Seok Rye Choi

Preclinical Properties of 18F-AV-45: A PET Agent for Aβ Plaques in the Brain

β-amyloid plaques (Aβ plaques) in the brain, containing predominantly fibrillary Aβ peptide aggregates, represent a defining pathologic feature of Alzheimer disease (AD). Imaging agents targeting the Aβ plaques in the living human brain are potentially valuable as biomarkers of pathogenesis processes in AD. (E)-4-(2-(6-(2-(2-(2-18F-fluoroethoxy)ethoxy)ethoxy)pyridin-3-yl)vinyl)-N-methyl benzenamine (18F-AV-45) is such as an agent currently in phase III clinical studies for PET of Aβ plaques in the brain. Methods: In vitro binding of 18F-AV-45 to Aβ plaques in the postmortem AD brain tissue was evaluated by in vitro binding assay and autoradiography. In vivo biodistribution of 18F-AV-45 in mice and ex vivo autoradiography of AD transgenic mice (APPswe/PSEN1) with Aβ aggregates in the brain were performed. Small-animal PET of a monkey brain after an intravenous injection of 18F-AV-45 was evaluated. Results: 18F-AV-45 displayed a high binding affinity and specificity to Aβ plaques (Kd, 3.72 ± 0.30 nM). In vitro autoradiography of postmortem human brain sections showed substantial plaque labeling in AD brains and not in the control brains. Initial high brain uptake and rapid washout from the brain of healthy mice and monkey were observed. Metabolites produced in the blood of healthy mice after an intravenous injection were identified. 18F-AV-45 displayed excellent binding affinity to Aβ plaques in the AD brain by ex vivo autoradiography in transgenic AD model mice. The results lend support that 18F-AV-45 may be a useful PET agent for detecting Aβ plaques in the living human brain.

18F Stilbenes and Styrylpyridines for PET Imaging of Aβ Plaques in Alzheimer’s Disease: A Miniperspective

Alzheimer’s disease (AD) is a neurodegenerative disease of the brain, characterized by a slowly progressive dementia. This insidious disease is growing in importance because it affects millions of older patients. Clinical symptoms of AD include cognitive decline, irreversible memory loss, disorientation and language impairment. Major neuropathology observations of postmortem AD brain include the presence of senile plaques containing β-amyloid (Aβ) aggregates and neurofibrillary tangles containing highly phosphorylated tau proteins (Figure 1A).1,2 Several genomic factors have been linked to AD. Familial AD (or early onset AD) has been reported to have mutations in genes encoding β-amyloid precursor protein (APP), presenilin 1, presenilin 2 and Apolipoprotein E (APOE).3 The exact mechanisms of these mutations, which lead to the development of AD, are not fully understood; however, formation of plaques comprised of Aβ peptide in the brain is a pivotal event in the pathology of Alzheimer’s disease. Significant evidence suggests that accumulation and aggregation of Aβ peptides may play a major causative role in AD pathogenesis.2,4 The excessive burden of Aβ, produced by various mechanisms, may represent the starting point of neurodegenerative events, and may initiate a cascade of events (β-amyloid cascade, Figure 1B) that includes gliosis, inflammatory changes, neuritic/synaptic change, tangles and transmitter loss.2 Currently, there is no definitive method to diagnose AD, except by postmortem evaluation and staining of the brain tissue, which demonstrates the existence of Aβ plaques. Figure 1 A. Processes (β-amyloid cascade) participating in AD pathogenesis. Aβ peptides produced by neurons aggregate into a variety of assemblies, some of which may impair synapses and neuronal dendrites. Build-up of pathogenic Aβ aggregates ... Recent reports have suggested that β-amyloid aggregates in the brain play a key role in a cascade of events leading to AD.2,5 Thus, the development of diagnostic imaging agents targeting Aβ aggregates is very important in the diagnosis and treatment of AD. Novel PET imaging agents specifically targeting the Aβ plaques may lead to early detection of AD pathology, differential diagnosis of patients with dementia, and for monitoring patients who are undergoing drug treatment designed to reverse the Aβ buildup in the brain. Indeed, diagnosis and treatment of AD have been hampered by the absence of reliable non-invasive markers for the underlying pathology. Diagnosis based on consensus criteria is approximately 81% sensitive and 70% specific by comparison to the gold standard of pathology at autopsy.6 In addition to errors of misdiagnosis in patients with AD, there is significant under diagnosis; approximately 10% of community dwelling elderly still have undiagnosed dementia, and community physicians may fail to diagnose up to 33% of mild dementia cases.7 Thus, there is a need for a biomarker that can be applied in the community setting and can help physicians separate those patients who do not have AD from those who have pathological signs and should be evaluated further. Additionally, there are a large number of patients who, upon comprehensive diagnostic testing, are found to have cognitive impairment but are not demented and thus, do not meet diagnostic criteria for AD (e.g., patients with mild cognitive impairment, MCI). Some, but not all of these patients will go on to develop AD within 3-5 years. A reliable biomarker might aid prognostic evaluation by documenting the presence or absence of AD related pathology. Based on the definitions of AD endorsed by the American Academy of Neurology, American Psychiatric Association (DSM-IV) and others, patients without abnormal amyloid plaque levels do not meet currently accepted neuropathological criteria for AD. This definition of AD, which includes amyloid plaques as a required feature, is supported by more than 100 years of autopsy data. Therefore, based on this widely-endorsed definition of AD, the use of a test for ruling-out the presence of amyloid plaque pathology in subjects with clinical signs and symptoms of cognitive impairment will, effectively, rule-out the diagnosis of AD, and lead to more careful evaluation and appropriate treatment for alternative causes of cognitive deficits. Moreover, the use of a test for ruling-in the presence of abnormal levels of Aβ plaques in the brain of subjects with signs and symptoms of cognitive impairment will lead to the selection of patients who warrant more detailed work-up for the possible diagnosis of AD or MCI. The differential diagnosis for AD includes a large number of other diseases. At early stages of disease (e.g. MCI), frequent confounds include cognitive impairment as a result of underlying depression, effects of CNS active medications, inadequately treated or end stage medical conditions affecting other organ systems, and even normal age-related changes. At later stages of disease, more common confounds include vascular dementia, frontal temporal lobar dementia (FTLD) complex, dementia with Lewy bodies (DLB) as well as rarer neurodegenerative diseases such as Creutzfeld Jacob Disease (CJD). Importantly, AD subjects will always have Aβ plaques, whereas amyloid is seen not at all, or only sporadically in most of these other diseases. In each case, appropriate prognosis and treatment requires accurate diagnostic assessment.

