Hans-Ulrich Gremlich

In vivo detection of amyloid-β deposits by near-infrared imaging using an oxazine-derivative probe

As Alzheimers disease pathogenesis is associated with the formation of insoluble aggregates of amyloid β-peptide, approaches allowing the direct, noninvasive visualization of plaque growth in vivo would be beneficial for biomedical research. Here we describe the synthesis and characterization of the near-infrared fluorescence oxazine dye AOI987, which readily penetrates the intact blood-brain barrier and binds to amyloid plaques. Using near-infrared fluorescence imaging, we demonstrated specific interaction of AOI987 with amyloid plaques in APP23 transgenic mice in vivo, as confirmed by postmortem analysis of brain slices. Quantitative analysis revealed increasing fluorescence signal intensity with increasing plaque load of the animals, and significant binding of AOI987 was observed for APP23 transgenic mice aged 9 months and older. Thus, AOI987 is an attractive probe to noninvasively monitor disease progression in animal models of Alzheimer disease and to evaluate effects of potential Alzheimer disease drugs on the plaque load.

Fluorescent Nanoprobes as a Biomarker for Increased Vascular Permeability: Implications in Diagnosis and Treatment of Cancer and Inflammation

This article describes the use of a fluorescent nanoprobe as a functional biomarker for the identification of increased vascular permeability in cancer/arthritis disease models. Synthesis of the fluorescent nanoprobe was achieved by passive loading of a fluorophore inside the nanoparticle using thin film hydration method. The outer layer of the nanoprobe was decorated with poly(ethylene glycol) arms to increase the bioavailability of the fluorophore. Stability studies of the nanoprobe showed that the particles were stable up to 70 days. The uptake and internalization of the fluorescent nanoprobe inside target cells was confirmed by fluorescence microscopy studies. Co-localization of the probe with the target tissue in vivo was unambiguously identified using intravital microscopy. Results from in vivo imaging studies showed that the particles had a long half-life in the circulation and passively targeted tumor or arthritic tissue. The increased and specific uptake of the fluorescent nanoprobe in tumor/arthritic tissue is attributed to an enhanced permeation and retention (EPR) effect. Use of an optical method to validate anti-inflammatory drugs in an arthritis disease model is demonstrated in this study. In general, this methodology could be used for detection of leaky vasculature in different pathological states.

Noninvasive assessment of gastric emptying by near-infrared fluorescence reflectance imaging in mice: pharmacological validation with tegaserod, cisapride, and clonidine.

Noninvasive near-infrared fluorescence reflectance imaging (FRI) is an in vivo technique to assess physiological and molecular processes in the intact organism. Here we describe a method to assess gastric emptying in mice. TentaGel beads with covalently bound cyanine dye (Cy5.5) conjugates as fluorescent probe were administered by oral gavage. The amount of intragastric beads/label was derived from the fluorescence signal intensity measured in a region of interest corresponding to the mouse stomach. The FRI signal intensity decreased as a function of time reflecting gastric emptying. In control mice, the gastric half-emptying time was in agreement with literature data. Pharmacological modulation of gastric motility allowed the evaluation of the sensitivity of the FRI-based method. Gastric emptying was either stimulated or inhibited by treatment with the 5-HT(4) receptor agonists tegaserod (Zelnorm) and cisapride or the alpha(2)-receptor agonist clonidine, respectively. Tegaserod and cisapride dose-dependently accelerated gastric emptying. In contrast, clonidine dose-dependently delayed gastric emptying. In conclusion, FRI using fluorescently labeled beads allows the reliable determination of gastric emptying as well as the assessment of pharmacological interventions. The technique thus offers the potential to characterize molecular targets and pathways involved in physiological regulation and pharmacological modulation of gastric emptying.

Investigating Pharmacology In Vivo Using Magnetic Resonance and Optical Imaging

The better and earlier a disease can be diagnosed and characterized, the greater the chance of being able to intervene in this process with a chemical entity. This is the rationale for the use of in vivo imaging techniques in the drug discovery and development process. In this article we address the value of two imaging modalities in this area, i.e. magnetic resonance imaging (MRI) and optical imaging. The multiparametric nature of MRI enables anatomical, functional, metabolic and, to a certain extent, also cellular and target-related information to be obtained noninvasively at high spatial resolution. This favours characterization of a disease state and the corresponding drug intervention. The noninvasiveness of MRI strengthens the link between preclinical and clinical pharmaceutical research. The high sensitivity of optical techniques enables molecular information to be obtained in vivo. Within pharmacological research, the main applications of optical techniques relate to the early drug discovery process and acquisition of target-related information. However, potential clinical applications of optical imaging are also emerging. The complementary character of both imaging modalities renders them useful in various portions of the drug discovery process, from early target selection and validation to clinical studies.

Role of Fourier transform infrared spectroscopy in the rehearsal phase of combinatorial chemistry : a thin-layer chromatography equivalent for on-support monitoring of solid-phase organic synthesis

The adaptation of diverse organic reactions to solid supports requires significant reaction optimization efforts. A convenient on-support analytical method functionally similar to TLC in solution chemistry is very advantageous. As a TLC-equivalent method, the single bead FTIR is a simple, sensitive, fast, and convenient analytical method to monitor SPOS without stopping the reaction or cleaving the product. As with TLC, single bead FTIR provides a wide range of information such as qualitative assessment, quantitative determination, and reaction kinetics. Studies with the single bead FTIR have not only provided a tool for daily monitoring of the solid-phase reactions, but a way to understand the properties of polymer-bound substrate and the nature of polymer-supported organic reactions. It has assisted in the selection of a wide range of reaction conditions rapidly for SPOS in the rehearsal phase of combinatorial chemistry. Due to its convenience and efficiency, FTIR internal reflection spectroscopy has evolved as a useful analytical methodology for monitoring of combinatorial chemistry reactions directly on polymer surface.

Chapter 7.1:In vivo Fluorescence Optical and Multi-Modal Imaging in Pharmacological Research: From Chemistry to Therapy Monitoring

The better and earlier a disease can be diagnosed and characterized, the higher the chance of being able to interfere in this process with a chemical entity. This is the rationale for the use of in vivo molecular imaging techniques in pharmaceutical research and development. The present review addresses the value of fluorescence optical imaging in this area. Through the administration of exogenous fluorochromes, molecular processes can be monitored non-invasively in vivo with operational simplicity, safety and high cost-effectiveness. This provides potential for obtaining information related to target validation, detection and characterization of pathology, and treatment evaluation. Although most applications are preclinical, in small rodents, a few clinical uses are also emerging. A key element for the success of the technique is the availability of an adequate fluorescent probe to address a given biological or pharmacological question of interest.