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Discovery of Signal Suppression by N-hydroxy Piperidine to Enable Activity-based Sensing by Chemical Exchange Saturation Transfer Magnetic Resonance ImagingTruong, Yen Janette 24 September 2020 (has links)
Fluorescent probes are useful tools for studying chemical biology, available in a wide variety of colours and applicable to different biochemical processes. One of their hallmarks is the ability to tune their chemistry and allow them to selectively “turn on” in response to different biomolecular targets of interest. However, fluorescence is largely limited by shallow tissue depth of penetration. Magnetic resonance imaging (MRI) can overcome the depth of penetration limitations to better map these biochemical processes and mechanisms with contrast agents. Chemical exchange saturation transfer (CEST) MRI is an alternative method to generating contrast in MR imaging that, like fluorescence, can provide multiplexed imaging by varying the chemical shift of the exchangeable proton on the contrast agent. Previously, a paramagnetic CEST agent containing two tetramethylpiperidinyloxyl (TEMPO) moieties was shown to reduce the CEST signal of a lanthanide complex due to T1 shortening effects on bulk water. Herein, we present a probe design strategy capable of suppressing the diamagnetic CEST (diaCEST) signal with the use of an N-hydroxy piperidine moiety. This discovery that N-hydroxy piperidine is capable of suppressing the diaCEST signal was applied to the study the activity of fibroblast activation protein-α (FAP). FAP is an enzyme involved in maintaining the tumour environment and its interactions can help understand tumour development, invasion and metastasis.
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Development and Application of CatalyCEST MRI Contrast Agents for the Study of Enzyme Activities in Tumor ModelsSinharay, Sanhita January 2016 (has links)
The in vivo detection of enzyme activity is a significant biomarker in tumorigenesis. Assessment of enzyme activity relative to enzyme concentration can serve as quite an accurate measurement of several disease states. Chemical Exchange Saturation Transfer (CEST) MRI is a non-invasive imaging technique that can be used to evaluate enzyme activity. Compared to other contrast agents CEST MRI agents have a slower chemical exchange rate and thus have greater specificity for detecting the intended biomarker. Chapter 1 provides an overview of the advances made in the field of molecular imaging for detection of cancer biomarkers. The molecular mechanism of each technique is explained with specific examples and advantages as well as disadvantages of each technique. Chapter 2 investigates the specific example of detection of an enzyme, γ-glutamyl transferase (GGT) in ovarian cancer tumor models using a catalyCEST MRI contrast agent. This chapter discusses the step-by step evaluation of the non-metallic contrast agent, from synthesis to evaluation of its catalytic efficiency with Michaelis Menten kinetics studies and finally in vivo GGT detection in ovarian tumor models of OVCAR-8 and OVCAR-3. Chapter 3 investigates the enzyme, Kallikrein-6 and its detection in HCT116 colon cancer tumor model. In addition to enzyme detection, enzyme inhibition using Antithrombin III inhibitor has also been explored within in vitro media and in vivo HCT116 tumor model. Chapter 4 introduces the catalyCEST agent for detection of sulfatase enzyme. This chapter discusses the synthesis of this agent and its ability to detect sulfatase in bacterial cell suspension and mammalian cell suspension. These examples portray catalyCEST MRI as a platform technology for enzyme activity detection. Finally in Chapter 5 future ideas have been proposed to improve the in vivo detection and broaden the applications of catalyCEST MRI in the field of enzyme studies.
