Molecular imaging of neurodegeneration by a novel cross-disease biomarker
Introduction
Diagnosis and management of neurodegenerative disorders such as Alzheimer's disease (AD), Parkinson's disease (PD), or amyotrophic lateral sclerosis (ALS) are paramount challenges in clinical neurology. While differing in clinical presentations, genetic predisposing factors or histopathological substrates, all these neurological disorders are characterized by progressive and relentless processes of loss of neuronal cell populations within the central nervous system (CNS), leading to severe neurological deficits.
Current diagnosis of these neurological disorders is based on clinical assessment of related symptoms and signs. However, these clinical features emerge only late in the disease course, and consequently, diagnosis of neurodegeneration is nearly always made rather late, thus confounding development and potential implementation of any effective neuroprotective strategy. At such late stage, the vast majority of target neuronal cells have already been lost. Therefore, novel strategies for early diagnosis of neurodegeneration are urgently needed, and are anticipated to play an important role in future disease management.
While each neurodegenerative disorder has its own distinctive characteristics, it is well recognized that there is an overlap between various disorders, both in clinical presentations and in histopathological features. For example, cognitive decline, the hallmark of AD, has also been associated with ALS (Woolley and Jonathan, 2008).As another example, Lewy bodies, one of the hallmarks of PD neuropathology, is also the hallmark of Lewy body dementia (Dodel et al., 2008). In addition, general neuropathological processes, such as abnormal protein aggregation, excitotoxicity, oxidative stress or apoptosis, have been shown to play a role in neuronal cell death across various neurodegenerative disorders (Goodall and Morrison, 2006, Parihar and Hemnani, 2004, Wood et al., 2003, Yacoubian and Standaert, 2008). Conceivably, these considerations call for the development of cross-disease molecular imaging agents, applicable as general probes for neurodegeneration, and which will be useful for early diagnosis or monitoring of treatment of various neurodegenerative disorders. However, such cross-disease biomarkers are currently not available in clinical neurological practice. The current molecular imaging tools in neurology are mostly pathology-specific, e.g., imaging amyloid plaque pathology in AD (Braskie et al., 2008, Ono et al., 2006, Ono et al., 2008) or dopaminergic neuronal loss in PD (Kassiou et al., 2009). The challenge of early diagnosis of other important neurodegenerative disorders such as ALS, or disorders with combined or overlapping features, is currently unaddressed.
Apoptosis, as one of the most fundamental biological processes, has been implicated in the etiology and pathogenesis of neurodegenerative disorders (Okouchi et al., 2007). ApoSense is a novel class of low molecular weight probes, developed for imaging of apoptosis in vivo. Previously, performance of the ApoSense compounds, labeled by either a fluorophore or a radio-isotope (3H or 18F) in imaging of apoptosis has been demonstrated in a wide array of pre-clinical models of disease, such as cancer, cerebral stroke, traumatic brain injury, or renal failure (Aloya et al., 2006, Cohen et al., 2007, Cohen et al., 2009, Damianovich et al., 2006, Grimberg et al., 2009, Reshef et al., 2007, Reshef et al., 2008a, Reshef et al., 2008b). 18F-ML-10, a positron emission tomography (PET) member of this family of biomarkers of apoptosis, has completed a Phase II clinical trial for imaging of neurovascular cell death in patients with acute ischemic cerebral stroke and is now in Phase II study for early detection of response of brain metastases to radiation therapy. All members of this novel class of molecular probes respond to a complex of cellular features, unique to cells in early apoptosis, which comprises irreversible loss of trans-membrane potential, permanent acidification of the outer plasma membrane and cytosol, and activation of membrane phospholipid scramblase system. In response to this set of cellular features, these biomarkers cross the cell membrane and accumulate within the cytoplasm of the apoptotic cell, from the early stages of the death program (Aloya et al., 2006, Cohen et al., 2007, Cohen et al., 2009, Damianovich et al., 2006, Grimberg et al., 2009, Reshef et al., 2007, Reshef et al., 2008a, Reshef et al., 2008b).
We now report on the performance of the ApoSense compound NST-729 (MW = 310; Fig. 1A) in targeting foci of disease in vivo, in pre-clinical models of AD (Tg2576 transgenic mice) and ALS (transgenic SOD-1 G93A mutation mice). In the reported proof-of-concept experiments, the probe was administered intravenously in vivo, followed by ex vivo demonstration of its uptake in relation with the characteristic neuropathology, utilizing the inherent fluorescent properties of the molecule.
Section snippets
Synthesis of NST-729 and the “mutated” analogue NST-727
NST-729 [n-(2-mercaptoethyl)-dansylamide MW = 310, Fig. 1A], was synthesized via didansylation of cystamine in homogeneous acetonitrile–water solution, in the presence of potassium carbonate. The didansyl derivative was then reduced in water–methanol solution by dithiothreitol, to afford NST-729 in 54% yield and high purity, as found by ESI-MS and 1H-NMR. In order to demonstrate structure/function relationship of this compound, we synthesized in addition to NST-729, also a “mutated” NST-729
AD model: Tg2576 transgenic mice
Examination of the brains of the Tg2576 transgenic mice revealed numerous amyloid plaques, being eosinophilic in the H&E staining, and which were distinctly stained with Congo red and visualized by polarized light (Figs. 3B, C). Such plaques were not found in the age-matched controls. Plaques were predominantly found in cortical areas, but also in the hippocampus and subcortical regions. Plaques were not found in the cerebellum. These amyloid plaques manifested marked uptake of NST-729 (Figs. 2
Discussion
Brain plasticity and neuronal tissue reserves enable preservation of neurological functions in situations of substantial loss of neuronal cell populations. Unfortunately, this important aspect of brain physiology often precludes early diagnosis of neurodegenerative disorders until late in the disease course, and by the time symptoms and signs emerge, only palliative treatments can be offered to the patients. Therefore, the development and implementation of more effective, neuroprotective
Acknowledgments
NST-729 is an investigational imaging agent developed by Aposense Ltd., (formerly NST Ltd.), and all authors receive personal compensation from Aposense Ltd. The authors would like to thank Dr. Ari Barzilai, from the Department of Neurobiochemistry, George S Wise Faculty of Life Sciences, Tel-Aviv University, Israel, for his assistance in the in vitro model.
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2014, Pharmacology and TherapeuticsCitation Excerpt :Imaging studies showed an age-dependent accumulation of Aβ in brain areas also affected in AD patients (Opazo et al., 2006; Muller et al., 2013) with the caveat that not each tracer effective in clinical practice gives a positive signal in this model (Kuntner et al., 2009; Snellman et al., 2013). Histological analyses of the brains of aged Tg2576 mice reveal similar changes to those found in AD patients: a large number of amyloid plaques associated with apoptosis (Shirvan et al., 2009; Wati et al., 2009; Shevchenko et al., 2012) dystrophic neurons (Woodhouse et al., 2009) and ubiquitin, as well as alpha-synuclein positive neurites (Yang et al., 2000). In the brain of aged transgenic Tg2576 mice, a significant reduction in the number of cholinergic (Apelt et al., 2002; Luth et al., 2003; Wenk et al., 2004; Watanabe et al., 2013) and adrenergic (Guerin et al., 2009) neurons was detected.
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These authors equally contributed to this paper.