In silico & In vitro screening for the identification of CSE modulators.

Postgraduate Thesis uoadl:3328488 38 Read counter

Κατεύθυνση Φαρμακευτική Χημεία
Library of the School of Science
Deposit date:
Basdeki Aikaterini
Supervisors info:
Εμμανουήλ Μικρός Καθηγητής Τμήμα Φαρμακευτικής ΕΚΠΑ,
Α. Παπαπετρόπουλος Καθηγητής Τμήμα Φαρμακευτικής ΕΚΠΑ,
Ι. Κ. Κωστάκης Αναπλ. Καθηγητής Τμήμα Φαρμακευτικής ΕΚΠΑ
Original Title:
Νέα μόρια-τροποποιητές της γ-λυάσης της κυσταθειονίνης (Cystathionine gamma-lyase, CSE). In silico & In vitro αξιολόγηση.
Translated title:
In silico & In vitro screening for the identification of CSE modulators.
Hydrogen sulfide (H2S), a colorless gas with a characteristic odor, has been known since 1713. For many years, studies were conducted exclusively on its toxicity, until its endogenous production in mammals was discovered. This discovery prompted the scientific community to investigate its function and its involvement in the various systems of the body. It is now recognized as the third gaseous transmitter along with nitric oxide (NO) and carbon monoxide (CO). That is, it is an enzymatically endogenously produced gas, free to penetrate cell membranes and engage with a variety of cellular targets. Cystathionine γ-lyase (CSE) is one of the three enzymes in the human body responsible for the biosynthesis of hydrogen sulphide. The other two are cystathionine β-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3MST). The basic substrate of the CSE enzyme to produce hydrogen sulfide in the human body is L-Cysteine, while the cofactor pyridoxal-5'-phosphate (PLP) is necessary for the enzyme reaction to take place. The CSE/H2S system is involved in a multitude of signaling pathways in human physiology and pathophysiology, making CSE a potential pharmacological target. Hydrogen sulfide is characterized by its dual nature. Depending on the system under consideration, higher concentration, or lower concentration than desired has been linked to pathophysiological conditions. For this reason, the development of both inhibitors and agonists of the enzyme is deemed necessary. Few molecules are available in the literature that are selective and potent active molecules that activate or inhibit the CSE enzyme. The aim of this work is to find such compounds.
To achieve this goal, in vitro and in silico techniques were combined. The chemical library of the Pharmacy department of National and Kapodistrian University of Athens, consists of approximately 2000 compounds. These include natural products and synthetic analogues. As a first step of the present work, an in vitro evaluation of approximately 470 synthetic compounds against the CSE enzyme was performed. For these experiments, the analytical colorimetric method Methylene Blue assay was used with CSE enzyme with TAG ligand (Glutathione-S-Transferase) which was overexpressed, isolated and purified. Of the compounds evaluated, two showed interesting activity against the enzyme. A dibenzofuran analogue showed dose-dependent inhibitory activity and a trimethoxy benzyl derivative agonistic activity. These two compounds were used as guide molecules to find compounds with similar characteristics, and possibly the same activity, through in silico methodologies. These can be divided into two parts. The first part involves finding similar compounds to the lead compounds through 2D and 3D similarity screening of the NCI chemical library. The second part involves evaluating the binding affinity to the molecular target, the CSE enzyme. These results were combined through a consensus method, to finally yield 39 molecules with candidate activity. At the same time, an attempt was made to organically synthesize three analogues of the trimethoxy benzyl derivative with minor conformational changes. The compounds resulting from the computational methods as well as from the organic synthesis were evaluated in vitro, but without yielding any significant result.
Main subject category:
Hydrogen sulfide, Cystathionine gamma-lyase, In vitro screening, In silico screening
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File access is restricted until 2026-05-26.

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File access is restricted until 2026-05-26.