Prospects and limitations of paramagnetic chemical exchange saturation transfer agents serving as biological reporters in vivo

The concept of using paramagnetic metal ion complexes as chemical exchange saturation transfer agents (paraCEST) for molecular imaging of various biological processes first appeared in the literature about 20 years ago. The first paraCEST agent was based on a highly shifted, inner‐sphere, slowly exchanging water molecule that could be activated at a frequency far away from bulk water, a substantial advantage for selective activation of the agent alone. Many other paraCEST agent designs followed that were based on activation of exchanging ‐NH or ‐OH proton on the chelate itself. Both types of paraCEST designs are attractive for molecular imaging because the rates of water molecule or ligand proton exchange can be designed to be sensitive to a biological or physiological property such as pH, enzyme activity, or redox. Hence, the intensity or frequency of the resulting CEST signal provides a direct readout of that property. Many molecular designs have appeared in the literature over the past 20 years, mostly reported as proof‐of‐concept designs but, unfortunately, only a few reports have explored the limitations of paraCEST agents for imaging a biological process in vivo. As a community, we now know that the sensitivity of paraCEST agents is lower than one might anticipate based upon simple chemical exchange principles and, in general, it appears the sensitivity of paraCEST agents is even lower in vivo than in vitro. In this short review, we address some of the factors that contribute to the limited sensitivity of paraCEST agents in vivo, offer some thoughts on approaches that could lead to dramatically improved paraCEST sensitivity, and challenge the scientific community to perform more in vivo experiments designed to test these ideas.