New radiotracer development to study immune cell mobilization of granzyme proteolytic activity
开发新的放射性示踪剂来研究免疫细胞动员颗粒酶蛋白水解活性
基本信息
- 批准号:10395587
- 负责人:
- 金额:$ 74.04万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2021
- 资助国家:美国
- 起止时间:2021-04-20 至 2025-03-31
- 项目状态:未结题
- 来源:
- 关键词:Amino AcidsAnimal ModelApoptoticBacteriaBacterial InfectionsBiochemistryBiodistributionBiological AssayBiological ModelsBiologyBypassCell DeathCell membraneCellsCoculture TechniquesCommunicable DiseasesComplexCytokine ActivationCytosolCytotoxic T-LymphocytesDataDevelopmentDiseaseDrug KineticsEndotoxinsEnzymesEscherichia coliExtracellular SpaceFluorescence Resonance Energy TransferFoundationsGoalsGranzymeHost DefenseHumanHuman ActivitiesImageImmuneImmune responseImmune systemImmunologyIn VitroInflammationInflammatory ResponseInterceptKlebsiella pneumoniaeKnockout MiceListeria monocytogenesLymphocyteMacrophage ActivationMasksMass Spectrum AnalysisMeasurementMeasuresMembraneMethodologyModelingMouse StrainsMusMycobacterium marinumMycobacterium tuberculosisMyositisNatureNerve DegenerationOrganismPathogenicityPeptide HydrolasesPeptidesPharmaceutical ChemistryPharmacologyPhospholipidsPositron-Emission TomographyProteolysisPseudomonas aeruginosaRadioactiveRadiolabeledResearchResolutionRodentRoleSafetySerine ProteaseSignal TransductionSpecificityStaphylococcus aureusTechnologyTestingTimeTryptaseViral PneumoniaVirus DiseasesXenobioticsanaloganti-cancerbasecancer cellclinical applicationclinically relevantclinically significantcombatcytotoxicdesigndosimetryenzyme activityexperimental studyextracellularhuman imagingimaging approachimaging probeimaging studyimmunological synapsein vivoin vivo imaginginterestlead candidatenovelnuclear imagingoptical imagingpathogenpathogenic virusperforinpre-clinicalpreclinical studyradiochemicalradiotracerresponsesystemic autoimmune diseasetomographytooltrafficking
项目摘要
Project Abstract:
The human granzymes are a somewhat mystical class of five serine proteases (A, B, H, K, M) that are expressed
and conditionally secreted by select lymphocytes like natural killer (NK) and cytotoxic T cells (CTL). Based
largely on data for A and B, granzymes have been historically regarded as pro-apoptotic effectors whereby they
are presented transiently by NK and CTLs at the immunological synapse with a problematic target cell (e.g.
cancer cell, pathogen infected cell), and immediately shuttled into the cytosol via perforin to initiate cell death.
However, emerging data has begun to challenge this canon and present a more complex model in which
secreted granzymes perform essential signaling functions in extracellular space, including activation of
macrophages as part of host defense. Interestingly, dysregulation of granzyme presentation and/or proteolytic
activity may also underlie the pathobiology of debilitating diseases like neurodegenerative or systemic
autoimmune diseases. Thus, more fully elaborating the biology of granzymes is an important unmet need, and
requires technologies to study granzymes in the most clinically relevant animal models and humans themselves.
To this end, we developed a novel imaging approach we term “restricted interaction peptides” (RIP) to detect
granzyme proteolytic activity in vivo with PET. Mechanistically, RIPs are administered as an inactive pro-form,
whereupon internal cleavage of the RIP by the target endoprotease releases a radiolabeled peptide that
immediately associates with nearby phospholipid membranes (i.e. the plasma membrane of the target cell).
Thus, accumulation of the radiolabeled cleavage product adjacent to the endoprotease provides a readout of the
relative units of enzyme activity within a region of interest. As preliminary data, we developed 64Cu-GB1, a RIP
that measures granzyme B activity on PET. Our proof of concept data show that 64Cu-GB1 detects the cytotoxic
pool of granzyme B activity trafficking to pathogenic target cells, but also an unexpected non-cytotoxic pool
elicited as part of an inflammatory response to an endotoxin. During this project, we will expand upon these
data in several important directions. First, we will test in Aim 1 if 64Cu-GB1 can be applied to study granzyme B
proteolytic activity in the spontaneous immune responses arising due to pathogen stimulation. These data will
be crucial to understanding the utility of this methodology beyond simply detecting pharmacologically augmented
anticancer immune responses. During Aim 2, we will expand the RIP toolkit to develop and study probes
targeting the tryptase proteolytic activities of granzymes A and K. During the final Aim, we will carry out the late
stage preclinical experiments required to judge the feasibility of human imaging with RIP probes. As a model
system, we will study our lead candidate 64Cu-GB1. If successful, this project will confer new translational
technologies at a crucial inflection point away from the classical view as simply pro-apoptotic effectors toward
multifaceted regulators of human immunology and host defense against pathogens.
