Tuberculosis (TB) continues to remain the world´s leading cause of death among infectious diseases, especially in economically less developed countries. Mycobacterium tuberculosis is the main causative agent of the disease since humans represent its only natural reservoir. Isoniazid is the first drug that was discovered to be effective against M. tuberculosis. It works as an inhibitor of InhA, an enzyme involved in the biosynthesis of mycolic acids, which are an important part of the mycobacterium cell wall. In combination with other first-line drugs, isoniazid (INH) is still part of the standard treatment therapy. The appearance of multi-drug resistant strains badly impacts the effectiveness of standard treatment regimes. However, it is also a driving force for new drug discovery. INH is a prodrug, that requires in vivo activation by the enzyme KatG, which converts it into the active form. In most of the INH-resistant strains, the resistance is caused by KatG mutation. Currently, drug discovery is focused on the development of direct inhibitors of InhA, that do not require prior activation for their activity.
In this master´s thesis, four analogues of the InhA direct inhibitor have been synthesised. Their design was based on the structure of the lead compound GSK, discovered by GlaxoSmithKline using high-throughput screening method. Our main purpose was to replace the methylthiazole group in the GSK compound with other heterocycles. We used the cooper(I)-catalyzed alkyne–azide Huisgen cycloaddition to combine pre-synthesized alkyne compounds with the 3-(azidomethyl)-1-(2,6-difluorobenzyl)-1H-pyrazole.
The inhibitory activity of the final compounds was determined by an inhibition assay, however none of the tested compounds exhibited and inhibitory activity towards InhA enzyme. Based on the structure comparison we have concluded that the amine group between pyrazole and thiadiazole in the GSK structure is likely crucial for the activity. The amine group forms important interactions in the enzyme active site, most importantly it enables the difluorophenyl and pyrazole heterocycles of the inhibitor to be properly placed in the hydrophobic pocket. Due to the synthetic pathway of the cooper(I)-catalyzed alkyne – azide Huisgen cycloaddition the critical amine group could not have been preserved in the structure of the newly-synthesized analogues.
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