Gustav J. Strijkers

MR molecular imaging and fluorescence microscopy for identification of activated tumor endothelium using a bimodal lipidic nanoparticle

In oncological research, there is a great need for imaging techniques that specifically identify angiogenic blood vessels in tumors on the basis of differences in the expression level of biomolecular markers. In the angiogenic cascade, different cell surface receptors, including the αvβ3‐integrin, are strongly expressed on activated endothelial cells. In the present study, we aimed to image angiogenesis by detecting the expression of αvβ3 in tumor bearing mice with a combination of magnetic resonance imaging (MRI) and fluorescence microscopy. To that end, we prepared MR‐detectable and fluorescent liposomes, which carry ∼700 αvβ3‐specific RGD peptides per liposome. RGD competition experiments and RAD‐conjugated liposomes were used as controls for specificity. In vivo, both RAD liposomes and RGD liposomes gave rise to signal increase on T1‐weighted MR images. It was established by the use of ex vivo fluorescence microscopy that RGD liposomes and RAD liposomes accumulated in the tumor by different mechanisms. RGD liposomes were specifically associated with activated tumor endothelium, while RAD liposomes were located in the extravascular compartment. This study demonstrates that MR molecular imaging of angiogenesis is feasible by using a targeted contrast agent specific for the αvβ3‐integrin, and that the multimodality imaging approach gave insight into the exact mechanism of accumulation in the tumor.

Nanoparticulate assemblies of amphiphiles and diagnostically active materials for multimodality imaging

Modern medicine has greatly benefited from recent dramatic improvements in imaging techniques. The observation of physiological events through interactions manipulated at the molecular level offers unique insight into the function (and dysfunction) of the living organism. The tremendous advances in the development of nanoparticulate molecular imaging agents over the past decade have made it possible to noninvasively image the specificity, pharmacokinetic profiles, biodistribution, and therapeutic efficacy of many novel compounds. Several types of nanoparticles have demonstrated utility for biomedical purposes, including inorganic nanocrystals, such as iron oxide, gold, and quantum dots. Moreover, natural nanoparticles, such as viruses, lipoproteins, or apoferritin, as well as hybrid nanostructures composed of inorganic and natural nanoparticles, have been applied broadly. However, among the most investigated nanoparticle platforms for biomedical purposes are lipidic aggregates, such as liposomal nanoparticles, micelles, and microemulsions. Their relative ease of preparation and functionalization, as well as the ready synthetic ability to combine multiple amphiphilic moieties, are the most important reasons for their popularity. Lipid-based nanoparticle platforms allow the inclusion of a variety of imaging agents, ranging from fluorescent molecules to chelated metals and nanocrystals. In recent years, we have created a variety of multifunctional lipid-based nanoparticles for molecular imaging; many are capable of being used with more than one imaging technique (that is, with multimodal imaging ability). These nanoparticles differ in size, morphology, and specificity for biological markers. In this Account, we discuss the development and characterization of five different particles: liposomes, micelles, nanocrystal micelles, lipid-coated silica, and nanocrystal high-density lipoprotein (HDL). We also demonstrate their application for multimodal molecular imaging, with the main focus on magnetic resonance imaging (MRI), optical techniques, and transmission electron microscopy (TEM). The functionalization of the nanoparticles and the modulation of their pharmacokinetics are discussed. Their application for molecular imaging of key processes in cancer and cardiovascular disease are shown. Finally, we discuss a recent development in which the endogenous nanoparticle HDL was modified to carry different diagnostically active nanocrystal cores to enable multimodal imaging of macrophages in experimental atherosclerosis. The multimodal characteristics of the different contrast agent platforms have proven to be extremely valuable for validation purposes and for understanding mechanisms of particle-target interaction at different levels, ranging from the entire organism down to cellular organelles.

