Optimizacija sinteze liganda za ubikvitin-ligazo E3 von Hippel-Lindau kot bistvenega gradnika himernih razgrajevalcev

Čižmek, Ula (avtor), Sosič, Izidor (mentor)

Klasični pristopi v farmacevtski kemiji so postavljeni pred izziv, saj približno 80 % človeškega proteoma s tovrstnimi strategijami ni mogoče ciljati, kar omejuje njihovo uporabnost pri zdravljenju številnih bolezenskih stanj. Posledično se je pojavila potreba po novejših pristopih, med katere spadajo tudi himerni razgrajevalci oziroma molekule PROTAC (angl. proteolysis-targeting chimeras). Molekule PROTAC so spojine, ki temeljijo na farmakološkemu modelu dogodka (angl. event-driven pharmacology), saj inducirajo proteolitično razgradnjo tarčnih proteinov in ne le njihovega zaviranja. Za doseganje razgradnje izkoriščajo v celicah prisoten ubikvitin-proteasomski sistem. Ta pristop je že pokazal številne prednosti v primerjavi s klasičnimi majhnimi molekulami, izmed katerih velja izpostaviti katalitično naravo delovanja, saj se lahko po uspešni razgradnji tarče molekula PROTAC odcepi z vezavnega mesta in svoj cikel razgradnje ponovi na naslednji molekuli tarče. Posledično so tovrstne spojine učinkovite pri sub-stehiometričnih količinah zaradi česar ni potrebe po velikih odmerkih učinkovine, kar vodi v manjše tveganje za stranske in neželene učinke. Prav tako imajo zaradi razgradnje tarče in ne le njenega zaviranja dolgotrajen farmakodinamski učinek, vežejo pa se lahko v kateri koli žep proteina, s čimer se poveča število dostopnih farmakoloških tarč. Himerni razgrajevalci so sestavljeni iz treh delov: liganda, ki se veže na tarčni protein, liganda za izbrano ubikvitin ligazo E3 in distančnika, ki ju povezuje. Večina znanih molekul PROTAC temelji na uporabi ligaz E3 Von Hippel-Lindau (VHL), cereblona (CRBN), zaviralca apoptoze (IAP) in »mouse double minute 2 homologue« (MDM2). V tem magistrskem delu smo optimizirali predhodno opisano sintezno pot dveh ligandov za VHL ligazo E3, saj je razvoj učinkovitega postopka sinteze pomemben korak ne le za nadaljnji razvoj himernih razgrajevalcev, temveč tudi za njihovo potencialno industrijsko uporabnost. Pri sintezi prvega liganda je bil primarni namen poleg poenostavitve sinteze in izboljšanja izkoristkov tudi sinteza večje količine benzilaminskega gradnika, ki je zelo uporaben pri sintezi novih in že uveljavljenih ligandov za VHL. Optimizirano sintezo prvega liganda smo nato aplicirali na sintezo strukturno podobnega liganda, ki ima večjo afiniteto do VHL. Optimizacija je pri sintezi prvega liganda uspela, saj smo v delu reakcije, ki je predstavljal največjo težavo, izkoristek izboljšali z 28 % na 41 %. Prav tako smo po aplikaciji optimizirane linearne sinteze na sintezo strukturno podobnega liganda dobili primerljive izkoristke z že znano konvergentno sintezo, s čimer smo nedvoumno pokazali optimizacijo sintezne poti.

• Jezik: Slovenski jezik
• Ključne besede: himerni razgrajevalci, ubikvitin-proteasomski sistem, von Hippel-Lindau, razgradnja proteinov, optimizacija sinteze
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2022
• PID: 20.500.12556/RUL-136246
• Datum objave v RUL: 21.04.2022

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Optimization of synthesis of ligand for the von Hippel-Lindau E3 ubiquitin ligase as an essential building block of proteolysis-targeting chimeras

Izvleček

Traditional approaches in pharmaceutical chemistry are facing a challenge as about 80 % of the human proteome is inaccessible with traditional small-molecule approaches, which limits their applicability in many diseases. Therefore, there is a need for novel approaches and medicinal chemistry modalities, such as targeted protein degradation with proteolysis-targeting chimeras (PROTACs). PROTACs are compounds that rely on the event-driven pharmacology model as they induce proteolytic degradation of target proteins by hijacking the naturally occurring ubiquitin-proteasome system. This approach has already shown a number of advantages, such as catalytic mode of action. Namely, after successful degradation of the targeted protein, PROTAC can undergo several cycles of ternary complex formation and protein degradation. PROTACs are effective at sub-stoichiometric quantities which results in lower concentrations needed for physiological effect. This fact translates to a lower risk of side effects. PROTACs also have a longer lasting pharmacodynamic effect due to the degradation of the target, rather than merely inhibiting it. They can also bind to any pocket of the protein, thus increasing the number of pharmacologically relevant targets that can be tackled with this methodology. PROTACs are composed of three elements: a ligand that binds to the target protein, a ligand for the selected ubiquitin ligase E3 and a linker that connects the two moieties. Most PROTACs are based on Von Hippel-Lindau (VHL) E3 ligase, cereblon (CRBN), inhibitor of apoptosis (IAP), and mouse double minute 2 homologue (MDM2). In this master's thesis we attempted to optimize the previously described synthetic pathway of two ligands for VHL as the development of an efficient synthesis process is an important step not only for further development of PROTACs but also for their potential industrial utility. In addition to simplification and yield increase, the primary purpose of the synthesis of the first ligand was also the preparation of multigram quantities of the benzylamine building block, which is a very useful precursor in the synthesis of VHL ligands. The optimized synthesis of first ligand was then applied to the second ligand, which is structurally similar, but has a higher affinity for VHL. Optimization was successful in the synthesis of first ligand, as in the part of the reaction that represented the greatest problem, the yield was improved from 28 % to 41 %. Furthermore, applying optimized linear synthesis to the synthesis of structurally similar ligand showed comparable yields to described convergent synthesis, thus further showcasing the importance of our work.

• Ključne besede: proteolysis-targeting chimeras, ubiquitin-proteasome system, von Hippel-Lindau, protein degradation, optimization of synthesis