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Razvoj zdravila je kompleksen in dolgotrajen proces, s katerim iščemo in razvijamo nove metode zdravil in zdravljenja za različne bolezni. Proces odkrivanja zdravila se običajno prične z določitvijo in ovrednotenjem biološke tarče, čemur sledijo različne tehnike za iskanje spojin zadetkov ter spojin vodnic. To lahko vključuje rešetanje velikih knjižnic kemičnih spojin, virtualno rešetanje ali oblikovanje novih spojin na podlagi strukture tarče. Ena izmed pomembnih tehnik pri odkrivanju zdravilnih učinkovin je masna spektrometrija, ki lahko zagotovi pomembne informacije o potencialnih kandidatih za zdravilne učinkovine. Pomembna tehnika je tudi afinitetna selekcijska masna spektrometrija, ki omogoča določitev majhnih molekul, ki se vežejo na biološko tarčo z visoko afiniteto in s tem omogoča razvoj bolj učinkovitih ter selektivnih zdravil. Klik kemija predstavlja pomembno orodje pri sestavljanju enostavnih molekul v kompleksnejše. Gre za učinkovito, hitro in selektivno sintetično metodo za spajanje molekularnih fragmentov pod blagimi reakcijskimi pogoji. Primer klik reakcije je z bakrom(I) katalizirana azid-alkin cikloadicija, ki smo jo uporabili tudi pri sinteznem delu magistrske naloge. Pri magistrski nalogi smo želeli razviti strategijo priprave knjižnice spojin, ne da bi pri tem izolirali posamezne spojine. Sintezni del naloge smo izvedli v treh delih. V prvem delu smo pripravili nekaj monoazidov in diazidov, v drugem delu smo pripravili nekaj monoalkinov in dialkinov, v zadnjem delu pa smo s klik reakcijo (z bakrom(I) katalizirano azid-alkin cikloadicijo) pripravili večje število 1,2,3-triazolov. Nastanek triazolov smo preverili s tekočinsko kromatografijo, sklopljeno z masno spektrometrijo. Končnim produktom smo določili tališče, na koncu pa smo jih testirali tudi na izoliranem encimu InhA in tako določili njihovo zaviralno aktivnost. Tekom magistrske naloge smo pripravili približno 300 spojin v relativno kratkem času (v dveh mesecih). Rezultati testiranj so pokazali, da nobeden od produktov sintez ni učinkovit zaviralec encima InhA. Uporaba afinitetne selekcijske masne spektrometrije tako ni bila smiselna. Za dosego boljših rezultatov bi v prihodnosti lahko uporabili večje število azidov in alkinov, s čimer bi povečali verjetnost zadetka. Prav tako bi bilo smiselno pripravljene zmesi testirati na več tarčah. Drug development involves a complex and lengthy process aimed at discovering and developing new treatment methods and drugs for diverse diseases. This process encompasses the identification and evaluation of biological targets, as well as techniques for identifying lead compounds. Mass spectrometry plays a vital role in drug discovery by providing essential information about potential drug candidates. In particular, affinity selection mass spectrometry enables the identification of small molecules that bind with high affinity to biological targets, facilitating the development of more effective and selective drugs. Click chemistry, exemplified by the copper(I)-catalyzed azide-alkyne cycloaddition, serves as a valuable tool for assembling simple molecules into more complex ones, offering an efficient, rapid, and selective synthetic method. This Master's thesis aimed to develop a strategy for preparing a compound library without isolating individual compounds. The synthesis process consisted of three parts: the preparation of monoazides and diazides, the synthesis of monoalkynes and dialkynes, and the generation of a larger number of 1,2,3-triazoles through the copper(I)-catalyzed azide-alkyne cycloaddition. The formation of triazoles was confirmed using liquid chromatography coupled with mass spectrometry. Additionally, the melting point of the final products (azides and alkynes) was determined, and their inhibitory activity was assessed by testing them on the isolated InhA enzyme. Therefore, affinity selection mass spectrometry was not used, since there was not any active reaction mixture. Approximately 300 compounds were successfully synthesized within a relatively short timeframe (two months) during the Master's thesis. However, none of the synthesized products exhibited effective inhibitory activity against the InhA enzyme. Future work could involve the use of a larger number of azides and alkynes to increase the likelihood of identifying promising compounds as well as testing the products on other enzymes. |