MRI contrast agents : current status and future perspectives

Magnetic Resonance Imaging (MRI) is increasingly used in clinical diagnostics, for a rapidly growing number of indications. The MRI technique is non-invasive and can provide information on the anatomy, function and metabolism of tissues in vivo. MRI scans of tissue anatomy and function make use of the two hydrogen atoms in water to generate the image. Apart from differences in the local water content, the basic contrast in the MR image mainly results from regional differences in the intrinsic relaxation times T(1) and T(2), each of which can be independently chosen to dominate image contrast. However, the intrinsic contrast provided by the water T(1) and T(2) and changes in their values brought about by tissue pathology are often too limited to enable a sensitive and specific diagnosis. For that reason increasing use is made of MRI contrast agents that alter the image contrast following intravenous injection. The degree and location of the contrast changes provide substantial diagnostic information. Certain contrast agents are predominantly used to shorten the T(1) relaxation time and these are mainly based on low-molecular weight chelates of the gadolinium ion (Gd(3+)). The most widely used T(2) shortening agents are based on iron oxide (FeO) particles. Depending on their chemical composition, molecular structure and overall size, the in vivo distribution volume and pharmacokinetic properties vary widely between different contrast agents and these largely determine their use in specific diagnostic tests. This review describes the current status, as well as recent and future developments of MRI contrast agents with focus on applications in oncology. First the basis of MR image contrast and how it is altered by contrast agents will be discussed. After some considerations on bioavailability and pharmacokinetics, specific applications of contrast agents will be presented according to their specific purposes, starting with non-specific contrast agents used in classical contrast enhanced magnetic resonance angiography (MRA) and dynamic contrast enhanced MRI. Next targeted contrast agents, which are actively directed towards a specific molecular target using an appropriate ligand, functional contrast agents, mainly used for functional brain and heart imaging, smart contrast agents, which generate contrast as a response to a change in their physical environment as a consequence of some biological process, and finally cell labeling agents will be presented. To conclude some future perspectives are discussed.

Magnetic and fluorescent nanoparticles for multimodality imaging

The development of nanoparticulate contrast agents is providing an increasing contribution to the field of diagnostic and molecular imaging. Such agents provide several advantages over traditional compounds. First, they may contain a high payload of the contrast-generating material, which greatly improves their detectability. Second, multiple properties may be easily integrated within one nanoparticle to allow its detection with several imaging techniques or to include therapeutic qualities. Finally, the surface of such nanoparticles may be modified to improve circulation half-lives or to attach targeting groups. Magnetic resonance imaging and optical techniques are highly complementary imaging methods. Combining these techniques would therefore have significant advantages and may be realized through the use of nanoparticulate contrast agents. This review gives a survey of the different types of fluorescent and magnetic nanoparticles that have been employed for both magnetic resonance and optical imaging studies.

Paramagnetic Lipid-Coated Silica Nanoparticles with a Fluorescent Quantum Dot Core : A New Contrast Agent Platform for Multimodality Imaging

Silica particles as a nanoparticulate carrier material for contrast agents have received considerable attention the past few years, since the material holds great promise for biomedical applications. A key feature for successful application of this material in vivo is biocompatibility, which may be significantly improved by appropriate surface modification. In this study, we report a novel strategy to coat silica particles with a dense monolayer of paramagnetic and PEGylated lipids. The silica nanoparticles carry a quantum dot in their center and are made target-specific by the conjugation of multiple alphavbeta3-integrin-specific RGD-peptides. We demonstrate their specific uptake by endothelial cells in vitro using fluorescence microscopy, quantitative fluorescence imaging, and magnetic resonance imaging. The lipid-coated silica particles introduced here represent a new platform for nanoparticulate multimodality contrast agents.

