Sinteza in vrednotenje knjižnice elektrofilnih fragmentov kot kovalentnih zaviralcev kaspaze 1

Kemperle, Žiga (avtor), Knez, Damijan (mentor), Meden, Anže (komentor)

Alzheimerjeva bolezen (AB) je nevrodegenerativna bolezen, pri kateri se pojavi postopna izguba spomina in poslabšanje kognitivnih funkcij. Poglavitni patološki znaki bolezni so plaki amiloida β (Aβ) in nevrofibrilarne pentlje, zelo pomembno vlogo pa ima tudi vnetje centralnega živčnega sistema – nevrovnetje. Prvi znak nevrovnetja je aktivacija mikroglije, kjer se pri provnetnem fenotipu M1 sproščajo citokini, ki aktivirajo sestavljanje inflamasoma NLRP3. Po aktivaciji inflamasoma NLRP3 se z odstranitvijo prodomene aktivira cisteinska provnetna kaspaza-1, ki cepi pro-IL-1β do njegove aktivne oblike IL-1β. Pri mišjih modelih AB farmakološko zaviranje kaspaze-1 preprečuje progresivno kopičenje plakov Aβ, zmanjšuje nevrovnetje in ohranja normalne koncentracije sinaptofizina v hipokampusu. Pri rešetanju knjižnice elektrofilnih fragmentov na UL FFA so bili odkriti trije zadetki kovalentnih bojnih glav – karbamoil fluorid, 1,3,4-oksatiazol-2-on ter imidazolid, ki so zavirali encimsko aktivnost kapsaze-1. V magistrski nalogi smo sintetizirali analoge dveh zadetkov – 1,3,4-oksatiazol-2-ona B ter karbamoil fluorida C, pri čemer smo spreminjali substituente na osnovnem ogrodju (substituenti na indolu pri analogih zadetka C) oziroma zamenjali fenilni obroč s furanom in njegovimi analogi (zadetek B). Dodatno smo na indolinu karbamoil fluoridno bojno glavo zamenjali še z nekaterimi drugimi, v literaturi opisanimi elektrofilnimi skupinami. Po postavitvi in optimizaciji sintezne poti smo uspeli pripraviti 23 spojin, za katere smo ovrednotili jakost zaviranja človeške kaspaze-1. Kaspazo-1 je zaviralo osem karbamoil fluoridov, preostali fragmenti so bili neaktivni. Na osnovi rezultatov lahko sklepamo, da je za zaviralno aktivnost ugodna predvsem prisotnost sterično majhnih 5- ali 6- substituentov (npr. fluor, nitro ali hidroksi), tolerirani pa so tudi estri acetata na mestu 3. Slednji predstavljajo tudi najboljše izhodišče za optimizacijo zaviralne aktivnosti in za rast fragmenta.

• Jezik: Slovenski jezik
• Ključne besede: Alzheimerjeva bolezen, mikroglija, kaspaza-1, kovalentni fragmenti
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2023
• PID: 20.500.12556/RUL-151840
• Datum objave v RUL: 22.10.2023

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Synthesis and evaluation of electrophile fragment library as covalent caspase 1 inhibitors

Izvleček

Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive memory loss and deterioration of cognitive functions. The main pathological features of the disease are amyloid-β (Aβ) plaques and neurofibrillary tangles. In addition, inflammation of the central nervous system (neuroinflammation) also plays a crucial role. The first sign of neuroinflammation is microglial activation, with the pro-inflammatory M1 phenotype releasing cytokines that activate the formation of NLRP3 inflammasome. Following NLRP3 inflammasome activation, proinflammatory cysteine caspase-1 is activated via prodomain removal and cleaves pro-IL-1β into its active form IL-1β. In mouse models of AD, pharmacological inhibition of caspase-1 prevented progressive accumulation of Aβ plaques, reduced neuroinflammation, and maintained normal concentrations of synaptophysin in the hippocampus. Screening of the electrophilic fragment library assembled at UL FFA identified three covalent warhead fragment hits - carbamoyl fluoride, 1,3,4-oxathiazol-2-one, and imidazolide - that inhibited the enzymatic activity of caspase-1. In this master's thesis, we synthesised analogues of two hits –1,3,4-oxathiazol-2-one B and carbamoyl fluoride C – by changing the substituents on the basic scaffold (substituents on indoline in the analogues of hit C) or by replacing the phenyl ring with furan and its analogues (hit B). In addition, the carbamoyl fluoride warhead on indoline was replaced by other known electrophilic groups from literature. After establishing and optimising the synthetic pathway, we prepared 23 compounds for which we determined the inhibitory potencies against human caspase-1. Caspase-1 was inhibited by eight carbamoyl fluorides, while the remaining fragments were inactive. The results suggest that inhibitory activity is favoured by the presence of sterically small substituents (e.g., fluorine, nitro or hydroxy) at positions 5 or 6 of indoline, whereas acetate esters at position 3 are also tolerated. The latter also provide the best starting point for optimisation of inhibitory activity and further fragment growth.

• Ključne besede: Alzheimer's disease, microglia, caspase-1, covalent fragments