4809 The current cancer therapeutic modalities have encountered tremendous challenges in treating... more 4809 The current cancer therapeutic modalities have encountered tremendous challenges in treating hypoxic cancer cells due to poor drug delivery for effective chemotherapy or low level of tumor oxygenation for effective radiation therapy. Bacteria can penetrate into hypoxic tumor regions and deliver therapeutic proteins, therefore treating cancer cells that usually escape from conventional cancer therapies. Recently, the attenuated Salmonella typhimurium strain was introduced with >10,000 virulence attenuation and ~1,000 tumor tropism in a variety of animal tumor models. The bacteria can be easily manipulated to introduce therapeutic genes and proteins for cancer treatment. However, in a human melanoma trial, the bacterial strain showed poor tumor tissue attraction, indicating that the bacteria injected i.v. in the cancer patients might be eliminated by human immune response. To resolve this issue, we hypothesize that genetic engineering of the attenuated Salmonella strain with t...
Proceedings of the IEEE 26th Annual Northeast Bioengineering Conference (Cat. No.00CH37114), 2000
A novel ENMR method-Convection Compensation ENMR (CC-ENMR)-has been developed, which permits sele... more A novel ENMR method-Convection Compensation ENMR (CC-ENMR)-has been developed, which permits selective detection of electrophoretic motion of ionic species. The convection generated ENMR spectral artifacts were removed by gradient moment nulling technique, whereas polarity switching of the applied DC electric field canceled out the flow compensation effect for ionic movement driven by the electric field.
High-resolution electrophoretic NMR (ENMR) resolves NMR spectra of mixtures on the basis of the e... more High-resolution electrophoretic NMR (ENMR) resolves NMR spectra of mixtures on the basis of the electrophoretic mobilities of contributing ions ( 1-4). This method is particularly convenient for the study of supramolecular systems, e.g., microemulsions, micelles, and vesicles. These systems tend to give large signals because of the large number of nuclei involved, the relatively long nuclear relaxation times, and the small mass diffusion coefficients. They are also polydisperse, and one of the aims of current research is to determine size and charge distributions for such particles. In this Communication we report the direct determination of velocity distributions for phospholipid vesicles in electric fields by a type of ENMR experiment previously suggested but not demonstrated (2). These vesicles consist of a single phospholipid bilayer in the form of a spherical shell that encloses an aqueous region ( 5). The ENMR experiments monitor both phospholipid protons in the bilayer and protons in entrapped molecules. Signals from water and from small molecules external to the vesicles are easily quenched by magnetic field gradients because of the large diffusion coefficients. The preliminary experiments reported here make use of the STE-ENMR sequence shown in Fig. 1 rather than the standard ENMR spin-echo sequence because T, b T2 for protons in the bilayer and because signals from entrapped molecules are degraded by the effects of J modulation (3). The new feature in this experiment is that 2D ENMR spectra are produced by Fourier transformations with respect to t2 and Edc rather than t2 and t I as in previously reported 2D NMR (2). The matched gradient pulses g, and g3 have areas G6, where G is the amplitude of the magnetic field gradient and 6 is the duration; and the gradient pulse gz in the time interval tl may be introduced to eliminate interference from the primary echo ( PE) and the secondary echoes ( SE lSE3). In the experiments described here it was more convenient to filter out unwanted coherence pathways by phase-cycling methods. The single-quantum coherence-transfer pathways for the stimulated-echo experiment are displayed in Ref. (6) with the exception of SE2 where p = 0 + 0 + + 1 + 1. Many different phase-cycling schemes are satisfactory. For sample I (described below) we cycled all three RF pulses with N,,* = 2 and ZV3 = 4 in the notation of Ref. (6) to eliminate PE, SEl-SE3, and various multiquantum pathways, while for sample II we cycled only RF pulse 3 with i’V3 = 4 to eliminate SE 1 -SE3 and experienced no problems.
Proceedings of the IEEE 26th Annual Northeast Bioengineering Conference (Cat. No.00CH37114)
Constant time electrophoretic NMR (CT-ENMR), an improved time incrementing method of ENMR, has be... more Constant time electrophoretic NMR (CT-ENMR), an improved time incrementing method of ENMR, has been developed to remove severe diffusion decay in the conventional time incrementing ENMR. The new method is useful to build complicated ENMR pulse sequences for structure characterization of proteins and protein interactions in solution.