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Development and Application of AcidoCEST MRI for Evaluating Tumor Acidosis in Pre-Clinical Cancer ModelsChen, Liu Qi January 2014 (has links)
Tumor acidosis is an important biomarker in cancer. We have developed a noninvasive imaging method, termed acidosis Chemical Exchange Saturation Transfer (acidoCEST) MRI to measure extracellular pH (pHe) in the tumor microenvironment. Chapter 1 introduces the importance of measuring tumor acidosis and presents various imaging modalities and their shortcoming to measure pHe. Chapter 2 describes the optimization of acidoCEST MRI for in vivo pHe measurement. The acidoCEST MRI protocol consists of a CEST-FISP acquisition and Lorentzian line shape fittings. We determined the optimal saturation time, saturation power and bandwidth, 5 sec, 2.8 µT and 90 Hz respectively. We also tried various routes of administration to increase contrast agent uptake in the tumor. We decided upon 200 µL bolus followed by 150 µL/hr infusion. The optimized acidoCEST MRI protocol was tested on a mammary carcinoma mouse model of MDA- MB-231. Our method can detect an increase in pHe in the bladder and tumor of the mice treated with bicarbonate. We used this optimized acidoCEST MRI method to measure pHe in lymphoma tumor model of Raji, Ramos and Granta 519 as described in Chapter 3. Pixel-wise pHe maps showed tumor heterogeneity. The pHe of Raji, Ramos and Granta 519 were determined to be mildly acidic with no significant difference. Chapter 4 describes the evolution of pixel-wise analysis in more detail. Besides the pHe map and spatial heterogeneity, we were able to determine the % contrast agent uptake. We monitored these biomarkers in two different mammary carcinoma mouse models, MDA- MB-231 and MCF-7 longitudinally and made comparisons between the different tumor models: MCF-7 were more acidic, more heterogeneous and faster growing than MDA- MB-231.
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Developing Responsive MRI Contrast Agents to Study Tumor BiologyHingorani, Dina Vinoo January 2014 (has links)
Enzymes are important biomarkers for determining tumor growth and progression. We have developed two molecules to image enzyme response by catalyCEST MRI. This technology allows for non-invasive detection of enzymes. A background of importance of measuring enzyme activity and MRI agents developed for this purpose have been covered in Chapter 1. We have synthesized a responsive paramagnetic Chemical Exchange Saturation Transfer (CEST) agent, called Tm-DO3A-cadaverine. This contrast agents has been successfully cross-linked to the protein albumin by the enzyme transglutaminase leading to the appearance of CEST at -9.2 ppm. The enzyme catalysis has been validated by measuring chemical exchange rates. We have shown that the position of the CEST peak is influenced by the conformation of the molecule depending on the neighboring amino acids to glutamine. This is the first example to show the appearance of CEST due to formation of a covalent bond. We have also synthesized a diamagnetic CEST agent with a large chemical shift dispersion to detect cathespin B activity. Upon enzyme mediated cleavage of PheArgSal, the aryl amide CEST peak at 5.3 ppm disappears. Taking a ratio of the CEST effects from salicylic acid at 9.5 ppm and aryl amide at 5.3 ppm we can detect enzyme activity. The salicylic acid moiety also undergoes some slow response due to enzyme action, as evident by the disappearance of CEST at 9.5 ppm. However, this proof of concept study is the first example of a DIACEST agent designed to measure enzyme activity using a ratio of two CEST effects from the same substrate. The last chapter highlights suggests improvements to the catalyCEST research. The appendix shows the use of bulk magnetic susceptibility measurements by NMR to determine bio-distribution of lanthanides in ex-vivo tissue.
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Machine learning applications for measuring pH using CEST MRIIcke, Ilknur 10 October 2019 (has links)
Non-invasive measurement of pH provides multiple potential benefits in oncology such
as better identifying the type of drug that can be more effective in chemotherapy, potentially identifying tumors that are more likely to metastasize and also better assessing the treatment effects. Chemical Exchange Saturation Transfer (CEST) Magnetic Resonance Imaging (MRI) is a versatile non-invasive technique for molecular imaging. AcidoCEST MRI techniques have been developed over the recent years to perform tumor pH measurements by utilizing a contrast agent for which chemical exchange saturation transfer effects depend on the pH of the microenvironment. Quantitative description of CEST MRI signals are generally done via modeling Bloch-McConnell equations by incorporating pH as a parameter or by fitting Lorentzian line shapes to observed z-spectra and then computing a log ratio of the CEST effects from multiple labile protons of the same molecule (ratiometric method). Modeling using Bloch-McConnell equations is complicated and requires careful inclusion of many scan parameters to infer pH. The ratiometric method requires contrast agents that have multiple labile protons, thus making it unsuitable to use for molecules with a single labile proton. Furthermore, depending on the pH, sometimes it might not be possible to numerically compute the ratio due to the inability of detecting signal peaks for certain labile protons.
