[Investigation of ligand-receptor interaction and biodistribution of a drug containing cattle retinal polypeptides in various administration routes]. / Issledovanie ligand-retseptornogo vzaimodeistviya i bioraspredeleniya pri razlichnykh rezhimakh vvedeniya lekarstvennogo sredstva, soderzhashchego polipeptidy setchatki glaz skota.

It is important to understand the features of the interaction of drug components with body receptors and obtain data on its distribution in various administration routes in recommended doses in order for its usage in clinical practice to be safe and effective. PURPOSE: To investigate in vitro the interactions of a drug consisting of water-soluble polypeptide fractions produced on animal retina with a wide range of receptor targets, and to assess its biodistribution in the organs of laboratory animals. MATERIAL AND METHODS: The biodistribution of the radioactively marked drug in different organs and tissues of laboratory mice in various routes of administration was studied at the National Research Centre «Kurchatov Institute¼. Evaluation of the ligand-receptor interaction of the drug was carried out in the laboratory at Eurofins Pharma Discovery Services by the method of competitive radioligand binding. RESULTS: A significant effect of the interaction of the polypeptide drug was revealed with different subtypes of glutamate receptors: AMPA, NMDA, and mGluR1. As a result of an in vivo test, we have obtained biodistribution data of the drug for intravenous, intramuscular and parabulbar administration, and the dynamics of drug accumulation in the tissues of the brain and eyes. CONCLUSION: According to the study results, the peptide drug binds to receptors associated with the loss of retinal ganglion cells. Interaction with these receptors potentially provides the test subject with neuroprotective effect. The content dynamics of the studied drug in the blood of animals depends on the route of administration and the amount of drug administered. At the time point of 0.5 hours for intravenous and intramuscular administration in the dose of 1.7 mg/kg, the studied drug has sufficiently high bioavailability in the tissues of the brain and eye. The data suggest that the main route of excretion of the studied drug is through kidneys.

AGGREGATION BY LECTIN-METHODICAL APPROACH FOR EFFECTIVE ISOLATION OF EXOSOMES FROM CELL CULTURE SUPERNATANT FOR PROTEOME PROFILING.

Exosomes are small membrane vesicles secreted by most cell types in vivo and in vitro. Exosomes are found in cell culture media, blood, urine, amniotic fluid, malignant ascite fluids and contain distinct subsets of microRNAs and proteins depending upon the tissue from which they are secreted. Thus exosomes constitute potential biomarkers of human diseases, such as cancer. A major bottleneck in the development of exosome-based diagnostic assays is the challenging purification of these vesicles; this requires time-consuming and instrument-based procedures. Isolation of exosomes can be a tedious, non-specific, and difficult process. Here, we provide a preparative technique for isolation of exosomes based on their ability to aggregate in the presence of lectins. The new method for lectin-based isolation of exosomes from cell culture media was developed as a sample preparation step for exosome-based protein biomarker research.

[Histone deacetylase inhibitors cause the TP53-dependent induction of p21/Waf1 in tumor cells carrying mutations in TP53].

p21/Waf1 protein is one of the main cell cycle arrest regulators and one of the most well-known transcriptional targets of TP53 protein. Here, we demonstrated the activation of expression of the p21/Waf1 gene when the cells were treated to sodium butyrate (NaBu)--one of the natural inhibitors of deacetylase, and investigated whether this phenomenon depends on the presence of functionally active TP53 protein. We compared the effect of the NaBu treatment on the human cell line with different TP53 mutation profile, including: wild-type TP53, single nucleotide substitutions, and the complete absence of TP53 gene. NaBu activated the TP53 protein via hyper acetylation at lysine residue K382, without significant changes in the level of protein expression. Western blotting demonstrated that the addition of NaBu triggers a significant increase in the p21/Waf1 protein level in both the TP53 wild-type cells and in the cells with single nucleotide substitutions in the domain responsible for the binding of TP53 protein to DNA. At the same time, no the p21/Waf1 protein induction was observed in the cells with complete deletion of the TP53 gene. However, NaBu was not able to induce the p2 1/Waf1 production when the expression of TP53 was transiently knocked down by the p53 siRNA. Overall, our results suggest that the NaBu-dependent induction of p21/Waf1 does require the presence of TP53 protein but unexpectedly it can occur regardless of mutational changes in the domain responsible for the TP53 binding to DNA. One of the hypothetical explanations is that NaBu increases the level of TP53 acetylation, and the modified protein is able to establish a new network of protein-protein interactions or trigger some conformational changes affecting the TP53-dependent transcriptional machinery even when its DNA binding ability is impaired.