Small molecule inhibitors of cyclic di-AMP signaling
SINTIM, HERMAN O
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Globally, it is estimated that more than 700,000 people die annually from infections caused by drug-resistant bacterial pathogens. Resistant strains of bacteria continue to be isolated in healthcare and community settings. At the same time, the antibiotic pipeline remains dry – exemplified by the paucity of new antibiotics introduced into clinical use. Consequently, antibiotic-resistant strains are rapidly spreading, and antibiotic-resistant infections persist. Additionally, the existing antibiotics target one of the common targets – DNA, RNA, protein and cell wall synthesis. There is an apparent need to identify antibacterial agents against novel targets to slow down the generation of resistance. Cyclic dinucleotides have emerged as central regulators of bacterial physiology. Particularly, cyclic di-AMP (c-di-AMP) regulates cell wall homeostasis, cell size, potassium ion transport, virulence and biofilm formation in various Gram-positive pathogens including Staphylococcus aureus, Enterococcus faecalis, Listeria monocytogenes and Streptococcus pneumoniae. It has been demonstrated that under standard laboratory conditions, deletion of the diadenylate cyclase genes that encode c-di-AMP synthesizing enzymes (diadenylate cyclase, DAC) was lethal in human pathogens like S. aureus and L. monocytogenes. Hence, DACs have been suggested as potential antibiotic targets. Thus far, the effect of c-di-AMP on bacterial physiology has been studied using genetic approaches whereby the key players of the second messenger signaling are deleted, inactivated or overexpressed to create conditions of varying intracellular c-di-AMP levels. However, these approaches are not amenable to drug development. Cell permeable small molecule modulator or c-di-AMP levels are required to validate the druggability of c-di-AMP signaling. This dissertation reports the identification of different small molecules that potently inhibit c-di-AMP synthesis. The cell permeable inhibitors possess the ability to decrease the intracellular concentration of c-di-AMP. Furthermore, the antibacterial activities of the cell permeable c-di-AMP synthesis inhibitors have been characterized. Efforts towards the development of antibiotics have also been discussed.