Advances in Magnetic and Optical Resonance Volume 13, 1989
In the following sections we review the principles of ENMR, practical considerations, and applica... more In the following sections we review the principles of ENMR, practical considerations, and applications. We place special emphasis on present limitations and possibilities for improving the method
The frequency-selective multiple-quantum-coherence (Sel-MQC) lactate (Lac) filter offers complete... more The frequency-selective multiple-quantum-coherence (Sel-MQC) lactate (Lac) filter offers complete lipid and water suppression in a single scan for robust in vivo detection of tumor Lac, even in the presence of abundant lipids. Conversion of the detected signal into accurate tissue concentrations of Lac requires knowledge of in vivo Lac T1 and T2 relaxation times. This work reports modifications to the Sel-MQC pulse sequence, T1- and T2-Sel-MQC, that facilitate relaxation measurements of Lac. The T1-Sel-MQC sequence combines an inversion prepulse with the Sel-MQC filter. The T2-Sel-MQC sequence incorporates a CH3-selective 180 degrees pulse during the MQ preparation period to overcome the J-modulation effects and allow the insertion of variable echo delays. The performance of these sequences was evaluated with the use of phantoms and subcutaneous murine tumor models in vivo. The present approach will allow investigators to correct for the relaxation-induced Lac signal loss in Sel-MQC experiments for the quantitative mapping of in vivo tumor Lac distribution.
Multidimensional electrophoretic NMR (ENMR) has been introduced to determine structures of coexis... more Multidimensional electrophoretic NMR (ENMR) has been introduced to determine structures of coexisting proteins and protein conformations in solution. Signals of different proteins are separated in a new dimension of electrophoretic flow according to their characteristic electrophoretic mobilities. The electrophoretic interferograms have been generated in the flow dimension in two approaches by incrementing either the amplitude or the duration of the electric field. The ENMR method of incrementing the duration of the electric field, however, introduces severe signal decays due to molecular diffusion and spin relaxation, limiting the effectiveness of the method. In this study, an improved method of constant-time multidimensional ENMR (CT-ENMR) has been proposed and successfully tested. The time delays between the magnetic field gradients and the RF pulses are kept constant in this new method so that the molecular diffusion and spin relaxation processes contribute to only a constant factor of signal amplitude. As an alternative approach of incrementing the amplitude of the electric field, this novel method significantly enhances our capability and potential in characterizing structural changes of interacting proteins during biological signaling processes. The CT-ENMR method is particularly useful in studies where the amplitude-incrementing of the electric field is not optimal. For example, the CT-ENMR method is superior when the electric field is applied in the direction not parallel to the static magnetic field B(0) to the xy-magnetization. The new method was successfully demonstrated with a sample solution containing 100 mM 4,9-dioxa-1,12-dodecanediamine and 100 mM L-aspartic acid in D(2)O.
Cancer cells display heterogeneous genetic characteristics, depending on the tumor dynamic microe... more Cancer cells display heterogeneous genetic characteristics, depending on the tumor dynamic microenvironment. Abnormal tumor vasculature and poor tissue oxygenation generate a fraction of hypoxic tumor cells that have selective advantages in metastasis and invasion and often resist chemo- and radiation therapies. The genetic alterations acquired by tumors modify their biochemical pathways, which results in abnormal tumor metabolism. An elevation in glycolysis known as the “Warburg effect” and changes in lipid synthesis and oxidation occur. Magnetic resonance spectroscopy (MRS) has been used to study tumor metabolism in preclinical animal models and in clinical research on human breast, brain, and prostate cancers. This technique can identify specific genetic and metabolic changes that occur in malignant tumors. Therefore, the metabolic markers, detectable by MRS, not only provide information on biochemical changes but also define different metabolic tumor phenotypes. When combined wi...
Publisher Summary Blood flow and oxygenation control the physiological environment within tumors ... more Publisher Summary Blood flow and oxygenation control the physiological environment within tumors and are major determinants of the outcome of nonsurgical forms of cancer therapy. One of the main pathological characteristics of tumor blood flow is its heterogeneity, which is reflected in heterogeneous oxygen, pH, and metabolite distribution. The formation of necrosis, either by limitation of blood flow and substrates or by actual vascular collapse, has also been observed. Because blood flow, oxygenation, and metabolism are closely linked, noninvasive nuclear magnetic resonance (NMR) spectroscopic methods measuring the metabolism have been applied to the study of solid tumors to investigate the relationship of tumor metabolism to blood flow and oxygenation, proliferation, and differentiation. Magnetic resonance imaging (MRI) methods have also been applied for the advantage of volume localization and to determine, noninvasively, the morphologic structure of solid tumors and tumor perfusion. This chapter discusses some of the typical morphological, metabolic, and physiological characteristics of solid tumors and their relationship to 1H, 13C, and NMR measurements. One of the major limitations of the application of NMR spectroscopic methods in vivo is low sensitivity.
