Dmitri Artemov

MR molecular imaging of the Her-2/neu receptor in breast cancer cells using targeted iron oxide nanoparticles

MR molecular imaging is an exciting new frontier in the biomedical applications of MR. One of the clinically relevant targets is the tyrosine kinase Her‐2/neu receptor, which has a significant role in staging and treating breast cancer. In this study Her‐2/neu receptors were imaged in a panel of breast cancer cells expressing different numbers of the receptors on the cell membrane. Commercially available streptavidin‐conjugated superparamagnetic nanoparticles were used as targeted MR contrast agent. The nanoparticles were directed to receptors prelabeled with a biotinylated monoclonal antibody and generated strong T2 MR contrast in Her‐2/neu‐expressing cells. The contrast observed in MR images was proportional to the expression level of Her‐2/neu receptors determined independently with FACS analysis. In these experiments, iron oxide nanoparticles were attached to the cell surface and were not internalized into the cells, which is a major advantage for in vivo applications of the method. Magn Reson Med 49:403–408, 2003.

Twist Overexpression Induces In vivo Angiogenesis and Correlates with Chromosomal Instability in Breast Cancer

Aggressive cancer phenotypes are a manifestation of many different genetic alterations that promote rapid proliferation and metastasis. In this study, we show that stable overexpression of Twist in a breast cancer cell line, MCF-7, altered its morphology to a fibroblastic-like phenotype, which exhibited protein markers representative of a mesenchymal transformation. In addition, it was observed that MCF-7/Twist cells had increased vascular endothelial growth factor (VEGF) synthesis when compared with empty vector control cells. The functional changes induced by VEGF in vivo were analyzed by functional magnetic resonance imaging (MRI) of MCF-7/Twist-xenografted tumors. MRI showed that MCF-7/Twist tumors exhibited higher vascular volume and vascular permeability in vivo than the MCF-7/vector control xenografts. Moreover, elevated expression of Twist in breast tumor samples obtained from patients correlated strongly with high-grade invasive carcinomas and with chromosome instability, particularly gains of chromosomes 1 and 7. Taken together, these results show that Twist overexpression in breast cancer cells can induce angiogenesis, correlates with chromosomal instability, and promotes an epithelial-mesenchymal-like transition that is pivotal for the transformation into an aggressive breast cancer phenotype.

Natural D -glucose as a biodegradable MRI contrast agent for detecting cancer

Modern imaging technologies such as CT, PET, SPECT, and MRI employ contrast agents to visualize the tumor microenvironment, providing information on malignancy and response to treatment. Currently, all clinical imaging agents require chemical labeling, i.e. with iodine (CT), radioisotopes (PET/SPECT), or paramagnetic metals (MRI). The goal was to explore the possibility of using simple D‐glucose as an infusable biodegradable MRI agent for cancer detection.

Molecular magnetic resonance imaging with targeted contrast agents

Magnetic resonance imaging (MRI) produces high‐resolution three‐dimensional maps delineating morphological features of the specimen. Differential contrast in soft tissues depends on endogenous differences in water content, relaxation times, and/or diffusion characteristics of the tissue of interest. The specificity of MRI can be further increased by exogenous contrast agents (CA) such as gadolinium chelates, which have been successfully used for imaging of hemodynamic parameters including blood perfusion and vascular permeability. Development of targeted MR CA directed to specific molecular entities could dramatically expand the range of MR applications by combining the noninvasiveness and high spatial resolution of MRI with specific localization of molecular targets. However, due to the intrinsically low sensitivity of MRI (in comparison with nuclear imaging), high local concentrations of the CA at the target site are required to generate detectable MR contrast. To meet these requirements, the MR targeted CA should recognize targeted cells with high affinity and specificity. They should also be characterized by high relaxivity, which for a wide variety of CA depends on the number of contrast‐generating groups per single molecule of the agent. We will review different designs and applications of targeted MR CA and will discuss feasibility of these approaches for in vivo MRI.

