A kinase-cGAS cascade to synthesize a therapeutic STING activator

The introduction of molecular complexity in an atom- and step-efficient manner remains an outstandinggoal in modern synthetic chemistry. Artificial biosynthetic pathways are uniquely able to address this challenge by using enzymesto carry out multiple synthetic steps simultaneously or in a one-pot sequence(1-3). Conducting biosynthesis exvivo further broadens its applicability by avoiding cross-talk with cellular metabolism and enabling the redesign of key biosynthetic pathwaysthrough the use of non-natural cofactors and synthetic reagents(4,5). Here we describe the discovery and construction of an enzymatic cascade to MK-1454, a highly potent stimulator of interferon genes (STING) activator under study as an immuno-oncology therapeutic(6,7) (ClinicalTrials.gov study NCT04220866). From two non-natural nucleotide monothiophosphates, MK-1454 is assembled diastereoselectively in a one-pot cascade, in which two thiotriphosphate nucleotides are simultaneously generated biocatalytically, followed by coupling and cyclization catalysed by an engineered animal cyclicguanosine-adenosine synthase (cGAS). For the thiotriphosphate synthesis, three kinase enzymes were engineered to develop a non-natural cofactor recycling system in which one thiotriphosphate serves as a cofactor in its own synthesis. This study demonstrates the substantial capacity that currently existsto use biosynthetic approachesto discover and manufacture complex, non-natural molecules.

Automated summary

Introducing molecular complexity in a efficient manner is a challenging goal in modern synthetic chemistry. Artificial biosynthetic pathways utilizing enzymes have the ability to carry out multiple synthetic steps simultaneously, enabling the discovery and production of complex, non-natural molecules. In this study, an enzymatic cascade was discovered and constructed for the synthesis of MK-1454, a potential immuno-oncology therapeutic. By engineering the enzymes involved, multiple steps of the synthesis were successfully carried out in a one-pot reaction.