Synthesis of piperidine-based C-terminal domain inhibitors of Hsp90 with anticancer activity

Rakava obolenja predstavljajo zelo pomemben javnozdravstveni, socialni in ekonomski problem, saj skupaj s kardiovaskularnimi boleznimi sodijo med najpogostejše vzroke smrti na svetu. Tako ne preseneča, da je eno glavnih področij, s katerim se dandanes ukvarjajo raziskovalci, odkrivanje novih učinkovin s protitumornim delovanjem. Protein toplotnega šoka Hsp90 je 90 kDa velik molekulski šaperon, sestavljen iz treh strukturnih domen: N-končne domene (NKD), srednje domene in C-končne domene (CKD). Pomembno vlogo ima pri celični homeostazi, sodeluje pa tudi pri pravilnem zvitju ter doseganju pravilne konformacije in s tem aktivacije več sto proteinov. Številni izmed le-teh so onkoproteini, ki so pomembni za doseganje za rakavo celico značilnih lastnosti. Hsp90 pomaga rakavim celicam premagati številna stresna stanja, njegova ekspresija pa je v rakavih celicah kar do 10-krat višja kot v normalnih celicah. Prvi odkriti zaviralci Hsp90 so bili zaviralci N-končne domene Hsp90, katerih glavna slabost je bila sprožitev odziva toplotnega šoka. Kmalu za tem je sledilo pomembno odkritje zaviralcev C-končne domene. Ker slednji niso privedli do odziva toplotnega šoka, je C-končna domena postala pomembna tarča v razvoju protitumornih učinkovin. V okviru magistrske naloge smo sintetizirali enajst potencialnih novih zaviralcev C-končne domene Hsp90. Izvedli smo modifikacijo levega in desnega dela referenčne spojine, medtem ko smo osrednji del v vlogi distančnika med aromatom na levi in bazičnim centrom na desni ohranili. Na skrajnem levem delu molekule smo proučili vpliv usmerjenosti amidne vezi ter vpliv vezave različnih substituentov na različna mesta na fenilu. Raziskali smo vpliv uvedbe piperidinskega skeleta na desni ter vpliv razdalje med bazičnim centrom na piperidinu in aromatom. Končne spojine smo s testom metabolne aktivnosti ovrednotili na celicah raka dojk MCF-7 ter na celicah limfomov U2932, SU-DHL-2, SU-DHL-10, WSU-DLCL2 in VAL. Vseh enajst spojin je na celicah MCF-7 izkazalo bistveno močnejše citotoksično delovanje v primerjavi z referenčno spojino, večina spojin tudi na celicah U2932, SU-DHL-2 in VAL, pri čemer pa smo na citotoksično delovanje spojin sklepali na osnovi učinka spojin na metabolno aktivnost celic. Na celicah SU-DHL-10 in WSU-DLCL2 so spojine izkazale šibkejše citotoksično delovanje napram referenčni spojini. Rezultati magistrske naloge nam tako dajejo pomembne informacije o odnosu med strukturo in delovanjem ter predstavljajo osnovo za nadaljnje raziskovanje.Cancer is a very important public health, social and economic problem, as it is one of the most common causes of death in the world, along with cardiovascular diseases. Thus, it is not surprising that one of the main research areas is the discovery of new active substances with anticancer activity. Heat shock protein Hsp90 is a 90 kDa molecular chaperone, which consists of three structural domains: the N-terminal domain (NTD), the middle domain and the C-terminal domain (CTD). The Hsp90 has an important role in cellular homeostasis, but it also participates in the correct protein folding to achieve bioactive conformation and thus the activation of hundreds of proteins. Many of these are oncoproteins, which are important for cancer cell-specific properties. Hsp90 helps cancer cells overcome many stressful situations and its expression in cancer cells is up to 10 times higher than in normal cells. The first Hsp90 inhibitors discovered were N-terminal domain inhibitors, the main disadvantage of which was induction of the heat shock response. This was soon followed by the important discovery of the allosteric C-terminal domain inhibitors. Since the latter did not induce the heat shock response, the C-terminal domain of Hsp90 became an important target in the development of anticancer drugs. In the context of this master’s thesis, we synthesized eleven potential new Hsp90 C-terminal domain inhibitors. We performed modification of the left and right parts of the reference compound, while the central part, acting as a spacer between the aromatic ring on the left and the basic center on the right, was retained. On the extreme left part of the molecule, we studied the influence of the orientation of the amide bond and the influence of the binding of various substituents to different sites on the phenyl ring. Moreover, we examined the influence of the introduction of the piperidine scaffold in the right part of the molecule and the influence of the distance between the basic center on the piperidine ring and the aromatic ring. The final compounds were evaluated with measuring of metabolic activity in MCF-7 breast cancer cells and in U2932, SU-DHL-2, SU-DHL-10, WSU-DLCL2 and VAL lymphoma cells. Compared to the reference compound, all synthesized compounds showed significantly stronger cytotoxic activity in MCF-7 cells, most of them also in U2932, SU-DHL-2 and VAL cells. Moreover, synthesized compounds showed weaker cytotoxic activity in SU-DHL-10 and WSU-DLCL2 cells compared to the reference. Therefore, the results of the master\u27s thesis provide us with important information about the structure-activity relationship and represent the basis for further optimization of Hsp90 C-terminal domain inhibitors