Sinteza derivatov kinazolina in izoksazolo[5,4-d]pirimidina z agonističnim delovanjem na Tollu podobni receptor 7

Vičič, Karolina (author), Sova, Matej (mentor)

Vnetni odziv predstavlja obrambni mehanizem, ki se aktivira v primeru izpostavljenosti škodljivim dejavnikom. V sklopu teh procesov imata ključno vlogo prirojeni in pridobljeni imunski sistem, ki se med drugim aktivirata tudi preko Tollu podobnih receptorjev (TLR). Do sedaj je znanih že 10 tipov TLR, za katere je značilno, da kot ligand prepoznajo različne manjše molekule in dele mikroorganizmov. V sklopu magistrske naloge smo se osredotočili na TLR7, ki ga aktivira enovijačna RNA, s tem pa povzroči kaskado procesov, ki vodi do protivnetnega, protivirusnega in protitumornega delovanja. Uporaba agonistov TLR7 bi tako lahko igrala ključno vlogo pri zdravljenju virusnih okužb, rakavih obolenj in alergijske astme. Pri našem raziskovalnem delu smo izhajali iz strukture znanega agonista TLR7 3-(4-fluorofenil)-4-(3-metilpiperidin-1-il)-6-(trifluorometil)izoksazolo[5,4-d]pirimidina. Z optimiziranim postopkom sinteze in izolacije smo pripravili pet analogov z izoksazolo[5,4-d]pirimidinskim skeletom in dva s kinazolinskim obročem. Vseh sedem končnih spojin smo okarakterizirali, jim določili čistost in jih biološko ovrednotili. Agonistično delovanje smo preverili na celični liniji HEK293, ki izraža človeški TLR7, aktivnim spojinam pa smo določili še citotoksičnost. Na podlagi rezultatov testiranj na celični liniji HEK293smo ugotovili, da kar štiri izmed sedmih končnih spojin izkazujejo agonistično delovanje, tri spojine pa niso bile aktivne. Za doseganje agonističnega delovanja so se kot najbolj optimalne izkazale spojine z izoksazolo[5,4-d]pirimidinskim skeletom, ki so na mestu 4 substituirane s 6-členskim obročem, ta pa z manjšo stransko skupino na mestu 3. Vse štiri aktivne spojine so se v testiranih koncentracijah izkazale za necitotoksične. Izmed vseh končnih spojin se je za najbolj obetavno izkazala spojina 8, saj ima najnižjo vrednost EC50 (EC50 = 11 μM), zaradi česar predstavlja dobro izhodišče za nadaljnje načrtovanje in sintezo agonistov TLR7.

• Language: Slovenian
• Keywords: Tollu podobni receptorji, TLR7, agonist, izoksazolo[5, 4-d]pirimidin, kinazolin
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2024
• PID: 20.500.12556/RUL-158750
• Publication date in RUL: 20.06.2024

Secondary language

• Language: English
• Title: Synthesis of quinazoline and isoxazolo[5,4-d]pyrimidine derivatives with Toll-like receptor 7 agonist activity

Abstract

The inflammatory response represents a defense mechanism that is activated when exposed to harmful factors. The innate and adaptive immune system plays a crucial role in this process, as it is also activated via Toll-like receptors (TLR). To date, 10 types of TLR have been identified. It is known that they recognize small molecules or microbial components as their ligands. The focus of our thesis was on TLR7, which is activated by single-stranded RNA. The activation triggers a cascade of responses leading to anti-inflammatory, antiviral and antitumor effects. Thus, TLR7 agonists could play a crucial role in the treatment of viral infections, cancers and allergic asthma. Our research was based on the structure of the known TLR7 agonist (3-(4-fluorophenyl)-4-(3-methylpiperidin-1-yl)-6-(trifluoromethyl)isoxazolo[5,4-d]pyrimidine). Five analogs with an isoxazolo[5,4-d]pyrimidine ring and two analogs with a quinazoline ring were prepared with the optimized synthetic and isolation procedures. All seven final compounds were characterized and biologically tested. Agonist activity was tested on the HEK293 cell line, expressing human TLR7. All active compounds were then tested for their cytotoxicity. Based on the results from biological evaluation, we found out that four out of the seven final compounds showed agonist activity, while three of them were not active. Compounds with an isoxazolo[5,4-d]pyrimidine scaffold substituted at position 4 by a six-membered ring, with a smaller group at position 3, were found to be the most optimal to obtain agonist activity. All four active compounds were not cytotoxic at any concentration. Out of all final compounds, compound 8 proved to be the most promising, as it has the lowest EC50 value (EC50 = 11 μM). Therefore, it represents a good starting point for further design and synthesis of TLR7 agonists.

• Keywords: Toll-like receptors, TLR7, agonist, isoxazolo[5, 4-d]pyrimidine, quinazoline