Načrtovanje, sinteza in vrednotenje heteroaromatskih dvojnih zaviralcev butirilholin esteraze in z mitogenom aktivirane protein kinaze p38α : raziskovalni podatki, obravnavani v doktorskem delu

Ferjančič Benetik, Svit (author), Obreza, Aleš (mentor), Košak, Urban (comentor)

S staranjem populacije postaja Alzheimerjeva bolezen (AB), najpogostejša nevrodegenerativna motnja in oblika demence, vse večji izziv za bolnike, svojce in zdravstvene sisteme. Pri postopnem upadanju holinergičnega prenosa pri bolnikih z AB ima pomembno vlogo butirilholin esteraza (BChE). Z mitogenom aktivirana protein kinaza p38α (p38α MAPK), ki je vključena v proces nevrovnetja, hiperfosforilacije proteina tau in v patofiziologijo amiloidnih plakov, pa je validirana tarča za upočasnjevanja nevrodegeneracije pri tej bolezni. V tej doktorski disertaciji smo računalniško načrtovali, sintetizirali in ovrednotili prvi strukturni razred dvojnih zaviralcev BChE/p38α MAPK kot možen pristop za zdravljenje AB. Z zamenjavo fenoksi skupine v spojini zadetku z benzilno smo dosegli π-π interakcijo s Trp430 v holin-vezavnem žepu BChE in ustrezno vezavo v hidrofobno regijo I p38α MAPK, kar je privedlo do spojin z nanomolarnimi jakostmi zaviranja obeh tarčnih encimov. Najobetavnejši spojini sta bili selektivni za p38α MAPK v panelu 103 kinaz in učinkoviti v in vivo modelih kognitivnih motenj. Z racionalnim načrtovanjem na podlagi kristalnih struktur prej omenjenih spojin v aktivnih mestih obeh encimov smo uspešno pripravili serijo karbamatnih plejotropnih predzdravil, ki po psevdo-ireverzibilni vezavi na Ser198 BChE sprostijo alkohol, ki nato kompetitivno zavira p38α MAPK. V želji po razvoju močnejših zaviralcev p38α MAPK, a ohranjeni visoki selektivnosti napram ostalim kinazam, smo načrtovali še eno serijo karbamatnih dvojnih zaviralcev BChE/p38α MAPK, ki se kovalentno vežejo v aktivno mesto BChE in so tudi alosterični zaviralci p38α MAPK. Z uvedbo karbamatne skupine smo pripravili spojine z nizko nanomolarno afiniteto za BChE in prokognitivnimi učinki v in vivo modelu kognitivnega upada. Ta sprememba strukture je povzročila izgubo pomembnih interakcij v alosteričnem vezavnem mestu p38α MAPK in s tem prenizko aktivnost za ovrednotenje kinazne selektivnosti.

• Language: Slovenian
• Keywords: butirilholin esteraza, z mitogenom aktivirana protein kinaza p38α, strukturno podprto načrtovanje, večtarčni ligandi, plejotropno predzdravilo, psevdo-ireverzibilni zaviralci, skopolaminski model, LPS model
• Typology: 2.20 - Complete scientific database of research data
• Organization: FFA - Faculty of Pharmacy
• Year: 2025
• PID: 20.500.12556/RUL-172004
• Data col. methods: Experiment: Laboratory
• Publication date in RUL: 05.09.2025

Licences

• License: CC BY 4.0, Creative Commons Attribution 4.0 International
• Link: http://creativecommons.org/licenses/by/4.0/
• Description: This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.

Secondary language

• Language: English
• Title: Design, synthesis and evaluation of heteroaromatic dual inhibitors of butyrylcholinesterase and p38α mitogen-activated protein kinase : research data underlying the doctoral dissertation

Abstract

With an aging population, Alzheimer’s disease (AD) — the most common neurodegenerative disorder and form of dementia — increasingly challenges patients, families, and healthcare systems. Butyrylcholinesterase (BChE) plays a key role in the progressive decline of cholinergic transmission observed in AD. Mitogen-activated protein kinase p38α (p38α MAPK), on the other hand is emerging as a validated target in neurodegeneration, tau hyperphosphorylation, and amyloid plaque pathology. In this doctoral dissertation, we computationally designed, synthesized, and evaluated first-in-class dual inhibitors of hBChE/p38α MAPK as potential anti-AD drugs. Replacing the phenoxy group of the hit compound with a benzyl group enabled π-π stacking with Trp430 of the BChE choline-binding pocket and favourable binding within p38α MAPK’s hydrophobic region I, achieving nanomolar inhibitory potencies on both targets. The two lead compounds showed selectivity for p38α MAPK in a panel of 103 kinases and were effective in vivo in two models of cognitive decline. Subsequently, using rational design based on the crystal structures of the complexes of these compounds with the target enzymes, we developed carbamate-based pleiotropic prodrugs of p38α MAPK inhibitors. These prodrugs are activated via pseudo-irreversible covalent binding to Ser198 of BChE, releasing an alcohol that competitively inhibits p38α MAPK. To enhance the affinity for p38α MAPK while retaining high selectivity over other kinases, we designed another series of dual carbamate inhibitors that covalently bind to BChE and act as allosteric p38α MAPK inhibitors. Introduction of the carbamate group produced compounds with low nanomolar affinity for BChE and procognitive effects in vivo, but also caused a loss of important interactions in the allosteric site of p38α MAPK and thus insufficient inhibitory potency for kinase selectivity evaluation.

• Keywords: butyrylcholinesterase, p38α mitogen-activated protein kinase, structure-based drug design, multitarget-directed ligands, pleiotropic prodrug, pseudo-irreversible inhibitors, scopolamine model, LPS model

Projects

• Funder: ARIS - Slovenian Research and Innovation Agency
• Project number: P1-0208
• Name: Farmacevtska kemija: načrtovanje, sinteza in vrednotenje učinkovin

• Funder: ARIS - Slovenian Research and Innovation Agency
• Project number: N1-0277
• Name: Raziskave multifunkcionalnih spojin, usmerjenih proti nevroinflamaciji in holinergičnemu pomanjkanju pri Alzheimerjevi bolezni