项目摘要:
人类颗粒酶是一个有点神秘的五种丝氨酸蛋白酶(A,B,H,K,M),
并由选择的淋巴细胞如自然杀伤细胞(NK)和细胞毒性T细胞(CTL)有条件地分泌。基于
主要基于A和B的数据,颗粒酶在历史上被认为是促凋亡效应物,
由NK和CTL在免疫突触处与有问题的靶细胞(例如,
癌细胞、病原体感染的细胞),并立即通过穿孔素穿梭到细胞质中以启动细胞死亡。
然而,新兴数据已经开始挑战这一标准,并提出了一个更复杂的模型,
分泌的颗粒酶在细胞外空间中执行必需的信号传导功能,包括激活
巨噬细胞作为宿主防御的一部分。有趣的是,颗粒酶呈递和/或蛋白水解的失调
活性也可能是使人衰弱的疾病,如神经退行性疾病或系统性疾病的病理学基础。
自身免疫性疾病因此,更充分地阐述颗粒酶的生物学是一个重要的未满足的需求,
需要技术来研究最临床相关的动物模型和人类本身的颗粒酶。
为此,我们开发了一种新的成像方法,我们称之为“限制性相互作用肽”(RIP)来检测
颗粒酶蛋白水解活性与PET。从机制上讲,RIP是作为一种无活性的前体形式给药的,
于是RIP被靶内切蛋白酶内部切割释放出放射性标记的肽,
立即与附近的磷脂膜(即靶细胞的质膜)结合。
因此,邻近内切蛋白酶的放射性标记的切割产物的积累提供了内切蛋白酶的读数。
在感兴趣的区域内的酶活性的相对单位。作为初步数据,我们开发了64 Cu-GB 1,
在PET上测量颗粒酶B活性。我们的概念验证数据表明,64 Cu-GB 1检测到细胞毒性
颗粒酶B活性池运输至致病性靶细胞,但也是非预期的非细胞毒性池
作为对内毒素的炎症反应的一部分引起的。在这个项目中,我们将扩大这些
在几个重要方向的数据。首先,我们将在目的1中检验~(64)Cu-GB_1能否用于研究颗粒酶B
在由于病原体刺激引起的自发免疫应答中的蛋白水解活性。这些数据将
对于理解这种方法的实用性至关重要,而不仅仅是检测增强的
抗癌免疫反应。在目标2中,我们将扩展RIP工具包以开发和研究探针
针对颗粒酶A和K的类胰蛋白酶蛋白水解活性。在最后的目标,我们将进行后期
阶段临床前实验需要判断的可行性与RIP探头的人体成像。为模特
系统,我们将研究我们的领先候选人64 Cu-GB 1。如果成功,该项目将提供新的翻译
技术在一个关键的转折点远离经典的观点,作为简单的促凋亡效应器,
人类免疫学和宿主防御病原体的多方面调节剂。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Charles Scott Craik其他文献
Charles Scott Craik的其他文献
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{{ truncateString('Charles Scott Craik', 18)}}的其他基金
Developing Antivirals Targeting Proteases and Polymerases of Coronaviruses, Picornaviruses and Bunyavirales
开发针对冠状病毒、小核糖核酸病毒和布尼亚病毒的蛋白酶和聚合酶的抗病毒药物
- 批准号:
10512628 - 财政年份:2022
- 资助金额:
$ 74.04万 - 项目类别:
New radiotracer development to study immune cell mobilization of granzyme proteolytic activity
开发新的放射性示踪剂来研究免疫细胞动员颗粒酶蛋白水解活性
- 批准号:
10231735 - 财政年份:2021
- 资助金额:
$ 74.04万 - 项目类别:
New radiotracer development to study immune cell mobilization of granzyme proteolytic activity
开发新的放射性示踪剂来研究免疫细胞动员颗粒酶蛋白水解活性
- 批准号:
10591415 - 财政年份:2021
- 资助金额:
$ 74.04万 - 项目类别:
Probing the Role of Chaperone-TPR Complexes in Tau Proteostasis
探讨分子伴侣-TPR 复合物在 Tau 蛋白质稳态中的作用
- 批准号:
10029781 - 财政年份:2020
- 资助金额:
$ 74.04万 - 项目类别:
Non-invasive Differentiation of Benign Lesions from Aggressive Pancreatic Cancer
良性病变与侵袭性胰腺癌的无创鉴别
- 批准号:
8823694 - 财政年份:2015
- 资助金额:
$ 74.04万 - 项目类别:
Extracellular Proteolysis as a Molecular Stratification Tool for Cancer
细胞外蛋白水解作为癌症的分子分层工具
- 批准号:
8829207 - 财政年份:2014
- 资助金额:
$ 74.04万 - 项目类别:
Antibodies for Characterizing the Structure and Function of Proteases
用于表征蛋白酶结构和功能的抗体
- 批准号:
8702411 - 财政年份:2014
- 资助金额:
$ 74.04万 - 项目类别:
Allosteric Inhibition of a Family of Proteolytic Enzymes
蛋白水解酶家族的变构抑制
- 批准号:
8577916 - 财政年份:2013
- 资助金额:
$ 74.04万 - 项目类别:
Allosteric Inhibition of a Family of Proteolytic Enzymes
蛋白水解酶家族的变构抑制
- 批准号:
8698774 - 财政年份:2013
- 资助金额:
$ 74.04万 - 项目类别:
Allosteric Inhibition of a Family of Proteolytic Enzymes
蛋白水解酶家族的变构抑制
- 批准号:
9039629 - 财政年份:2013
- 资助金额:
$ 74.04万 - 项目类别:
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