Sinteza analogov 2-(3-(4-fluorofenil)-4-oksoizoksazolo[5,4-d]pirimidin-5(4H)-il)-N-fenilacetamida kot zaviralcev indolamin 2,3-dioksigenaze 1

Kožuh, Eva (author), Sova, Matej (mentor)

Človeški imunski sistem je kompleksen sistem različnih celic, tkiv in organov. Zadolžen je za odgovor na različne notranje ali zunanje grožnje, ena izmed njih predstavlja razvoj rakavih celic. Na transformacijo normalnih celic v rakave lahko vpliva več dejavnikov, tako zunanjih kot notranjih, pomembno vlogo pri širjenju raka pa ima tudi znotrajcelični citosolni encim indolamin 2,3-dioksigenaza 1 (IDO1), ki sodeluje v kinureninski razgradni poti (KP) esencialne aminokisline triptofan (Trp). Ekspresija IDO1 v celicah tako vpliva na razmerje med koncentracijo Trp in metabolitov KP, ki imajo imunosupresivno delovanje in zavirajo odziv imunskega sistema na rakave celice. Število na novo odkritih zaviralcev IDO1 se povečuje, saj velika farmacevtska podjetja vlagajo v njihov razvoj, veliko od njih se jih je znašlo že v kliničnih raziskavah. V okviru magistrske naloge smo sintetizirali pet končnih spojin, ki smo jim ovrednotili zaviralno delovanje na IDO1. Vseh pet spojin ima enak skelet, ki temelji na zadetku iz virtualnega rešetanja 3-(4-fluorofenil)izooksazolo[5,4-d]pirimidin- 4(5H)-onu. Spojine smo analitsko ovrednotili in zaviralno aktivnost preverili s pomočjo biokemijskega testiranja na osnovi merjenja fluorescence. Od sintetiziranih spojin, ki se med seboj razlikujejo v substituciji na fenilnem obroču, sta le dve izkazovali signifikantno zaviralno delovanje. Spojinam 12 in 13 smo določili in izračunali vrednost IC50 v nizkem mikromolarnem območju, zato lahko sklepamo, da se cikloheksilni obroč ali trifluorometilna skupina, ki sta vezani na fenilacetamidni fragment, najbolje prilegata v žep B v aktivnem mestu encima IDO1, kar smo potrdili tudi z molekulskim sidranjem spojine 12 v kristalno strukturo encima IDO1. Z ustrezno optimizacijo predstavlja spojina 12 dobro izhodišče za razvoj močnega zaviralca encima IDO1, ki bi ga lahko v prihodnosti uporabljali kot protitumorno učinkovino.

• Language: Slovenian
• Keywords: indolamin 2, 3-dioksigenaza 1 (IDO1), zaviralci, rak, imunski sistem, kinureninska razgradna pot, triptofan
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2020
• PID: 20.500.12556/RUL-119871
• Publication date in RUL: 12.09.2020

Secondary language

• Language: English
• Title: Synthesis of 2-(3-(4-fluorophenyl)-4-oxoisoxazolo[5,4-d]pyrimidin-5(4H)-yl)-N-phenylacetamide analogs as indoleamine 2,3-dioxygenase 1 inhibitors

Abstract

Human immune system is a complex system of different cells, tissues and organs. It responds appropriately to internal or external threats, one of which is the development of cancer cells. Many internal and external factors can affect the transformation of a normal cell to a cancerous one and among them is an intracellular cytosolic enzyme indoleamine 2,3-dioxygenase 1 (IDO1) has an important role in cancer metastasis. IDO1 is involved in the degradation pathway of the essential amino acid tryptophan (Trp), known as Kynurenine pathway (KP). IDO1 expression in cells regulates the ratio between Trp concentration and concentration of metabolites of the KP, which have immunosuppressant activity, thus blocking the immune system response to cancer cells. The number of newly discovered IDO1 inhibitors is increasing every day since big pharma companies are investing in their development, and many of them are already present in clinical trials. In our Master thesis we synthesized 5 final compounds, which were evaluated for their inhibitory activity. All five synthesized compounds possess the same scaffold, which is based on a virtual screening hit compound 3-(4- fluorophenyl)[1,2]isoxazolo[5,4-d]pyrimidin-4(5H)-on. All compounds were confirmed by different analytical methods and their inhibitory activity was evaluated by biochemical assay based on fluorescence measurement. Among synthesized compounds, which differ in substitution on the phenyl ring, only two showed significant inhibitory activity. IC50 values for compounds 12 in 13 was determined in low micromolar range, thus we could assume that cyclohexyl ring or trifluoromethyl group fits into the pocket B in the enzyme’s active site, which was confirmed by molecular docking of compound 12 into the crystal structure of IDO1.With appropriate optimization compound 12 represents the important starting point for the development of potent IDO1 inhibitor, which could be used as potential antitumor agent.

• Keywords: indoleamine 2, 3-dioxygenase 1 (IDO1), inhibitors, cancer, immune system, kynurenine pathway, tryptophan