LAUSR

At the beginning of the millennium, the first Chemical Exchange Saturation Transfer (CEST) contrast agents were bio-organic molecules. However, later, metal based CEST agents (paraCEST agents) took center stage. This didn't last too long as paraCEST agents showed limited translational potential. In contrast, the CEST field gradually became dominated by metal free CEST agents. One branch of research stemming from the original work by van Zijl and colleagues1 is the development of CEST agents based on polypeptides. Indeed, in the past two decades tremendous progress was achieved in this field. This includes the design of novel peptides as biosensors, genetically encoded recombinant as well as synthetic reporters. This was a result of extensive characterization and elucidation of the theoretical requirements for rational designing and engineering of such agents. Here we will give an extensive overview of the evolution of more precise protein based CEST agents, review the rationalization of enzyme-substrate pairs as CEST contrast enhancers, discuss the theoretical considerations to improve peptide selectivity, specificity and enhance CEST contrast. Moreover, we will discuss the strong influence of synthetic biology on the development of the next generation of protein based CEST contrast agents.