Our main projects currently are:
Intracellular targeting (enzyme and mRNA targeting)
In recent years several, mainly intracellularly targeted CA were developed by our group. Enzymes (model: β-galactosidase) and mRNA (model: mRNA of red fluorescent protein DsRed) were chosen as cellular targets. Although these CA efficiently enhanced the contrast in MR images of labeled cells (even when low micromolar concentrations were applied), a high unspecific background signal in non-targeted cells prevented a successful in vivo application so far. The intracellular localization of the CA (e.g. cytosol vs. endosomes) is not only crucial for a specific interaction and accumulation in the target cells but also for the efficacy of MR contrast enhancement. Even a significantly higher accumulation can be insufficient to increase contrast in MR images when the CA is predominantly located in endosomes or lysosomes.
A new approach to reduce the unspecific background signal in non-targeted cells is followed by using 19F MRI and an enzyme targeted probe where the 19F signal will be only detectable after the action of the enzyme. The synthesized probe was successfully tested in solution and the proof of concept in cells and in vivo in a tumor model expressing β-galactosidase is in progress.
To circumvent endosomal entrapment we were also looking for new vector molecules (e.g. cell penetrating peptides, CPP) showing optimized cellular distribution. This study was leading to a novel cysteine rich peptide, CyLoP-1, which is able to transport efficiently various cargo molecules (e.g. the bioactive pro-apoptotic SmacN7-peptide) into the cytosol. When coupled to a Gadolinium chelate it is a potent MR CA strongly reducing relaxivity quenching effects due to endosomal entrapment.
In parallel, non-peptide delivery systems (e.g. lipid based systems like cholesterol) were tested for mRNA targeted imaging probes. Whilst uptake efficiency could be further increased compared to CPP conjugates the cellular distribution and thus the specificity were not altered. However, the synthesis of peptide nucleic acids, used to target mRNA, could be optimized further.
Recently we initiated a new project in collaboration with the University of Durham, UK. Here we developed CA responsive for the metabotropic glutamate receptor mGluR5 in the brain. These CA based on highly stable Gadolinium chelates coupled to known antagonists of this receptor. The goal is the non-invasive visualization of the receptors in the brain and potentially a more direct monitoring of neuronal activity by imaging extracellular glutamate fluctuations during activation.
Silica-based platforms for MR and multi modality imaging probes
One drawback of MRI is its relatively low sensitivity. A large number of contrast producing moieties is required to obtain a significant change in image contrast. To achieve a higher local accumulation, macromolecular or nanoparticle-based platforms can be used to increase the number of MR reporters per molecule. In collaboration with the University of Tübingen SiO2-based systems (silsesquioxanes, nonporous nanoparticles) functionalized with Gadolinium chelates were developed. The relatively small silsesquioxanes allow the coupling of up to eight chelates by retaining a pharmacokinetic profile of a low molecular weight CA. They undergo a slow degradation process under physiological conditions to mono-chelates  readily excreted via the kidneys. In vivo, a rapid accumulation in the gastrointestinal tract after i.v. injection into mice was observed not present when the same concentration of Gadolinium as GdDOTA, a commercially available CA used in the clinics, was injected.
Silica-based nanoparticles (NPs) are much larger (~ 50 – 100 nm in diameter) and thus will show different biodistribution and pharmacokinetics. However, a large number of imaging reporters can be conjugated per particle and different functionalities (e.g. targeting sensors, vectors for cellular uptake) can be introduced.
NPs with a high payload of Gadolinium were synthesized and tested for their ability to enhance MR contrast in vitro . These NPs were further functionalized with fluorophores for optical imaging and a CPP to enhance cellular uptake. Such NPs can be taken up by cells and a initial in vivo study on the biodistribution after i.v. injection showed a fast accumulation particularly in lungs and liver, typical for nanoparticles.
A different type of NPs (ferromagnetic metallic cobalt with a functionalized carbon coating) is used for in vivo targeting of β-cells in pancreatic islets of Langerhans in mice. A β-cell specific single chain antibody fragment (SCA), developed by our collaborators, was coupled to these NPs to obtain a targeted T2-CA. In combination with the unprecedented spatial resolution which is achievable at our 16.4T animal scanner and optimizing imaging sequences and image processing by our MRI experts, it was possible for the first time to visualize single islets in vivo in a freely breathing mouse after i.v. injection. This study particularly exemplifies the great advantage to have CA developers (chemists, biologists) and MR physicists working close together in one group.
Understanding brain connectivity is important to enhance the understanding of brain function and disease. To correlate brain anatomy to the functional outcome and to follow changes e.g. with development, aging or learning, non-invasive longitudinal studies are needed. MRI is providing an excellent measure to perform dynamic investigations of brain connectivity by using MR active neuroanatomical tracers. In a collaboration with former and present members of the Dept. Logothetis we are helping to develop such novel tracer molecules by providing information about the uptake efficacy into and the transport of the probes inside neural cells along their processes. Two classes of tracers, for short term (1 - 3 days) and long term (7 - 14 days) studies, were successfully tested in vitro and in vivo.
Molecular in vivo imaging of cellular therapeutics – CeTheProbes
This work is in parts performed in the frame of the joint research project of the MPI for Biological Cybernetics (coordinator), Eberhard-Karls University Tübingen, and Hannover Medical School, funded by the German Federal Ministry of Education and Research (BMBF, FKZ 01EZ0813)