Design and Characterization of Two Bifunctional Cryptophane A‐Based Host Molecules for Xenon Magnetic Resonance Imaging Applications

Abstract

Cryptophanes have been shown to be a promising tool in xenon magnetic resonance imaging (MRI) applications. The affinity of xenon for molecular host structures such as cryptophane‐A (CrA) and the large xenon chemical shift that can be detected in the presence of such hosts has motivated the synthesis of a number of cryptophane‐based MRI biosensors. Bifunctional biosensors that incorporate both a fluorophore and a cryptophane moiety are flexible tools for evaluating the intracellular localization and quantifying cellular uptake/binding of such xenon hosts in conjunction with testing their MRI capabilities. In this study, we compare the performance of a new compound that bears tetramethylrhodamine (TAMRA), CrA‐PEG‐TAMRA (PEG=polyethylene glycol), with our previously synthesized compound in which fluorescein (FAM) was the optical reporter. We demonstrate that these biosensors are suitable candidates for MRI‐based cell‐tracking studies; specifically, the new addition of a rhodamine derivative will allow for multiplexing experiments and showed slightly improved NMR spectroscopic behavior. Importantly, both biosensors display equally good cell‐labeling efficiency, which facilitates the acquisition of high‐quality xenon magnetic resonance images in two different cell lines. This study describes a simple synthetic approach for the production of biologically compatible bifunctional compounds. These biosensors can be used as they are for cell labeling but can also serve as building blocks for further functionalization and cryptophane‐based xenon biosensor design with MRI and fluorescence multiplexing options.

Magnetic resonance imaging with hyperpolarized agents: methods and applications

“…The development of bifunctional cryptophane-A host molecules with attached fluorophores for both HP 129 Xe MRI and optical imaging has also created a means for improved contrast agent characterization and utility through multimodal and multiscale imaging [234]. Furthermore, the range of chemical shifts (~300 ppm) exhibited by HP 129 Xe in different bound states creates the potential for tracking multiple biosensors simultaneously (i.e., “multiplexing”) and has been successfully tested in vitro by Klippel et al .…”

NMR Hyperpolarization Techniques of Gases

“…However, they are not an absolute requirement since the hydrophobic character of CrA can mediate membrane-association and therefore enables certain types of cellular labeling. [156] Sensors with highly specific binding motifs ( e.g. , antibodies [148, 151] or bioorthogonal reaction partners [153] ) only require sample-averaged concentrations of 10 −8 M for MRI.…”

“…[157] This property proved primarily useful to identify cell-associated cages in the first live-cell experiments. [150, 156a] A closer look in combination with FRET data revealed partitioning coefficients on the order 10 2 –10 3 for different membrane compositions. [158] This work also initiated a new class of Hyper-CEST experiments for investigating membrane fluidity and integrity: Due to the accelerated exchange, Xe signals from membrane-embedded cages do only differ marginally in chemical shift.…”

“…[170] The first imaging experiments with the latter substance also included the first multi-channel read-out of different host classes. [156b] Similar host compartments such as gas vesicles [171] and bacterial spores [172] will be discussed separately in the following section.…”

“…Spin-echo encoding is an alternative for cases where T 2 * relaxation makes EPI encoding impractical. [156a] …”