Chitosan-based systems for molecular imaging

Molecular imaging enables the non-invasive assessment of biological and biochemical processes in living subjects. Such technologies therefore have the potential to enhance our understanding of disease and drug activity during preclinical and clinical drug development. Molecular imaging allows a repetitive and non-invasive study of the same living subject using identical or alternative biological imaging assays at different time points, thus harnessing the statistical power of longitudinal studies, and reducing the number of animals required and cost. Chitosan is a hydrophilic and non-antigenic biopolymer and has a low toxicity toward mammalian cells. Hence, it has great potential as a biomaterial because of its excellent biocompatibility. Conjugated to additional materials, chitosan composites result in a new class of biomaterials that possess mechanical, physicochemical and functional properties, which have potential for use in advanced biomedical imaging applications. The present review will discuss the strengths, limitations and challenges of molecular imaging as well as applications of chitosan nanoparticles in the field of molecular imaging.

Structure and magnetization of arrays of electrodeposited Co wires in anodic alumina

We have produced arrays of Co nanowires in anodic porous alumina filters by means of electrodeposition. The structure and magnetization behavior of the wires was investigated with nuclear magnetic resonance (NMR) and magnetization measurements. NMR shows that the wires consist of a mixture of fcc and hcp texture with the (0001) texture of the hcp fraction oriented preferentially perpendicular to the wires. The magnetization direction is determined by a competition of demagnetizing fields and dipole–dipole fields and can be tuned parallel or perpendicular to the wires by changing the length of the wires.

Determination of mouse skeletal muscle architecture using three-dimensional diffusion tensor imaging

Muscle architecture is the main determinant of the mechanical behavior of skeletal muscles. This study explored the feasibility of diffusion tensor imaging (DTI) and fiber tracking to noninvasively determine the in vivo three‐dimensional (3D) architecture of skeletal muscle in mouse hind leg. In six mice, the hindlimb was imaged with a diffusion‐weighted (DW) 3D fast spin‐echo (FSE) sequence followed by the acquisition of an exercise‐induced, T2‐enhanced data set. The data showed the expected fiber organization, from which the physiological cross‐sectional area (PCSA), fiber length, and pennation angle for the tibialis anterior (TA) were obtained. The values of these parameters ranged from 5.4–9.1 mm2, 5.8–7.8 mm, and 21–24°, respectively, which is in agreement with values obtained previously with the use of invasive methods. This study shows that 3D DT acquisition and fiber tracking is feasible for the skeletal muscle of mice, and thus enables the quantitative determination of muscle architecture. Magn Reson Med 53:1333–1340, 2005.

Relaxivity of liposomal paramagnetic MRI contrast agents

Paramagnetic liposomes, spherical particles formed by a lipid bilayer, are able to accommodate a high payload of Gd-containing lipid and therefore can serve as a highly potent magnetic resonance imaging contrast agent. In this paper the relaxation properties of paramagnetic liposomes were studied as a function of composition, temperature and magnetic field strength. The pegylated liposomes with a diameter of approximately 100 nm were designed for favorable pharmacokinetic properties in vivo. The proton relaxivity, i.e. the T1 relaxation rate per mmol of Gd(III) ions, of liposomes with unsaturated DOPC phospholipids was higher than those with saturated DSPC lipids. Addition of cholesterol was essential to obtain monodisperse liposomes and led to a further, although smaller, increase of the relaxivity. Nuclear magnetic relaxation dispersion measurements showed that the relaxivity was limited by water exchange. These results show that these paramagnetic liposomes are very effective contrast agents, making them excellent candidates for many applications in magnetic resonance imaging.

Synergistic targeting of alphavbeta3 integrin and galectin-1 with heteromultivalent paramagnetic liposomes for combined MR imaging and treatment of angiogenesis

Effective and specific targeting of nanoparticles is of paramount importance in the fields of targeted therapeutics and diagnostics. In the current study, we investigated the targeting efficacy of nanoparticles that were functionalized with two angiogenesis-specific targeting ligands, an alpha(v)beta(3) integrin-specific and a galectin-1-specific peptide. We show in vitro, using optical techniques and MRI, that the dual-targeting approach produces synergistic targeting effects, causing a dramatically elevated uptake of nanoparticles as compared to single ligand targeting.