NMR and NIR Detection of Phospholipase Activity In Vivo

体内磷脂酶活性的 NMR 和 NIR 检测

• 批准号: 7812187
• 负责人: EDWARD J DELIKATNY
• 金额: $ 28.22万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2013-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7812187
• 关键词: Achievement; Address; Alzheimer' s Disease; Animals; Antineoplastic Agents; Arthritis; Autoimmune Process; Biochemical; Biochemical Pathway; Biomedical Engineering; Cancer cell line; Catalysis; Cell Line; Cells; Characteristics; Choline; Coupled; Data; Detection; Development; Disease; Early Diagnosis; Enzymes; Equilibrium; Event; Fluorescence; Glycerol; Goals; Human; Hydrolysis; Image; Imaging Techniques; In Situ; In Vitro; Inflammatory; Kinetics; Length; Lipids; Magnetic Resonance; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Malignant Neoplasms; Malignant neoplasm of prostate; Measurement; Measures; Metabolic; Metabolism; Methods; Modality; Modeling; Molecular; Monitor; Non-Hodgkin' s Lymphoma; Organ; Pathology; Pathway interactions; Pharmacotherapy; Phospholipase; Phospholipase A2; Phospholipase C; Phospholipids; Phosphorylcholine; Physiological; Positioning Attribute; Process; Protein Isoforms; Recurrence; Reporter; Research; Sensitivity and Specificity; Specificity; Spectrum Analysis; Staging; Structure; Techniques; Testing; Therapeutic; Tissues; Tube; United States National Institutes of Health; Vertebral column; Whole Organism; Xenograft Model; bacteriochlorin; base; biological systems; clinical Diagnosis; design; fluorophore; imaging modality; imaging probe; improved; in vivo; lipid metabolism; molecular imaging; optical imaging; phosphodiester; phosphoethanolamine; phosphomonoester; prototype; public health relevance; response; small molecule; tumor; tumor progression; tumor xenograft

DESCRIPTION (provided by applicant): The overall aim of this application is to develop methods to image the expression and activity of phospholipases in vivo. A multi-modality approach with sequential magnetic resonance imaging/spectroscopy (MRI/MRS) and optical imaging using near infrared (NIR) fluorophores will be employed to obtain information that is not available from either technique separately. The key hypothesis to be addressed is that in vivo activation of phospholipases can be measured by determining the changes in MR-visible lipid metabolite levels together with measuring fluorescence released by enzymatic hydrolysis of a self- quenching NIR phospholipid fluorophore. We will compare and validate the MR spectroscopy with results obtained using NIR optical imaging in tumor models in which changes in choline metabolite levels associated with tumor progression and response to therapy are well known. Since phospholipases have been implicated in a wide range of pathologies including cancer, arthritis and Alzheimer's disease, this study will supply critical in vivo data on the specific biochemical pathways involved in these diseases. The hypothesis will be tested by the achievement of the following specific aims: Aim 1): Design and synthesis of chiral lipid-based self-quenching fluorophores containing a stable pyropheophorbide NIR fluorophore, coupled to a dark quencher. The fluorophore will be attached to different positions of the glycerol backbone to interrogate the activity of different phospholipase isoforms. The quencher linkage will be attached at different points on the glycerol backbone to distinguish between phospholipase types. Structure-activity studies will be performed by synthetically inserting linkers of various length to determine the effects of steric hindrance on probe activity. Aim 2): In vitro characterization of the synthesized probes. This will include test-tube demonstration of the sensitivity and specificity of the probes to various phospholipase types and isoform and detailed measurement of kinetics of catalysis and inhibition. We will further characterize the delivery and activation of these self- quenching fluorophores in cultured cell lines. Aim 3): Demonstration of in vivo delivery, bio-distribution and subsequent activation of phospholipases in tumor xenograft models of prostate cancer and non-Hodgkin's lymphoma, followed by dual modality molecular imaging using 1H MR spectroscopy and optical imaging. These data will supply critical information on the specific lipid catabolic pathways involved in the onset and subsequent treatment of disease. PUBLIC HEALTH RELEVANCE: To develop near-infrared fluorescent beacons for the in vivo detection of phospholipase activity via dual optical imaging and MR spectroscopy.

描述（由申请人提供）：本申请的总体目标是开发对磷脂酶的体内表达和活性进行成像的方法。将采用顺序磁共振成像/光谱（MRI/MRS）和使用近红外（NIR）荧光团的光学成像的多模态方法来获得单独使用任一技术无法获得的信息。要解决的关键假设是，磷脂酶的体内活化可以通过测定MR可见脂质代谢物水平的变化以及测量由自猝灭NIR磷脂荧光团的酶水解释放的荧光来测量。我们将比较和验证MR光谱与肿瘤模型中使用NIR光学成像获得的结果，其中胆碱代谢物水平的变化与肿瘤进展和对治疗的反应是众所周知的。由于磷脂酶已被牵连在广泛的病理，包括癌症，关节炎和阿尔茨海默氏病，这项研究将提供关键的体内数据的具体生化途径参与这些疾病。该假设将通过实现以下具体目标来检验：目标1）：设计和合成手性基于脂质的自猝灭荧光团，其含有稳定的焦脱镁叶绿酸NIR荧光团，与暗猝灭剂偶联。荧光团将连接到甘油骨架的不同位置，以询问不同磷脂酶同种型的活性。猝灭剂键将连接在甘油主链上的不同点处以区分磷脂酶类型。将通过合成插入不同长度的接头来进行结构-活性研究，以确定空间位阻对探针活性的影响。目的2）：合成探针的体外表征。这将包括试管演示探针对各种磷脂酶类型和亚型的灵敏度和特异性，以及催化和抑制动力学的详细测量。我们将进一步表征这些自猝灭荧光团在培养细胞系中的递送和激活。目标3）：证明磷脂酶在前列腺癌和非霍奇金淋巴瘤的肿瘤异种移植模型中的体内递送、生物分布和随后的活化，随后使用1H MR光谱和光学成像进行双模态分子成像。这些数据将提供关键信息的具体脂质分解代谢途径参与发病和随后的治疗疾病。公共卫生关系：开发近红外荧光信标，用于通过双光学成像和MR光谱法在体检测磷脂酶活性。