedatin increased activities of the antiradical and antioxidant defense systems, stimulated fry development by increasing body weight and length, and increased the number of nucleoli in the myocardium and liver of young fish.

We studied the effect of thymodepressin on migration and adhesion of mouse hemopoietic CD34+ cells under normal conditions and under the effect of granulocytic CSF. It was found that the peptide reduced the absolute number of CD34+ hemopoietic cells in the peripheral blood, increased the percent of cells bound to fibronectin and expressing receptor for integrin β1 (CD29+) in the bone marrow of mice under normal conditions and after stimulation with granulocytic CSF, and reduced the relative number of cells carrying CXCR4 receptor for stromal factor-1 (CD184+) in the bone marrow (CD34+CD184+) and blood (CD184+) of mice stimulated with granulocytic CSF. The results suggest that thymodepressin can inhibit migration of CD34+ cells from bone marrow into peripheral blood under conditions of normal and granulocytic CSF-stimulated hemopoiesis.

Treatment with synthetic arginine-containing dermorphin analogue sedatin (100 mg/kg, 5 intraperitoneal injections) stimulated DNA synthesis in the gastric mucosa and decreased spontaneous and induced chemiluminescence in homogenates of the stomach from albino rats. Non-arginine sedatin analogue in the same dose had little effect on DNA synthesis and free radical oxidation. Fivefold treatment with NO synthase inhibitor L-NAME (9.3×10−5 mol/kg) suppressed DNA synthesis in the gastric mucosa. Sedatin did not modulate DNA synthesis against the background of L-NAME administration.

The influence of Glu-Trp (EW) synthetic dipeptide isomers on hemopoietic progenitor cells and certain immune response reactions is determined by their optical and chemical properties. Thus, the all L-amino acid containing dipeptides L-Glu-L-Trp and L-gammaGlu-L-Trp have no effect on proliferation of committed and pluripotent CFU-S in intact bone marrow. The optical isomers of the Glu residue are an essential determinant of the EW dipeptide biological activity. The inversion of the amino acid optical form imparts suppressor properties: D-Glu-D-Trp,D--gammaGlu-D-Trp, D-Glu-L-Trp and D-gammaGlu-L-Trp inhibit proliferation of hemopoietic progenitors in intact bone marrow. The type of the peptide bond between L-Glu and Trp is another important factor for the biological activity of the L-Glu-containing peptides. Unlike L-Glu-D-Trp with alpha-peptide bond, the dipeptide L-gammaGlu-D-Trp with gamma-peptide bond stimulates CFU-S-8 proliferation in intact bone marrow. The diverse effects of the EW optical isomers on hemopoietic progenitors underlie the radioprotective properties of the D-Glu-containing dipeptides and the radiotherapeutic ones of the L-Glu dipeptides. In animals, pre-irradiation injection of D-Glu-D-Trp, D-gammaGlu-D-Trp, D-Glu-L-Trp, D-gammaGlu-L-Trp, or post-irradiation injection of L-Glu-L-Trp, L-gammaGlu-L-Trp promoted regeneration of the hemopoietic progenitor population.

We studied the effects of optical (dd-, ll-, dl-, and ld-dipeptides with α-bond, EW) structural isomers and cyclic (dd-, ll-, dl-, and ld-dipeptides with γ-bond, iEW) analogs of Glu-Trp synthetic dipeptide on the population of normal hemopoietic stem cells. Dipeptides containing lGlu (lGlu-lTrp, lGlu-dTrp) injected to mice were inert towards committed bone marrow CFU-S; dGlu-containing dipeptides (dGlu-dTrp, dGlu-lTrp) inhibited the growth of CFU-S-8; and LiGlu-dTrp stimulated these cells. Inhibitory or stimulatory effects of optical and chemical isomers of Glu-Trp dipeptide are determined by optical orientation and nature of peptide bond of Glu residue. The effects of cyclic and mixed peptides towards colony formation are similar to those of the corresponding linear dipeptides.

Studies of cooperative effects of D- or L-isomers of iEW dipeptide and neuraminidase on the number of splenic CFU, comparison of the peptide effects on CD34+ cells and splenic CFU, and evaluation of the effects of iEW D- or L-isomers on parameters of T-cell activation showed that interactions of L-(iEW) and D-(iEW) with CD34+ surface receptors are realized through the same mechanism, while their different biological effects on hemopoietic precursor cells in vivo can be explained by their different influence on T-component of the microenvironment.