Problems of Biological Medical and Pharmaceutical Chemistry

The review article discusses the currently used preclinical research methods of angioprotective activity of biologically active substances. Published data indicate the social and economic significance of chronic venous pathologies as a common group of diseases affecting a significant part of the population, characterized by a major risk of complications that require prompt correction. One of the main methods of complex treatment is pharmacotherapy with phlebotropic agents, due to their complex effect on various links of this pathology: increasing vascular tone, reducing pain, improving lymphatic drainage, beneficially affecting all stages of inflammation, membrane-stabilizing and antioxidant effect. The creation and introduction of new and effective medicines of phleboprotective action, including herbal origin, is one of the priority areas of pharmacology and medicine. However, preclinical trials, as one of the long stages of drug development, need optimization. The purpose of the publication: systematization of methods for determining angioprotective activity, consisting of anti-inflammatory, antioxidant, capillary-strengthening activity. The review contains information about the main stages of the preclinical phase of testing – from the search for promising compounds to the developed and used preliminary pharmacological tests to determine biological activity.

Introduction. Regulatory documentation is an important tool for ensuring the safety and efficacy of new drugs. Pharmaceutical articles (PM) must contain validated methods for determining authenticity (identification) indicators. The validation procedure allows for the timely identification of shortcomings in the process of developing new methods and their improvement at early stages.

The aim of study is to develop and validate a method for determining a new biologically active compound 5-methyl-N-phenyl-7-(4-fluorophenyl)-4,7-dihydrotetrazolo [1,5-a]pyrimidine-6-carboxamide by spectrophotometry in the UV region. For the validation assessment of the determination method, the following characteristics were selected: specificity, linearity, correctness, precision under repeatability conditions (convergence).

Material and methods. The following samples and solvents were used in the experiment: substance (colorless crystalline powder); DMF solvent; UV-1800 spectrophotometer, manufactured by Shimadzu, Japan.

Results. To assess the specificity, the absorption spectrum of the substance was obtained in the selected range in the UV region. The spectrum shows an absorption maximum at 285 nm. To establish the specific absorption index, sample solutions were prepared in the concentration range from 0.00060% to 0.0010% and it was equal to 270. The linearity of the method was established based on the study of the dependence of optical density on the content at 5 concentration levels from 80 to 120%. The acceptance criterion is the correlation coefficient, it is 0.9954. The validated method is considered correct if the values accepted as true lie within the confidence intervals of the corresponding average analysis results. The determination was carried out at 3 concentration levels (80%, 100%, 120%) by determining the content of the analyzed compound in model solutions. The acceptance criterion is the average percentage of openness and its average value should be within 100±5%, it is within the range from 97.5% to 102.0%. The repeatability of the analytical method was assessed based on six independent results obtained under the same regulated conditions in one laboratory. The calculated value of the variation coefficient is 1.5%, and the relative error of determination is 0.54% with a confidence probability of p=95%, which indicates the precision of the method under repeatability conditions.

Conclusion. The value of the specific absorption index of a new biologically active compound has been established, which can be used to establish authenticity (identification) and determine the quantitative content when developing a pharmacopoeial article. A validation assessment of the developed method has been carried out.

In recent years, there has been an increase in cases of drug contamination with ethylene glycol (EG) and diethylene glycol (DEG) – toxic polyatomic alcohols that pose a serious threat to human health even at minimal concentrations. These hazardous substances may be present in medications as impurities due to the adulteration of raw materials or the use of substandard excipients. In response to this global threat, international regulatory agencies and national pharmacopoeias have introduced requirements mandating testing for EG and DEG impurities. This article provides a detailed review of current regulatory approaches to controlling these impurities at global and national levels. Special attention is given to the analysis of key regulatory documents and recent initiatives. The article examines WHO medical alerts (2022–2024), which mandate the testing of all batches of potentially hazardous excipients, as well as FDA warning letters (2023–2024) addressed to manufacturers violating GMP rules in raw material quality control. It also describes the FDA’s industry guidance on testing excipients most susceptible to EG and DEG contamination, along with the WHO methodology for monitoring these impurities in finished dosage forms. A particular focus is placed on harmonized pharmacopoeial requirements, including general chapters from various pharmacopoeias (US, Russian, European, Belarusian, EAEU, Uzbekistan, and Kazakhstan) that regulate the determination of EG and DEG in ethoxylated substances and establish maximum permissible limits – no more than 0.1% for each. Notably, the article mentions a new general pharmacopoeial monograph introduced in the Pharmacopoeia of the Republic of Uzbekistan, which allows for the control of EG and DEG impurities in finished dosage forms. Special emphasis is placed on risk minimization strategies at all stages of drug manufacturing and the need for a comprehensive approach to drug safety. The analysis of international experience and modern regulatory practices helps identify key directions for ensuring drug safety, including harmonizing standards, strengthening quality control, and implementing advanced analytical methods.