4809 The current cancer therapeutic modalities have encountered tremendous challenges in treating... more 4809 The current cancer therapeutic modalities have encountered tremendous challenges in treating hypoxic cancer cells due to poor drug delivery for effective chemotherapy or low level of tumor oxygenation for effective radiation therapy. Bacteria can penetrate into hypoxic tumor regions and deliver therapeutic proteins, therefore treating cancer cells that usually escape from conventional cancer therapies. Recently, the attenuated Salmonella typhimurium strain was introduced with >10,000 virulence attenuation and ~1,000 tumor tropism in a variety of animal tumor models. The bacteria can be easily manipulated to introduce therapeutic genes and proteins for cancer treatment. However, in a human melanoma trial, the bacterial strain showed poor tumor tissue attraction, indicating that the bacteria injected i.v. in the cancer patients might be eliminated by human immune response. To resolve this issue, we hypothesize that genetic engineering of the attenuated Salmonella strain with t...
Proceedings of the IEEE 26th Annual Northeast Bioengineering Conference (Cat. No.00CH37114), 2000
A novel ENMR method-Convection Compensation ENMR (CC-ENMR)-has been developed, which permits sele... more A novel ENMR method-Convection Compensation ENMR (CC-ENMR)-has been developed, which permits selective detection of electrophoretic motion of ionic species. The convection generated ENMR spectral artifacts were removed by gradient moment nulling technique, whereas polarity switching of the applied DC electric field canceled out the flow compensation effect for ionic movement driven by the electric field.
High-resolution electrophoretic NMR (ENMR) resolves NMR spectra of mixtures on the basis of the e... more High-resolution electrophoretic NMR (ENMR) resolves NMR spectra of mixtures on the basis of the electrophoretic mobilities of contributing ions ( 1-4). This method is particularly convenient for the study of supramolecular systems, e.g., microemulsions, micelles, and vesicles. These systems tend to give large signals because of the large number of nuclei involved, the relatively long nuclear relaxation times, and the small mass diffusion coefficients. They are also polydisperse, and one of the aims of current research is to determine size and charge distributions for such particles. In this Communication we report the direct determination of velocity distributions for phospholipid vesicles in electric fields by a type of ENMR experiment previously suggested but not demonstrated (2). These vesicles consist of a single phospholipid bilayer in the form of a spherical shell that encloses an aqueous region ( 5). The ENMR experiments monitor both phospholipid protons in the bilayer and protons in entrapped molecules. Signals from water and from small molecules external to the vesicles are easily quenched by magnetic field gradients because of the large diffusion coefficients. The preliminary experiments reported here make use of the STE-ENMR sequence shown in Fig. 1 rather than the standard ENMR spin-echo sequence because T, b T2 for protons in the bilayer and because signals from entrapped molecules are degraded by the effects of J modulation (3). The new feature in this experiment is that 2D ENMR spectra are produced by Fourier transformations with respect to t2 and Edc rather than t2 and t I as in previously reported 2D NMR (2). The matched gradient pulses g, and g3 have areas G6, where G is the amplitude of the magnetic field gradient and 6 is the duration; and the gradient pulse gz in the time interval tl may be introduced to eliminate interference from the primary echo ( PE) and the secondary echoes ( SE lSE3). In the experiments described here it was more convenient to filter out unwanted coherence pathways by phase-cycling methods. The single-quantum coherence-transfer pathways for the stimulated-echo experiment are displayed in Ref. (6) with the exception of SE2 where p = 0 + 0 + + 1 + 1. Many different phase-cycling schemes are satisfactory. For sample I (described below) we cycled all three RF pulses with N,,* = 2 and ZV3 = 4 in the notation of Ref. (6) to eliminate PE, SEl-SE3, and various multiquantum pathways, while for sample II we cycled only RF pulse 3 with i’V3 = 4 to eliminate SE 1 -SE3 and experienced no problems.
Proceedings of the IEEE 26th Annual Northeast Bioengineering Conference (Cat. No.00CH37114)
Constant time electrophoretic NMR (CT-ENMR), an improved time incrementing method of ENMR, has be... more Constant time electrophoretic NMR (CT-ENMR), an improved time incrementing method of ENMR, has been developed to remove severe diffusion decay in the conventional time incrementing ENMR. The new method is useful to build complicated ENMR pulse sequences for structure characterization of proteins and protein interactions in solution.
