The discovery of penicillin began the age of antibiotics, which was a turning point in human healthcare. However, to this day, microbial infections are still a concern throughout the world, and the rise of multidrug-resistant organisms is an increasing challenge. To combat this threat, diagnostic imaging tools could be used to verify the causative organism and curb inappropriate use of antimicrobial drugs. Nuclear imaging offers the sensitivity needed to detect small numbers of bacteria in situ. Among nuclear imaging tools, radiolabeled antibiotics traditionally have lacked the sensitivity or specificity necessary to diagnose bacterial infections accurately. One reason for the lack of success is that the antibiotics were often chelated to a radiometal. This was done without addressing the ramifications of how the radiolabeling would impact probe entry to the bacterial cell, or the mechanism of binding to an intracellular target. In this review, we approach bacterial infection imaging through the lens of bacterial specific molecular targets, their intracellular or extracellular location, and discuss radiochemistry strategies to guide future probe development.

Temporal control of protein levels in cells and living animals can be used to improve our understanding of protein function. In addition, control of engineered proteins could be used in therapeutic applications. PRoteolysis-TArgeting Chimeras (PROTACs) have emerged as a small-molecule-driven strategy to achieve rapid, post-translational regulation of protein abundance via recruitment of an E3 ligase to the target protein of interest. Here, we develop several PROTAC molecules by covalently linking the antibiotic trimethoprim (TMP) to pomalidomide, a ligand for the E3 ligase, Cereblon. These molecules induce degradation of proteins of interest (POIs) genetically fused to a small protein domain, E. coli dihydrofolate reductase (eDHFR), the molecular target of TMP. We show that various eDHFR-tagged proteins can be robustly degraded to 95% of maximum expression with PROTAC molecule 7c. Moreover, TMP-based PROTACs minimally affect the expression of immunomodulatory imide drug (IMiD)-sensitive neosubstrates using proteomic and biochemical assays. Finally, we show multiplexed regulation with another known degron-PROTAC pair, as well as reversible protein regulation in a rodent model of metastatic cancer, demonstrating the formidable strength of this system. Altogether, TMP PROTACs are a robust approach for selective and reversible degradation of eDHFR-tagged proteins in vitro and in vivo.

The rational design of foldable and functionalizable peptidomimetic scaffolds requires the concerted application of both computational and experimental methods. Recently, a new class of designed peptoid macrocycle incorporating spiroligomer proline mimics (Q-prolines) has been found to pre-organize when bound by monovalent metal cations. To determine the solution-state structure of these cation-bound macrocycles, we employ a Bayesian inference method (BICePs) to reconcile enhanced-sampling molecular simulations with sparse ROESY correlations from experimental NMR studies. The BICePs approach circumvents the need for bespoke force field parameterization, instead relying on experimental restraints to help narrow the possible set of cis/trans amide isomers in solution. Conformations predicted to be most populated in solution were then simulated in the presence of explicit cations to yield trajectories with observed binding events, revealing a highly-preorganized all-trans amide conformation, whose formation is likely limited by the slow rate of cis/trans isomerization. Interestingly, this conformation differs from a racemic crystal structure solved in the absence of cation. Free energies of cation binding computed from distance-dependent potentials of mean force suggest Na+ has higher affinity to the macrocycle than K+, with both cations binding much more strongly in acetonitrile than water. The simulated affinities are able to correctly rank the extent to which different macrocycle sequences exhibit preorganization in the presence of different metal cations and solvents, suggesting our approach is suitable for solution-state computational design.