Introduction. Renal cell carcinoma (RCC) is one of the most aggressive tumors of the urinary system. Immune checkpoints such as PD-1 and PD-L1 play an important role in regulating the antitumor immune response. Serum levels of soluble forms of PD-1 (sPD-1) and PD-L1 (sPD-L1) may have some clinical significance in RCC, but their prognostic value remains poorly understood.

The aim – to analyze the relationship between serum levels of sPD-1 and sPD-L1 in patients with renal cell carcinoma and overall survival rates.

Material and methods. The study of sPD-1 and sPD-L1 levels was performed in the blood serum before treatment in 105 patients with RCC (63 men and 42 women) aged 33 to 81 years (median 60 years) at various stages of the disease (I – 55, II – 12, III – 17,
IV – 21). Clear cell renal carcinoma was detected in 90 patients, chromophobe in 6, papillary type I in 6, papillary type II in 3, with the differentiation degree of G1 (9 tumors), G2 – 60, G3 –18, G4 – 12. The concentration of sPD-L1 and sPD-1 was determined in the blood serum of patients with renal neoplasms before treatment using the Human PD-L1 Platinum ELISA and Human PD-1 ELISA kits (Affimetrix, eBioscience, USA) according to the manufacturer's instructions. To identify differences between values, one-way and multivariate analysis of variance (ANOVA) were applied. Survival curve analysis was performed using the Kaplan-Meier method, and survival curve comparisons were conducted using the Log-Rank test with Bonferroni correction for multiple comparisons. A multivariate analysis of the impact of factors on overall survival was carried out using the Cox proportional hazards regression model. Differences were considered statistically significant at p<0.05. All calculations were performed on a personal computer using the statistical software packages "Statistica for Windows" and SPSS.

Results. Serum sPD-L1 levels were associated with worse overall survival in patients with RCC. Patients with sPD-L1 concentration ≥ 42.6 pg/mL demonstrated a significant reduction in 5-year overall survival compared to patients with low levels of this marker (p=0.0012). In multivariate analysis, sPD-L1 was an independent prognostic factor along with tumor size and the presence of metastases. A significant difference in overall survival in groups of patients with RCC was also shown when dividing patients by the sPD-1 level equal to 76 pg/ml, which confirms the significance of studying the content of soluble forms of these proteins in the blood serum of patients with RCC.

Conclusions. High serum sPD-L1 levels are associated with poor prognosis in RCC and can be used as a potential biomarker for patient stratification. Further studies are needed to confirm the clinical significance of sPD-L1 and develop personalized treatment strategies for RCC.

The article is dedicated to Viktor Vasilyevich Banin, Corresponding Member of the Russian Academy of Sciences, Doctor of Medical Sciences, Professor, an outstanding Soviet and Russian morphologist, who worked for many years at the 2nd MOLGMI (RSMU), Deputy Head of the Research and Educational and Methodological Center for Biomedical Technologies of the All-Russian Research Institute of Medicinal and Aromatic Plants (R&D Center for Biomedical Technologies of the FGBNU VILAR, Moscow) and currently continues his scientific and teaching activities at the A.I. Evdokimov Moscow State Medical University. The article presents information about the main stages of his scientific and practical activities in Russia and abroad, scientific areas of fundamental and applied research. The main scientific works of V.V. Banin are presented, their role in the development of domestic and foreign science, the training of highly qualified scientific personnel is shown.