Advances in Magnetic and Optical Resonance Volume 13, 1989
In the following sections we review the principles of ENMR, practical considerations, and applica... more In the following sections we review the principles of ENMR, practical considerations, and applications. We place special emphasis on present limitations and possibilities for improving the method
The frequency-selective multiple-quantum-coherence (Sel-MQC) lactate (Lac) filter offers complete... more The frequency-selective multiple-quantum-coherence (Sel-MQC) lactate (Lac) filter offers complete lipid and water suppression in a single scan for robust in vivo detection of tumor Lac, even in the presence of abundant lipids. Conversion of the detected signal into accurate tissue concentrations of Lac requires knowledge of in vivo Lac T1 and T2 relaxation times. This work reports modifications to the Sel-MQC pulse sequence, T1- and T2-Sel-MQC, that facilitate relaxation measurements of Lac. The T1-Sel-MQC sequence combines an inversion prepulse with the Sel-MQC filter. The T2-Sel-MQC sequence incorporates a CH3-selective 180 degrees pulse during the MQ preparation period to overcome the J-modulation effects and allow the insertion of variable echo delays. The performance of these sequences was evaluated with the use of phantoms and subcutaneous murine tumor models in vivo. The present approach will allow investigators to correct for the relaxation-induced Lac signal loss in Sel-MQC experiments for the quantitative mapping of in vivo tumor Lac distribution.
Multidimensional electrophoretic NMR (ENMR) has been introduced to determine structures of coexis... more Multidimensional electrophoretic NMR (ENMR) has been introduced to determine structures of coexisting proteins and protein conformations in solution. Signals of different proteins are separated in a new dimension of electrophoretic flow according to their characteristic electrophoretic mobilities. The electrophoretic interferograms have been generated in the flow dimension in two approaches by incrementing either the amplitude or the duration of the electric field. The ENMR method of incrementing the duration of the electric field, however, introduces severe signal decays due to molecular diffusion and spin relaxation, limiting the effectiveness of the method. In this study, an improved method of constant-time multidimensional ENMR (CT-ENMR) has been proposed and successfully tested. The time delays between the magnetic field gradients and the RF pulses are kept constant in this new method so that the molecular diffusion and spin relaxation processes contribute to only a constant factor of signal amplitude. As an alternative approach of incrementing the amplitude of the electric field, this novel method significantly enhances our capability and potential in characterizing structural changes of interacting proteins during biological signaling processes. The CT-ENMR method is particularly useful in studies where the amplitude-incrementing of the electric field is not optimal. For example, the CT-ENMR method is superior when the electric field is applied in the direction not parallel to the static magnetic field B(0) to the xy-magnetization. The new method was successfully demonstrated with a sample solution containing 100 mM 4,9-dioxa-1,12-dodecanediamine and 100 mM L-aspartic acid in D(2)O.
Cancer cells display heterogeneous genetic characteristics, depending on the tumor dynamic microe... more Cancer cells display heterogeneous genetic characteristics, depending on the tumor dynamic microenvironment. Abnormal tumor vasculature and poor tissue oxygenation generate a fraction of hypoxic tumor cells that have selective advantages in metastasis and invasion and often resist chemo- and radiation therapies. The genetic alterations acquired by tumors modify their biochemical pathways, which results in abnormal tumor metabolism. An elevation in glycolysis known as the “Warburg effect” and changes in lipid synthesis and oxidation occur. Magnetic resonance spectroscopy (MRS) has been used to study tumor metabolism in preclinical animal models and in clinical research on human breast, brain, and prostate cancers. This technique can identify specific genetic and metabolic changes that occur in malignant tumors. Therefore, the metabolic markers, detectable by MRS, not only provide information on biochemical changes but also define different metabolic tumor phenotypes. When combined wi...
Publisher Summary Blood flow and oxygenation control the physiological environment within tumors ... more Publisher Summary Blood flow and oxygenation control the physiological environment within tumors and are major determinants of the outcome of nonsurgical forms of cancer therapy. One of the main pathological characteristics of tumor blood flow is its heterogeneity, which is reflected in heterogeneous oxygen, pH, and metabolite distribution. The formation of necrosis, either by limitation of blood flow and substrates or by actual vascular collapse, has also been observed. Because blood flow, oxygenation, and metabolism are closely linked, noninvasive nuclear magnetic resonance (NMR) spectroscopic methods measuring the metabolism have been applied to the study of solid tumors to investigate the relationship of tumor metabolism to blood flow and oxygenation, proliferation, and differentiation. Magnetic resonance imaging (MRI) methods have also been applied for the advantage of volume localization and to determine, noninvasively, the morphologic structure of solid tumors and tumor perfusion. This chapter discusses some of the typical morphological, metabolic, and physiological characteristics of solid tumors and their relationship to 1H, 13C, and NMR measurements. One of the major limitations of the application of NMR spectroscopic methods in vivo is low sensitivity.
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