Sinteza in biološko vrednotenje halogeniranih piperidinskih zaviralcev proteina Hsp90 z antiproliferativnim delovanjem

Ahmić, Azra (avtor), Tomašič, Tihomir (mentor), Zajec, Živa (komentor)

Rak je v zahodni civilizaciji drugi najpogostejši vzrok smrti. Njegova pojavnost se vsako leto povečuje, a se hkrati umrljivost, kot posledica zgodnjega odkrivanja bolezni in večje uspešnosti zdravljenja, počasi manjša. Rakave celice se v sklopu kancerogeneze spopadajo z več izzivi, med katerimi predstavljajo največjo oviro njihovi napačno zviti lastni proteini (onkoproteini), ki so v normalnih razmerah podvrženi razgradnji. Rak v telesu ustvarja stresne pogoje, kar vodi do aktivacije in indukcije proteinov toplotnega šoka, ki onkoproteine stabilizirajo, jim omogočajo pridobitev funkcionalnosti ter s tem nadaljnji razvoj tumorja. Proteini toplotnega šoka v osnovi skrbijo za proteostazo in doseganje nativne konformacije proteinov klientov, a je njihovo delovanje med kancerogenezo spremenjeno ter izkoriščeno s strani rakavih celic. Mnogi onkoproteini, ključni za maligno transformacijo, so klienti proteina toplotnega šoka 90 (Hsp90), zato predstavlja Hsp90 idealno tarčo za razvoj novih protitumornih učinkovin. V sklopu magistrske naloge smo proučili vpliv halogenih atomov na zaviralno delovanje alosteričnih piperidinskih zaviralcev C-končne domene Hsp90. Izhajali smo iz spojine z zaviralnim delovanjem, pri kateri smo ohranili strukturne elemente pomembne za tvorbo interakcij s Hsp90. Spreminjali smo aromatski del izhodne spojine, na katerega smo vpeljevali različne halogene atome ter tako pripravili štiri nove spojine. Le-te smo biološko ovrednotili na človeški celični liniji raka dojke MCF-7 s testom MTS ((3-(4,5-dimetiltiazol-2-il)-5-(3-karboksimetoksifenil)-2-(4-sulfonil)-2H-tetrazolijeva sol), s pomočjo katerega smo spojinam določili vrednosti IC50. Na podlagi rezultatov smo ovrednotili vpliv halogenov na sposobnost tvorbe interakcij s tarčo, jih primerjali s predhodno sintetiziranimi spojinami ter določili odnos med strukturo in delovanjem. Rezultati testiranja so pokazali, da je substitucija aromata levega dela molekule na mestih meta in para ključna za zaviralno delovanje v nizkih mikromolarnih koncentracijah. Spojina 17 je imela najnižjo vrednost IC50 (IC50 = 5,58 ± 0,75 µM) ter je tako izkazovala najmočnejše delovanje. V primerjavi z izhodno spojino TZZ-11 imata spojini 15 in 17 močnejše delovanje, spojini 19 in 11 pa delujeta šibkeje, kar je vodilo do ugotovitve, da sta kot aromatska substituenta levega dela molekule najboljša halogena atoma klor ter brom, ki po naših predvidevanjih zaradi svoje velikosti ter sposobnosti tvorbe ustrezno močne halogene vezi tvorita najmočnejše interakcije s tarčo.

• Jezik: Slovenski jezik
• Ključne besede: rak, molekularni šaperoni, protein toplotnega šoka, Hsp90, piperidin, zaviralec
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2022
• PID: 20.500.12556/RUL-137357
• Datum objave v RUL: 14.06.2022

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Synthesis and biological evaluation of halogenated piperidin-based Hsp90 inhibitors with antiproliferative activity

Izvleček

Cancer is the second leading cause of death in Western civilisation. Its incidence is higher every year but due to early detection of the disease and more successful treatment options, its mortality rate is decreasing. Cancer cells face many challenges during their development in the human body. One of the biggest obstacles are misfolded proteins (oncoproteins) that are normally subject to degradation. The formation of cancer cells (carcinogenesis) creates a stressful environment leading to the activation and induction of heat shock proteins, which promote cancer development and growth by stabilizing oncoproteins, allowing them to reach their full potential. Heat shock proteins are fundamentally responsible for proteostasis and client protein folding but during carcinogenesis their mechanism of action is altered and abundantly utilised by cancer cells. Many oncoproteins responsible for cancer development interact with heat shock protein 90 (Hsp90). Hsp90 therefore represents a promising target for the development of new cytotoxic agents. In this master's thesis, we investigated the influence of halogen atoms on the inhibitory potential of new allosteric piperidine-based inhibitors of the Hsp90 C-terminal domain. The basis of our research was a compound with known inhibitory properties. We retained the structural elements responsible for the interaction with Hsp90 and focused solely on the modification of the aromatic ring on the left side by introducing different halogen atoms. We prepared four new compounds, which we biologicaly evaluated with the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) metabolic assay on MCF-7 human breast cancer cells and determined their IC50 values. This provided insightful information, which helped evaluate the ability of the halogens to form halogen bonds with the target, compare our compounds with other Hsp90 inhibitors and establish a structure-activity relationship. The test results led to the conclusion that substitution of the aromatic ring in both meta and para positions is critical for inhibition in the low micromolar range. Compound 17 had the lowest IC50 value (IC50 = 5.58 ± 0.75 µM) and is therefore the most potent inhibitor. Compared to compound TZZ-11, which served as the reference compound, both compounds 15 and 17 were found to be more potent, while compounds 19 and 11 were not. This led to the conclusion that chlorine and bromine atoms are the best aromatic substituents. This is due to their atom size, which enables them to form the strongest interactions with the target.

• Ključne besede: cancer, molecular chaperones, heat shock protein, Hsp90, piperidine, inhibitor