Our aim here is to develop a machine learning based method that learns the CEST signal patterns from observed z-spectra on temperature and concentration-controlled contrast agent phantoms independent of the type of the contrast agent. Our results indicate that the machine learning method provides more general and accurate prediction of pH in comparison to the ratiometric method based on the phantom CEST dataset. Our method is more general in the sense that it does not require explicit modeling of signal peaks that are dependent on the type of contrast agent. We also describe a state of the art variational autoencoder based algorithm extending our machine learning method to measure tumor pH in vivo using AcidoCEST MRI on mouse tumor models.
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Rapid and Quantitative MRI of Chemical Exchange and Magnetization TransferShah, Tejas Jatin 30 July 2010 (has links)
No description available.
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Développement de l’imagerie métabolique par IRM-CEST : application à la maladie de Huntington / Development of metabolic imaging using CEST-MRI : application to Huntington’s diseasePépin, Jérémy 16 February 2018 (has links)
La maladie de Huntington (MH) est une maladie neurodégénérative héréditaire qui affecte le cerveau. Cette maladie est caractérisée par des signes cliniques tels que la dépression, la démence ainsi que des troubles moteurs s’aggravant au fil du temps. Ces déficiences sont dues à une augmentation anormale de la taille des répétitions CAG dans le gène codant la protéine huntingtine. Celle-ci s’accumule dans les cellules cérébrales et entraine leur mort. Des études antérieures ont démontré que le profil métabolique mesuré en spectroscopie RMN ¹H pouvait être altéré chez les patients atteints de cette maladie ainsi que des atrophies majeures de certaines structures du cerveau. Des hypothèses impliquant des défauts du métabolisme énergétique ont été avancées pour expliquer en partie la physiopathologie de la maladie. Les acteurs du métabolisme pourraient ainsi constituer des biomarqueurs d’intérêt. A l'aide d'une modalité d'IRM prometteuse appelée CEST (Chemical Exchange Saturation Transfer : Transfert de Saturation par Echange Chimique) il est possible de détecter des protons labiles faiblement concentrés qui sont classiquement indétectables en IRM. Il devient ainsi possible de cartographier in vivo la distribution de métabolites comme le glutamate (qui est un neurotransmetteur) ou le glucose (qui est le carburant des cellules) qui sont potentiellement impliqués dans les maladies neurodégénératives. Les développements méthodologiques effectués lors de cette thèse ont ensuite été appliqués à des modèles de rongeurs de la maladie de Huntington (souris KI140, souris R6/1, rats BACHD) afin d'identifier les biomarqueurs potentiels de la pathologie et d'évaluer la pertinence de ces méthodes IRM innovantes. L’ensemble de ces résultats et des méthodes mises en place durant cette thèse montrent le potentiel de l’imagerie CEST pour l’étude des maladies neurodégénératives. / Huntington's disease (HD) is a inherited neurodegenerative disease affecting the brain. This disease is characterized by clinical symptoms such as psychiatric, cognitive and motor disorders worsening over time. These deficiencies are due to an abnormal increase in the size of the CAG repeats in the gene encoding the huntingtin protein. Thisaccumulates in the brain cells and causes their death. Previous studies have shown that the metabolic profile measured in ¹H NMR spectroscopy can be altered in patients with this disease as well as major atrophy of certain structures of the brain. Hypotheses involving defects in energy metabolism have been advanced to explain partially the pathophysiology of the disease. The metabolic actors could thus be biomarkers of interest. Using a promising MRI modality called Chemical Exchange Saturation Transfer (CEST), it is possible to detect low-concentrated labile protons that are classically undetectable in MRI. It thus becomes possible to map in vivo the distribution of metabolites such as glutamate (which is a neurotransmitter) or glucose (which is the fuel of cells) which are potentially involved in neurodegenerative diseases. The methodological developments carried out during this thesis were then applied to rodent models of Huntington's disease (KI140 mice, R6/1 mice, BACHD rats) in order to identify potential biomarkers of the pathology and to evaluate the relevance of these innovative MRI methods. All of these results and methods implemented during this thesis show the potential of CEST imaging for the study of neurodegenerative diseases.