Twist contributes to hormone resistance in breast cancer by downregulating estrogen receptor-α

The role of estrogen receptor-α (ER) in breast cancer development, and as a primary clinical marker for breast cancer prognosis, has been well documented. In this study, we identified the oncogenic protein, TWIST1 (Twist), which is overexpressed in high-grade breast cancers, as a potential negative regulator of ER expression. Functional characterization of ER regulation by Twist was performed using Twist low (MCF-7, T-47D) and Twist high (Hs 578T, MDA-MB-231, MCF-7/Twist) expressing cell lines. All Twist high expressing cell lines exhibited low ER transcript and protein levels. By chromatin immunoprecipitation and promoter assays, we demonstrated that Twist could directly bind to E-boxes in the ER promoter and significantly downregulate ER promoter activity in vitro. Functionally, Twist overexpression caused estrogen-independent proliferation of breast cells, and promoted hormone resistance to the selective estrogen receptor modulator tamoxifen and selective estrogen receptor down-regulator fulvestrant. Importantly, this effect was reversible on downregulating Twist. In addition, orthotopic tumors generated in mice using MCF-7/Twist cells were resistant to tamoxifen. These tumors had high vascular volume and permeability surface area, as determined by magnetic resonance imaging (MRI). Mechanistically, Twist recruited DNA methyltransferase 3B (DNMT3B) to the ER promoter, leading to a significantly higher degree of ER promoter methylation compared with parental cells. Furthermore, we demonstrated by co-immunoprecipitation that Twist interacted with histone deacetylase 1 (HDAC1) at the ER promoter, causing histone deacetylation and chromatin condensation, further reducing ER transcript levels. Functional re-expression of ER was achieved using the demethylating agent, 5-azacytidine, and the HDAC inhibitor, valproic acid. Finally, an inverse relationship was observed between Twist and ER expression in human breast tumors. In summary, the regulation of ER by Twist could be an underlying mechanism for the loss of ER activity observed in breast tumors, and may contribute to the generation of hormone-resistant, ER-negative breast cancer.

Optimization of SABRE for polarization of the tuberculosis drugs pyrazinamide and isoniazid

Hyperpolarization produces nuclear spin polarization that is several orders of magnitude larger than that achieved at thermal equilibrium thus providing extraordinary contrast and sensitivity. As a parahydrogen induced polarization (PHIP) technique that does not require chemical modification of the substrate to polarize, Signal Amplification by Reversible Exchange (SABRE) has attracted a lot of attention. Using a prototype parahydrogen polarizer, we polarize two drugs used in the treatment of tuberculosis, namely pyrazinamide and isoniazid. We examine this approach in four solvents, methanol-d4, methanol, ethanol and DMSO and optimize the polarization transfer magnetic field strength, the temperature as well as intensity and duration of hydrogen bubbling to achieve the best overall signal enhancement and hence hyperpolarization level.

Molecular and functional imaging of cancer: Advances in MRI and MRS

Publisher Summary This chapter presents an overview of the endogenous and exogenous magnetic resonance (MR) contrast mechanisms utilized in characterizing tumor vasculature. Every contrast mechanism for probing the tumor vasculature, including the use of exogenous MR contrast agents, is in some way a result of the changes in the MR signal intensity brought about by changes in tissue relaxation times. Noninvasive multinuclear magnetic resonance imaging (MRI) and MR spectroscopic imaging (MRSI) provide a wealth of spatial and temporal information on tumor vasculature, metabolism, and physiology. The most commonly used MR contrast agents (CA) are paramagnetic gadolinium chelates. These agents are tightly bound complexes of the rare earth element gadolinium and various chelating agents. A simple linear compartment model, describing uptake of the contrast agent from plasma, postulates a negligible reflux of the contrast agent from the interstitium back to the blood compartment. Blood concentrations of the CA can be approximated to be constant for the duration of the MR experiment, and under these conditions, contrast uptake is a linear function of time. The MR detection of cellular targets is also elaborated.

