Sinteza in vrednotenje derivatov kinolin-2-ona kot zaviralcev N-acetilglukozaminil transferaze

Gruden, David (author), Tomašič, Tihomir (mentor)

Glikoziltransferaze so encimi, ki pripenjajo sladkorne komponente na številne akceptorske (makro)molekule v celicah, najpogosteje preko tvorbe vezi z nukleofilnim kisikom v njihovi strukturi. Za svoje delovanje potrebujejo donorske substrate aktivirane s fosfatnimi izstopajočimi skupinami, ki so v večini primerov nukleozidni difosfati. O-β-N-acetil-D-glukozaminil transferaza (OGT) je edina glikoziltransferaza, ki katalizira prenos N-acetilglukozaminske skupine (GlcNAc) iz donorskega substrata UDP-GlcNAc na različne serinske (Ser) in treoninske (Thr) aminokislinske ostanke celičnih proteinov. Tovrstna posttranslacijska modifikacija (PTM) proteinov ima pomembno vlogo v številnih celičnih procesih. Različne motnje v PTM proteinov z GlcNAc so opazili tudi pri kroničnih boleznih, kot so diabetes, rak, srčno-žilne bolezni in Alzheimerjeva bolezen. Zaradi velikega vpliva OGT na pravilno delovanje celic in razvoj omenjenih kroničnih bolezni, bi imela sinteza učinkovitih in selektivnih zaviralcev tega encima velik pomen pri nadaljnjem proučevanju in zdravljenju teh bolezni. V tem magistrskem delu smo načrtovali in sintetizirali potencialne zaviralce OGT, ki smo jih osnovali na kinolin-2-onskem ogrodju, katerega so raziskovalci na Fakulteti za farmacijo v Ljubljani odkrili z virtualnim rešetanjem na podlagi strukture vezavnega mesta OGT. Fragmenti s tem ogrodjem so šibki zaviralci OGT, ki se vežejo v vezavno mesto uridinske skupine donorskega substrata. S sintezo nove knjižnice zaviralcev s kinolin-2-onskim ogrodjem smo želeli izboljšati fizikalno-kemijske lastnosti teh fragmentov in proučiti odnos med strukturo potencialnih zaviralcev in njihovim delovanjem na encimu OGT. Na osnovni kinolin-2-onski skelet smo pripenjali različne aromatske in alifatske aminske substituente z reakcijo reduktivnega aminiranja. S tem smo povečali molekulo in proučili vplive, ki jih imajo različni substituenti na zaviralne aktivnosti končnih spojin. Končnim spojinam smo nato preverili zaviralno aktivnost na encimu z direktnim določanjem aktivnosti na podlagi fluorescence. Iz preliminarnih rezultatov testiranja smo ugotovili, da bo potrebno omenjeno metodo dodatno optimizirati, preden bo primerna za natančno in zanesljivo testiranje končnih spojin.

• Language: Slovenian
• Keywords: O-β-N-acetil-D-glukozaminil transferaza, encim, OGT, UDP-GlcNAc, zaviralci, kinolin-2-onski skelet
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2021
• PID: 20.500.12556/RUL-127791
• Publication date in RUL: 23.06.2021

Secondary language

• Language: English
• Title: Synthesis and evaluation of quinolin-2-one derivatives as N-acetylglucosaminyl transferase inhibitors

Abstract

Glycosyltransferases (GT) are enzymes that catalyze the attachment of a sugar moiety to many different acceptor molecules in cells, in most cases through a glycosidic bond with nucleophilic oxygen of the acceptor. They use donor substrates activated with phosphate leaving groups, most commonly nucleoside diphosphates. O-β-N-acetyl-D-glucosaminyl transferase (OGT) is the only GT that catalyzes the addition of the N-acetylglucosamine (GlcNAc) moiety from the donor substrate UDP-GlcNAc to serine (Ser) and threonine (Thr) residues of acceptor proteins. This post-translational modification (PTM) plays an important role in a variety of cellular processes. Abnormal levels of O-GlcNAcylation have been found in chronic diseases such as diabetes, cancer, cardiovascular diseases, and Alzheimer's disease. Due to the important role that OGT plays in normal cell functions and in the pathogenesis of various chronic diseases, the synthesis of potent and selective inhibitors would greatly impact the future understanding and treatment of these diseases. In this Master's thesis, we designed and synthesized potential OGT inhibitors based on the quinolin-2-one scaffold discovered by researchers at the Faculty of Pharmacy in Ljubljana by structure-based virtual screening. Fragments based on this core are weak OGT inhibitors that target the uridine binding site of the donor substrate. By synthesizing a new library of inhibitors with this core, we aimed to improve the physicochemical properties of the fragments and explore their structure-activity relationships (SAR). We coupled various aromatic and aliphatic amines to the quinolin-2-one core by reductive amination. In this way, we enlarged the molecules and investigated the effect that different substituents have on the potency of inhibition. The final compounds were then tested for their inhibitory activity against OGT using a direct fluorescence activity assay. The preliminary results indicated that future modifications of the assay are needed in order to obtain more accurate and reliable results.

• Keywords: O-β-N-acetyl-D-glucosaminyl transferase, enzyme, OGT, UDP-GlcNAc, inhibitors, quinolin-2-one core