Abstract

   Development of an effective and safe antioxidant compound is still challenging in the last few decades. There has been an increasing interest in the role of reactive oxygen species (ROS) in food, drugs, and even living system. Free radical formation is associated with the normal natural metabolism of aerobic cells. They are inevitably exposed to reactive oxygen species formed as oxygen metabolites. Oxidative stress which is largely characterized by reactive oxygen and nitrogen species is implicated in the development of a number of chronic and degenerative diseases such as atherosclerosis, cancer, cirrhosis, diabetes, wound healing and aging. Free radicals are highly reactive and therefore can attack membrane lipids, generating carbon radicals and peroxy radicals, which cause lipid peroxidation. Therefore, scientists in various disciplines have become more interested in naturally-occurring antioxidants as well as in related synthetic derivatives that could provide active components which prevent or reduce the impact of oxidative stress. Іn order to study the effect of various substituents in the molecules on the nature of thepharmacological activity of thiazolo[4,5-b]pyridin-2-ones, a series of new compounds were synthesized based on the previously obtained 5,7-dimethyl-3Hthiazolo[4,5-b]pyridin-2-one. The high electrophilicity of the N3 position in 5,7-dimethyl-3H-thiazolo[4,5-b]pyridin- 2-one allows to use of its functionalization as a convenient method for obtaining various N3-substituted derivatives to expand the range of thiazolo[4,5-b]pyridines. In particular, an NH center with a mobile hydrogen atom at the N3 position in 5,7-dimethyl-3H-thiazolo[4,5-b]pyridin-2-one allows conducting a synthesis based on 3-substituted derivatives. This conversion was carried out through the stage of obtaining potassium salt. Several chloroacetamides were tested as alkylating agents, which allowed to obtain the corresponding 2-(5,7-dimethyl-2-oxo-thiazolo[4,5-b]pyridin-3-yl)-N-aryl-acetamides (1–6). Methods of quantitative elemental analysis, mass spectrometry, and 1H NMR spectroscopy were used to confirm the structure and individuality of the synthesized substances. Interpretation of the spectra revealed that the signals for protons of all structural units were observed in their characteristic ranges.

Mounting research has been performed in the recent decades focusing on natural and low-molecular-weight synthetic antioxidants discovering as key molecules that control oxidative damage and its pathway to disease.

Oxidative stress is a phenomenon resulting from the imbalance between oxidation-reduction processes, in particular, the formation and accumulation reactive oxygen species (ROS) and reactive nitrogen species (RNS) in cells and tissues, and the ability of the antioxidant defence system of the organism to eliminate these by-products. Oxidative stress develops under the influence of external or internal factors and leads to oxidative modification of biomolecules, in particular lipids, proteins and DNA [6]. One- and two-electron oxidation-reduction reactions, as an integral part of aerobic metabolism, often lead to free radicals’ in vivo formation. Molecular oxygen reduction processes include the stepwise single electron reduction of O₂ results in such ROS generation as superoxide anion radical (O₂^•-), hydrogen peroxide (H₂O₂) and hydroxyl radical (HO^•). H₂O₂ is produced as a result of two-electron O₂ reduction. Reactive nitrogen species include mainly nitric oxide (NO^•), nitrogen dioxide (NO₂^•) and peroxynitrite (ONOO⁻), as well as non-radicals such as nitrous acid HNO₂ and N₂O₄ (dinitrogen tetroxide).

At lower concentrations ROS/RNS have beneficial effects and indulged in different physiological processes such as redox regulation, mitogenic responses, cellular signaling pathways, and an immune function while at a higher level, these reactive species generate nitrosative and oxidative stress.

In modern research the two main types of antioxidants are distinguished: (1) the primary antioxidants, or free radical scavengers, which are able to break the chain reaction; (2) the secondary, or preventive, antioxidants, for which the action mechanisms may include the deactivation of metals, inhibition of lipid hydroperoxides by interrupting the production of undesirable volatiles, the regeneration of primary antioxidants, and the elimination of singlet oxygen. The methods of the antioxidant capacity determining are commonly classified into two main groups, based on the reaction mechanisms involved in free radicals’ reduction process: (a) hydrogen atom transfer (HAT) reactions; and (b) transfer reactions of a single electron (SET). 

Nowadays the discovery of effective antioxidant agents among low-molecular-weight organic molecules is a recent problem that requires new methodological approaches implementation, while it is also the society relevant task [1]. Both thiazole and pyridine scaffolds are of the highest priority in modern medicinal chemistry [2, 3]. Numerous reports concerning variety biological effects possessed by thiazolopyridine derivatives have been currently published including their discovery as potent antioxidant agents [4].