Magnetic Resonance Spectroscopy in Metabolic and Molecular Imaging, and Diagnosis of Cancer

Since its discovery in the 1940s, magnetic resonance (MR) spectroscopy (MRS) has developed into a major technique used by chemists to elucidate molecular structures. The underlying principle of MRS is the generation of radiofrequency (RF) signals by magnetic nuclear spins that are excited with a specific RF to precess in an external magnetic field B0. The magnetic resonance frequency ω0 is linearly dependent on B0 and the gyromagnetic ratio of the nucleus γ, as ω0 = γB0. The MR signal intensity depends on the concentration of nuclear spins, the magnetic field strength B0, and the gyromagnetic ratio γ of these spins. The magnetization signal in MRS is characterized by two rate constants, the spin-lattice (or longitudinal relaxation time) T1, and the spin-spin (or tranverse relaxation time) T2. Since the resonance frequency of a particular nucleus is dependent upon its molecular structure, an important aspect of MRS is the ability to distinguish a nucleus with respect to its environment in the molecule. Because the molecular structure-based frequency shift and the resonance frequency are directly proportional to the strength of the magnetic field, the frequency shift is converted into a field-independent dimensionless value known as the chemical shift. Since the frequency shifts are extremely small in comparison to the resonance frequency, the chemical shift is expressed in parts per million (ppm). The chemical shift is typically reported relative to a reference resonance frequency. MRS therefore provides information about the chemical environment of the nuclear spin such as number of chemical bonds, neighboring nuclei, and overall chemical structure. As a result, each peak in an MR spectrum has a characteristic chemical shift that is dependent upon the chemical structure of the metabolite or compound, and a peak area that is proportional to the concentration of the compound. Scalar spin-spin interactions, or J-couplings, produce fine multiplet structures that can be used to further analyze the chemical structure of a given molecule. Within the past two decades, the same principles of chemical shifts, magnetic moments, relaxation rates, and deriving concentrations from peak integrals have been applied in several preclinical and clinical studies to advance cancer discovery, diagnosis, and treatment. Incorporating imaging techniques with MRS has resulted in the development of MR spectroscopic imaging (MRSI) where the chemical information is spatially phase encoded,1–3 providing images of specific chemical compounds such as metabolites, reporter probes, labeled substrates, or drugs. The purpose of this article is to review recent developments and examples of the use of multi-nuclear MRS in cancer, and its integration with multi-modality imaging in cancer discovery and treatment. The ability of cancer cells to adapt and survive treatments, and the collateral damage to normal cells as a result of several cancer treatments, continue to make the successful treatment of cancer a major challenge for the twenty first century. Tumor recurrence and metastasis are the leading causes of morbidity and mortality from cancer and, despite major advances in cancer research and treatment, cancer continues to evade cure. This is not surprising given the complexities of a tumor, and the genomic plasticity of cancer cells and stromal cells that are co-opted within the tumor. A schematic of the different components of a tumor is shown in Figure 1. Figure 1 Schematic of the components of a tumor. Cancer cells are embedded within the extracellular matrix (ECM). The ECM consists of a complex meshwork of structural extracellular proteins. The tumor microenvironment (TME) contains the ECM and stromal cells such ... Physiological conditions such as hypoxia and acidic extracellular pH (pHe) that exist in the tumor microenvironment, the interactions between cancer cells and stromal cells such as endothelial cells, fibroblasts and macrophages, the extracellular matrix, and the numerous secreted factors and cytokines cumulatively influence progression, aggressiveness, and response of the disease to treatment. Hypoxia, in particular, is a major cause of radio- and chemo-resistance in cancer cells. Because of the remarkable ability of cancer cells to adapt and survive, finding effective treatments against cancer depends upon identifying and attacking targets and pathways critically important for the cancer cell. Multi-nuclear MRS provides unique opportunities for molecular and functional imaging of cancer in preclinical and clinical studies, and for imaging interactions between cancer cells and stromal cells. Some of these applications of MRS, the nuclei commonly studied, and the information that can be obtained are summarized in Table 1. From this table it is apparent that multi-nuclear noninvasive MRS methods have wide-ranging applications in cancer that can translate from bench to beside. The chemical structures of various compounds referred to in this review article are summarized in Tables 2 and ​and3.3. Table 3 also provides details regarding administration and dose of the contrast agents described here. Table 1 Nuclei commonly studied in order of sensitivity of detection and some of their preclinical (+) and clinical (*) applications in cancer. The intensity of the MR signal depends on the concentration of nuclear spins, and the gyromagnetic ratio γ ... Table 2 Chemical structures of metabolites referred to in this review article. Table 3 Chemical structures of reporter molecules referred to in this review article. The past decade has seen major advances in sequence design, development of novel reporter probes, and technological advances that have significantly increased the uses of MRS in molecular and functional imaging applications in oncology. Some of the recent applications of 1H, 13C, 31P, and 19F MRS in preclinical models of cancer are reviewed, and examples of biomedical MRS applications for each nucleus are shown in Figure 2. New developments, such as hyperpolarization of spins to increase the sensitivity of detection of the MR signal of 13C-labeled substrates are discussed. Advantages and limitations of the spectroscopic techniques and challenges for the future are outlined. Figure 2 Examples of multi-nuclear MRS applications. From top to bottom: Representative in vivo single-voxel 1H MRS of an MDA-MB-231 breast tumor xenograft model obtained at 4.7T, and ex vivo high-resolution 1H MRS of a water-soluble MDA-MB-231 cell extract obtained ... The use of MR biomarkers such as the total choline (tCho) signal and perfusion are already being explored clinically for characterizing tumors and following treatment response.4–7 The elevation of choline compounds presents a unique target to exploit for molecular targeting; such targeting can be imaged noninvasively with MRS.8,9 Both pharmacological and molecular approaches are being developed to target choline metabolism, specifically choline kinase activity, which is the first step in choline phospholipid biosynthesis. The interaction between cancer cells and the tumor microenvironment is providing new insights into the etiology and progression of cancer. For example, oxygen partial pressure (pO2) in tumors can be imaged by both 1H and 19F MRSI.10,11 Oxygen is sensed by administration of reporter molecules through changes in the spin-lattice relaxation rate. Integrating MRSI with MRI and other imaging modalities such as optical and nuclear imaging is providing useful insights into the dynamics between hypoxia and the tumor extracellular matrix (ECM), vascularization, extracellular pH, interstitial fluid transport, and metabolism in preclinical models.12–14 These insights can be exploited to find effective treatment strategies.

