Novel Adjuvant Strategies to overcome Antimicrobial Resistance

Antimicrobial resistance (AMR) is a natural phenomenon that occurs when bacteria, viruses, fungi, and parasites develop the ability to survive when exposed to antimicrobials. If no urgent action is taken, the number of people who will die due to AMR will increase up to 10 million each year by 2050. In this scenario, the discovery of new treatments has become as relevant as ever before. Besides the “classical” discovery approach, new strategies may encompass the development of new antibiotic adjuvants, which are chemical entities characterized by weak or absent antibiotic activity but that, co-administered with antibiotics, may boost their efficacy against MDR bacterial strains. Since such drugs would not inhibit any crucial cell functions of the bacteria, another advantage would be the reduction of the selective pressure for the development of resistance. Therefore, the combination between existing drugs and adjuvants could result in a winning strategy in the search of new treatments against infectious diseases difficult to eradicate due to resistance. At this regard, we have focused our attention on unexplored or underexplored “non-essential” targets, in particular metabolic enzymes such as those involved in cysteine biosynthesis. Strong evidence has proved that suppression or reduction of cysteine biosynthesis leads to a decrease in bacterial fitness and a reduction of virulence. In bacteria, cysteine biosynthesis is carried out by the reductive sulfate assimilation pathway (RSAP), which is absent in mammals. The last two enzymes of this pathway are O- acetylserine sulfhydrylase (OASS) and Serine acetyltransferase (SAT). Thus, the aim of this thesis was focused on the design and development of small molecules as potential antimicrobial adjuvants and inhibitors of SAT and OASS. Different approaches were investigated to inhibit cysteine production. Regarding the development of potential SAT inhibitors, the study started from a virtual screening previously conducted in our lab that disclosed two chemical series of hit compounds as potential SAT inhibitors. Further investigation was conducted to establish the Structure-Activity Relationships (SAR) of these hits and evaluate the biological properties towards SAT. In the search of novel hit compounds as OASS inhibitors, two fragment-like derivatives discovered through a scaffold hopping approach were further investigated. The straightforward synthetic route established to synthesize the fragment derivatives allowed us to obtain a variously substituted set of molecules. Two fragment derivatives showed increased activity compared to that of the lead compound and can be considered interesting derivatives for further chemical optimization. Additionally, the evidence reported in the literature regarding the ability of triazole ring to target OASS prompted us to initiate a preliminary investigation on the potential activity of the benzotriazole core (BTA) on the cysteine pathway. BTA derivatives were synthesized and biologically evaluated. The preliminary analysis showed the ability of these derivatives to inhibit the synthesis of cysteine in-vitro and in-cell assays.