Sinteza in vrednotenje N-(2-benziloksi)fenilpirolamidnih zaviralcev proteina toplotnega šoka 90

Sopčič, Ema (avtor), Zidar, Nace (mentor), Dernovšek, Jaka (komentor)

Število novih primerov raka in umrljivost zaradi raka se v Sloveniji v zadnjih letih povečuje. Ker lahko s pravočasnim začetkom in pravilno izbiro zdravljenja rakavih obolenj bolnikom omogočimo ozdravitev ali podaljšanje časa do napredovanja bolezni, potekajo številne raziskave za odkrivanje novih zdravilnih učinkovin, ki zavirajo nenormalno in nenadzorovano rast rakavih celic. Predmet teh raziskav so tudi zaviralci proteina toplotnega šoka 90 (Hsp90), saj so proteini toplotnega šoka v rakavih celicah bolj izraženi kot v zdravih celicah in jim tako omogočijo preživetje v neugodnih pogojih tumorskega mikrookolja. Z zaviranjem Hsp90, preko vpliva na proteine, ki so od tega šaperona odvisni, hkrati zavremo več različnih signalnih poti in tako zavremo rast rakavih celic. V okviru magistrske naloge smo sintetizirali in ovrednotili serijo potencialnih zaviralcev C-končne domene Hsp90. Pri sintezi naših spojin smo izhajali iz spojin, ki so bile predhodno pripravljene na Katedri za farmacevtsko kemijo Fakultete za farmacijo Univerze v Ljubljani in so poleg delovanja na DNA-topoizomerazo IIα pri testiranju in vitro izkazale tudi delovanje na proteine toplotnega šoka Hsp90. Naše spojine so imele N-(2-benziloksi)fenilpirolamidno osnovno strukturo. Vse pripravljene spojine iz naše serije smo fizikalno-kemijsko ovrednotili. Pri tem smo med drugim uporabili jedrsko magnetno resonanco, infrardečo spektroskopijo, masno spektrometrijo in tekočinsko kromatografijo visoke ločljivosti. Spojine smo na celični liniji raka dojke MCF-7 ovrednotili tudi biološko. V sklopu biološkega testiranja na celični liniji raka dojke MCF-7 smo končnim spojinam 8, 10, 12, 14, 16, 23 in 24 določili aktivnost s testom MTS in jo podali v obliki srednje zaviralne koncentracije (IC50). Večina testiranih spojin je izkazala močno zaviralno delovanje na rast/proliferacijo celične linije raka dojke MCF-7 z IC50 vrednostmi v nizkem mikromolarnem oz. visokem nanomolarnem območju. Najboljšo zaviralno aktivnost je izkazala spojina 23, z IC50 vrednostjo 0,85 μM. Sledili sta ji spojini 24 in 14 z IC50 vrednostma 0,92 μM in 0,94 μM. Rezultati biološkega testiranja so pokazali, da imajo nekoliko bolj fleksibilne molekule boljše zaviralno delovanje, saj je v teh molekulah položaj bazičnega centra najverjetneje bolj ugoden za vezavo na C-končno domeno Hsp90 kot pri bolj rigidnih molekulah. V splošnem so se novi N-(2-benziloksi)fenilpirolamidi, ki smo jih pripravili v okviru magistrske naloge, izkazali kot spojine z obetavnim delovanjem na celično linijo raka dojke MCF-7.

• Jezik: Slovenski jezik
• Ključne besede: rak, Hsp90, zaviralec, C-končna domena Hsp90, N-(2-benziloksi)fenilpirolamid
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2022
• PID: 20.500.12556/RUL-142951
• Datum objave v RUL: 06.12.2022

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Synthesis and evaluation of N-(2-benzyloxy)phenylpyrrolamide inhibitors of heat shock protein 90

Izvleček

The number of new cancer cases and cancer mortality have increased in Slovenia in recent years. With the early treatment and proper selection of cancer therapy, patients can be cured or the time to progression of the disease can be prolonged. Therefore, many researchers are looking for new agents that inhibit the abnormal and uncontrolled growth of cancer cells. Inhibitors of heat shock protein 90 are being intensively investigated, because heat shock proteins are more highly expressed in cancer cells than in normal cells and enable them to survive in adverse conditions of the tumor microenvironment. Inhibition of the heat shock protein 90 downregulate several signalling pathways simultaneously by acting on proteins that depend on this chaperone and thus reduces the growth of cancer cells. In this work, we synthesised and evaluated a series of potential inhibitors of the C-terminal domain of Hsp90. We started from compounds previously prepared at the Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ljubljana, which have shown activity on Hsp90 in in vitro assays in addition to their effect on DNA topoisomerase Iiα. All of our compounds had a N-(2-benzyloxy)phenylpyrrolamide basic structure. All compounds were investigated physicochemically using nuclear magnetic resonance spectroscopy, infrared spectroscopy, mass spectrometry, and high-resolution liquid chromatography. The compounds were also evaluated biologically using the MCF-7 breast cancer cell line. As a part of the biological testing, compounds 8, 10, 12, 14, 16, 23, and 24 were evaluated on the MCF-7 breast cancer cells using the MTS assay. Activity was expressed as mean inhibitory concentration (IC50). Most of the tested compounds showed potent inhibitory effects on the MCF-7 cell viability with IC50 values in the low micromolar or high nanomolar range. Compound 23 showed the best inhibitory activity with a mean inhibitory concentration of 0.85 μM. This was followed by compounds 24 and 14 with mean inhibitory concentrations of 0.92 μM and 0.94 μM, respectively. The results of biological testing showed that slightly more flexible molecules have a better inhibitory effect because the position of the basic centre in these molecules is more favourable for binding to the C-terminal domain of Hsp90 than in more rigid molecules. Overall, the new N-(2-benzyloxy)phenylpyrolamides prepared in the master’s thesis have shown promising effect on the viability of MCF-7 breast cancer cell line.

• Ključne besede: cancer, Hsp90, inhibitor, Hsp90 C-terminal domain, N-(2-benzyloxy)phenylpyrrolamide