One of the antioxidant action mechanisms can be exerted through the inhibition enzymes’ activity which are responsible for reactive oxygen species (ROS) producing, thereby reducing oxidative stress. The objective of the precent study was to fulfil molecular docking studies of novel 3H-thiazolo[4,5-b]pyridine-2-one derivatives towards lipoxygenase (LOX) as one of ROS-producing enzymes. LOX-5 is the enzyme that catalyses the oxidation of polyunsaturated fatty acids to form lipid hydroperoxides, which can lead to membrane damage and inflammation.

The objects of the precent research were three series of condensed 3H-thiazolo[4,5-b]pyridine derivatives, synthesized at Danylo Halytsky Lviv National Medical University [5-7]: (a) the 1^st series included N³ substituted derivatives containing phenyl, propanenitrile, propanoic acid and phenylpropanamide moieties as N³-substituents while they comprised hydroxyl group at C⁵ position; (b) the 2^nd series comprised compounds incorporated substituted phenyl diazonium or alkyl substituents at C⁶ position of the core scaffold; (c) the 3^rd series included derivatives incorporated chloroacetate, chlorobenzoate, benzyloxybenzoate, methoxyphenyl acrylate and substituted phenyltriazolecarboxylate moieties as the substituents at C⁵ position of the core scaffold (Fig. 1). The antioxidant activity evaluation of all tested compounds was reported as a spectrophotometric DPPH assay based on the ability of antioxidant drug candidates possess free radical scavenging potency. Compounds of all three series were found to exhibit moderate and low antioxidant effects.

Probing the action mechanism of 3H-thiazolo[4,5-b]pyridin-2-one derivatives as antioxidants was performed through molecular docking studies towards lipoxygenase. Docking studies, filtering and poses grouping according to the Estimated Affinity towards the biotarget were carried out using SeeSar13.1.0 software (BioSolveIT, Sankt Augustin, Germany) [8]. The crystal structures of lipoxygenase were downloaded from Protein Data Bank using the Protein Mode of the software. Two structures of LOX-5 were downloaded: PDB entry 3O8Y - a 2.39 Å resolution structure of LOX-5 without ligands; PDB entry 6N2W – a 2.71 Å resolution structure of LOX-5 with co-crystallized ligand NDGA (Masoprocol, 4-[(2r,3s)-3-[(3,4-dihydroxyphenyl)methyl]-2-methylbutyl]benzene-1,2-diol).

Actuality. Discovery of innovative drug candidates is one of the relevant tasks of modern medical and pharmaceutical science. The thiazole core is a versatile scaffold for the development of new drugs and biologically active agents [1]. The recent publications, dealing with synthesis and pharmacological screening of thiazole-bearing heterocycles and their activity mechanisms [2-4], indicate the relevance of further drug discovery among novel derivatives of the specified chemical class.

The objectives. The precent work is devoted to the systematic study and generalization of the main trends in the field of potential antimicrobial drug candidates development of among condensed thiazolo[4,5-b]pyridine derivatives.

Materials and methods. The complex of general scientific methods of searching and systematizing literary references, analysis and comparison of information from various sources was used, followed by generalization of new and promising directions.

Results. Thiazolo[4,5-b]pyridine derivatives have been extensively evaluated as antimicrobial agents. Abd El-Mahmoud reported antimicrobial and antifungal screening for 5-amino-2-thioxo-2,3,4,7-tetrahydro-1,3-thiazolo-[4,5-b]pyridine-6-carbonitriles [5]. All compounds showed moderate antimicrobial activity against Staphylococcus aureus (St). Some compounds from this series showed moderate antimicrobial activity against Bacillus subtilis (B.C.) and Proteus vulgaris (P. vulgaris). The lead compound was highly active against Aspergillus flavus (A. flavus).

El-Sofany et al.reported in vitro antimicrobial and antifungal activity evaluation of novel spiro[cyclohexane-1,2'-thiazolo[4,5-b]pyridine derivatives [6]. SAR analysis revealed that open-chain sugar moieties introduction played a significant role in antimicrobial activity increasing. It was also shown that acetylated aldoses were less active than non-acylated ones and the activity increased by increasing the number of -OH groups rather than -OAc groups in the open-chain sugar nucleus.

Lozynsky et al. reported antimicrobial activity in vitro evaluation for 5,7-substituted 3H-thiazolo[4,5-b]pyridin-2-ones [7]. It was shown that the antibacterial effect did not depend on the substituents at C⁵ and C⁷ positions. The presence of amide fragment was favorable for antimicrobial potency.

Conclusions. The current trends in discovery of new active pharmaceutical ingredients among condensed thiazolo[4,5-b]pyridine derivatives exhibiting antimicrobial activity was based on structural modification strategy of the core condensed scaffold at C², C⁵, C⁶, and C⁷ positions followed by pharmacological screening.

Acknowledgement: O. K. thanks Universidad San Pablo CEU for a Postdoctoral Contract for Ukrainian Researchers 2022-2024.