Photoacoustic tomography has emerged as a promising alternative to MRI and X-ray scans in the clinical setting due to its ability to afford high-resolution images at depths in the cm range. However, its utility has not been established in the basic research arena owing to a lack of analyte-specific photoacoustic probes. To this end, we have developed acoustogenic probes for copper(II)-1 and -2 (APC-1 and APC-2, a water-soluble congener) for the chemoselective visualization of Cu(II), a metal ion which plays a crucial role in chronic neurological disorders such as Alzheimer's disease. To detect Cu(II), we have equipped both APCs with a 2-picolinic ester sensing module that is readily hydrolyzed in the presence of Cu(II) but not by other divalent metal ions. Additionally, we designed APC-1 and APC-2 explicitly for ratiometric photoacoustic imaging by using an aza-BODIPY dye scaffold exhibiting two spectrally resolved NIR absorbance bands which correspond to the 2-picolinic ester capped and uncapped phenoxide forms. The normalized ratiometric turn-on responses for APC-1 and APC-2 were 89- and 101-fold, respectively.

In an era of personalized medicine, the clinical community has become increasingly focused on understanding diseases at the cellular and molecular levels. Magnetic resonance imaging (MRI) is a powerful imaging modality for acquiring anatomical and functional information. However, it has limited applications in the field of molecular imaging due to its low sensitivity. To expand the capability of MRI to encompass molecular imaging applications, we introduced bioresponsive Gd-(III)-based magnetic resonance contrast agents (GBCAs) in 1997. Since that time, many research groups across the globe have developed new examples of bioresponsive GBCAs. These contrast agents have shown great promise for visualizing several biochemical processes, such as gene expression, neuronal signaling, and hormone secretion. They are designed to be conditionally retained, or activated, in vivo in response to specific biochemical events of interest. As a result, an observed MR signal change can serve as a read-out for molecular events. A significant challenge for these probes is how to utilize them for noninvasive diagnostic and theranostic applications. This Perspective focuses on the design strategies that underlie bioresponsive probes, and describes the key advances made in recent years that are facilitating their application in vivo and ultimately in clinical translation. While the field of bioresponsive agents is embryonic, it is clear that many solutions to the experimental and clinical radiologic problems of today will be overcome by the probes of tomorrow.

Formaldehyde (FA), in the 0.2-0.4 mM range, is produced and maintained endogenously via enzymatic pathways. At these levels, FA can promote cell proliferation as well as mediate memory formation. Once elevated, FA stress is known to induce cognitive impairments, memory loss, and neurodegeneration owing to its potent DNA and protein cross-linking mechanisms. Optical imaging is a powerful noninvasive approach used to study FA in living systems; however, biocompatible chemical probes for FA are currently lacking. Herein, we report the design, synthesis, and biological evaluation of Formaldehyde Probe 1 (FP1), a new fluorescent indicator based on the 2-aza-Cope sigmatropic rearrangement. The remarkable sensitivity, selectivity, and photostability of FP1 has enabled us to visualize FA in live HEK293TN and Neuroscreen-1 cells. We envision that FP1 will find widespread applications in the study of FA associated with normal and pathological processes.

A peptide derived from Aβ17–36 crystallizes to form trimers that further associate to form higher-order oligomers. The trimers consist of three highly twisted β-hairpins in a triangular arrangement. Two trimers associate face-to-face in the crystal lattice to form a hexamer; four trimers in a tetrahedral arrangement about a central cavity form a dodecamer. These structures provide a working model for the structures of oligomers associated with neurodegeneration in Alzheimer’s disease.