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MRI kontrastní látky využívající přenosu saturace / MRI contrast agents based on saturation transferKrchová, Tereza January 2012 (has links)
Title: MRI Contrast Agents Based on Saturation Transfer Author: Bc. Tereza Krchová Department: Department of Inorganic Chemistry, Faculty of Science, Charles University in Prague Supervisor: doc. RNDr. Jan Kotek, Ph.D. Supervisor's email: modrej@natur.cuni.cz Abstract: The aim of this thesis was to synthesize macrocyclic ligands (based on the DO3A and DO2A framework) with exchangeable protons on the aminic coordinating groups that could be (after the complexation with suitable paramagnetic metal ions) potential contrast agents based on saturation transfer, so called CEST contrast agents (Chemical Exchange Saturation Transfer). Two ligands H3L1 and H2L2 with one and two aminoethyl groups have been synthesized. The structurally similar ligands H3L1 Me and H3L1 2Me with one exchangeable proton and without it have been also prepared. The protonation constants of the ligands H3L1 and H2L2 have been determined by pH 1 H NMR and potentiometric titrations. The stability constants of metal ion complexes with H3L1 and H2L2 have been also determined by potentiometry. The CEST effect of Eu3+ and Yb3+ complexes with H3L1 , H3L1 Me and H3L1 2Me in solution have been studied. The residence lifetime of the inner-sphere water molecule has been determined for Gd-L1 complex. Keywords: CEST, MRI, aminoethyl, DO3A, DO2A, 17 O...
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Development and Evaluation of Exchange Rate Measurement MethodsRandtke, Edward Alexander January 2013 (has links)
Exchange rate determination allows precise modeling of chemical systems, and allows one to infer properties relevant to tumor biology such as enzyme activity and pH. Current exchange rate determination methods found via Contrast Enhanced Saturation Transfer agents are not effective for fast exchanging protons and use non-linear models. A comparison of their effectiveness has not been performed. In this thesis, I compare the effectiveness of current exchange rate measurement methods. I also develop exchange rate measurement methods that are effective for fast exchanging CEST agents and use linear models instead of non-linear models. In chapter 1 I review current exchange rate measurement methods. In chapter 2 I compare several of the current methods of exchange rate measurement, along with several techniques we develop. In chapter 3 I linearize the Quantifying Exchange through Saturation Transfer (QUEST) measurement method analogously to the Omega Plot method, and compare its effectiveness to the QUEST method. In chapter 4, I compare the effectiveness of current exchange rate theories (Transition State Theory and Landau-Zener theory) in the moderate coupling regime, and propose our own combined Eyring-Landau-Zener theory for this intermediate regime. In chapter 5 I discuss future directions for method development and experiments involving exchange rate determination.
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Characterization of a Novel Promoter Region for the Enteropathogenic Escherichia coli Type III Secretion System Chaperone Gene cesTBrouwers, Erin 05 December 2011 (has links)
Enteropathogenic Escherichia coli (EPEC) is an enteric pathogen that causes potentially fatal infantile diarrhea. A type III secretion system is employed by EPEC to inject bacterial effector proteins directly into host intestinal epithelial cells. The multivalent chaperone, CesT, interacts with nine effectors and is a significant contributor to EPEC pathogenesis. A putative transcriptional promoter region was identified directly upstream of cesT. In silico analyses identified conserved elements that suggest the cesT promoter is recognized by ?70. Using transcriptional fusions to lux reporter genes I showed that the cesT promoter region is active under conditions known to induce virulence-gene expression. I conclude that the cesT promoter is active early during an in vitro assay, and regulated by different mechanisms than those affecting the Ptir operon promoter.
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