Magnetic resonance imaging of cell surface receptors using targeted contrast agents.

Over the past decade MR (magnetic resonance) imaging has emerged as one of the major modalities for noninvasive functional imaging. Recent advances in the development of targeted MR contrast agents have added significantly to the capabilities of MR imaging. In particular, the use of targeted contrast agents to report on the expression of cell surface receptors, combined with the functional capabilities of MR imaging, together provide unique opportunities to understand receptor-mediated pathways. In this article we have reviewed current MRI strategies used to visualize receptor expression, the potential advantages and drawbacks of these strategies, and novel areas of focus for the future.

Tumor angiogenesis, vascularization, and contrast-enhanced magnetic resonance imaging.

Angiogenesis, the process by which new blood vessels are generated, occurs during wound healing, in the female reproductive system during ovulation and gestation, and during embryonic development. The process is carefully controlled with positive and negative regulators, because several vital physiological functions require angiogenesis. The consequences of abnormal angiogenesis are either excessive or insufficient blood vessel growth. Ulcers, strokes, and heart attacks can result from the absence of angiogenesis normally required for natural healing, whereas excessive blood vessel proliferation may favor tumor growth and dissemination, blindness, and arthritis. In this review, the process of angiogenesis and the characteristics of the resulting tumor vasculature are outlined. Contrast-enhanced magnetic resonance imaging techniques that currently are available for basic research and clinical applications to study various aspects of tumor angiogenesis and neovascularization are discussed.