Effective cancer therapy largely depends on inducing apoptosis in cancer cells via chemotherapy and/or radiation. Monitoring apoptosis in real-time provides invaluable information for evaluating cancer therapy response and screening preclinical anticancer drugs. In this work, we describe the design, synthesis, characterization and in vitro evaluation of caspase probe 1 (CP1), a bimodal fluorescence-magnetic resonance (FL-MR) probe that exhibits simultaneous FL-MR turn-on response to caspase-3/7. Both caspases exist as inactive zymogens in normal cells but are activated during apoptosis and are unique biomarkers for this process. CP1 has three distinct components: a DOTA-Gd(III) chelate that provides the MR signal enhancement, tetraphenylethylene as the aggregation induced emission luminogen (AIEgen), and DEVD peptide which is a substrate for caspase-3/7. In response to caspase-3/7, the water-soluble peptide DEVD is cleaved and the remaining Gd(III)-AIEgen (Gad-AIE) conjugate aggregates leading to increased FL-MR signals. CP1 exhibited sensitive and selective dual FL-MR turn-on response to caspase-3/7 in vitro and was successfully tested by fluorescence imaging of apoptotic cells. Remarkably, we were able to use the FL response of CP1 to quantify the exact concentrations of inactive and active agents and accurately predict the MR signal in vitro. We have demonstrated that the aggregation-driven FL-MR probe design is a unique method for MR signal quantification. This probe design platform can be adapted for a variety of different imaging targets, opening new and exciting avenues for multimodal molecular imaging.

Amyloid diseases such as Alzheimer's disease, Parkinson's disease, and type II diabetes share common features of toxic soluble protein oligomers. There are no structures at atomic resolution of oligomers formed by full-length amyloidogenic peptides and proteins, and only a few structures of oligomers formed by peptide fragments. The paucity of structural information provides a fundamental roadblock to understanding the pathology of amyloid diseases and developing preventions or therapies. Here, we present the X-ray crystallographic structures of three families of oligomers formed by macrocyclic peptides containing a heptapeptide sequence derived from the amyloidogenic E chain of β 2 -microglobulin (β 2 m). Each macrocyclic peptide contains the heptapeptide sequence β 2 m63-69 and a second heptapeptide sequence containing an N-methyl amino acid. These peptides form β-sheets that further associate into hexamers, octamers, and dodecamers: the hexamers are trimers of dimers; the octamers are tetramers of dimers; and the dodecamers contain two trimer subunits surrounded by three pairs of β-sheets. These structures illustrate a common theme in which dimer and trimer subunits further associate to form a hydrophobic core. The seven X-ray crystallographic structures not only illustrate a range of oligomers that a single amyloidogenic peptide sequence can form, but also how mutation can alter the size and topology of the oligomers. A cocrystallization experiment in which a dodecamerforming peptide recruits a hexamer-forming peptide to form mixed dodecamers demonstrates that one species can dictate the oligomerization of another. These findings should also be relevant to the formation of oligomers of full-length peptides and proteins in amyloid diseases.

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Real-time detection of bio-event in whole animals provides essential information for understanding biological and therapeutic processes. Magnetic resonance (MR) imaging represents a non-invasive approach to generating three-dimensional anatomic images with high spatial–temporal resolution and unlimited depth penetration. We have developed several self-immolative enzyme-activatable agents that provide excellent in vivo contrast and function as gene expression reporters. Here, we describe a vast improvement in image contrast over our previous generations of these bioresponsive agents based on a new pyridyl-carbamate Gd(III) complex. The pyridyl-carbamate-based agent has a very low MR relaxivity in the “off-state” (r 1 = 1.8 mM–1 s–1 at 1.41 T). However, upon enzymatic processing, it generates a significantly higher relaxivity with a Δr 1 = 106% versus Δr 1 ∼ 20% reported previously. Single X-ray crystal and nuclear magnetic relaxation dispersion analyses offer mechanistic insights regarding MR signal enhancement at the molecular scale. This work demonstrates a pyridyl-carbamate-based self-immolative molecular platform for the construction of enzymatic bio-responsive MR agents, which can be adapted to a wide range of other targets for exploring stimuli-responsive materials and biomedical applications.