Correlation of amyloid PET ligand florbetapir F 18 binding with Aβ aggregation and neuritic plaque deposition in postmortem brain tissue.

BackgroundFlorbetapir F 18 (18F-AV-45) is a positron emission tomography imaging ligand for the detection of amyloid aggregation associated with Alzheimer disease. Earlier data showed that florbetapir F 18 binds with high affinity to &bgr;-amyloid (A&bgr;) plaques in human brain homogenates (Kd=3.7 nM) and has favorable imaging pharmacokinetic properties, including rapid brain penetration and washout. This study used human autopsy brain tissue to evaluate the correlation between in vitro florbetapir F 18 binding and A&bgr; density measured by established neuropathologic methods. MethodsThe localization and density of florbetapir F 18 binding in frozen and formalin-fixed paraffin-embedded sections of postmortem brain tissue from 40 patients with a varying degree of neurodegenerative pathology was assessed by standard florbetapir F 18 autoradiography and correlated with the localization and density of A&bgr; identified by silver staining, thioflavin S staining, and immunohistochemistry. ResultsThere were strong quantitative correlations between florbetapir F 18 tissue binding and both A&bgr; plaques identified by light microscopy (Silver staining and thioflavin S fluorescence) and by immunohistochemical measurements of A&bgr; using 3 antibodies recognizing different epitopes of the A&bgr; peptide. Florbetapir F 18 did not bind to neurofibrillary tangles. ConclusionsFlorbetapir F 18 selectively binds A&bgr; in human brain tissue. The binding intensity was quantitatively correlated with the density of A&bgr; plaques identified by standard neuropathologic techniques and correlated with the density of A&bgr; measured by immunohistochemistry. As A&bgr; plaques are a defining neuropathologic feature for Alzheimer disease, these results support the use of florbetapir F 18 as an amyloid positron emission tomography ligand to identify the presence of Alzheimer disease pathology in patients with signs and symptoms of progressive late-life cognitive impairment.

[18F](2S,4S)-4-(3-Fluoropropyl)glutamine as a Tumor Imaging Agent

Although the growth and proliferation of most tumors is fueled by glucose, some tumors are more likely to metabolize glutamine. In particular, tumor cells with the upregulated c-Myc gene are generally reprogrammed to utilize glutamine. We have developed new 3-fluoropropyl analogs of glutamine, namely [18F](2S,4R)- and [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 3 and 4, to be used as probes for studying glutamine metabolism in these tumor cells. Optically pure isomers labeled with 18F and 19F (2S,4S) and (2S,4R)-4-(3-fluoropropyl)glutamine were synthesized via different routes and isolated in high radiochemical purity (≥95%). Cell uptake studies of both isomers showed that they were taken up efficiently by 9L tumor cells with a steady increase over a time frame of 120 min. At 120 min, their uptake was approximately two times higher than that of l-[3H]glutamine ([3H]Gln). These in vitro cell uptake studies suggested that the new probes are potential tumor imaging agents. Yet, the lower chemical yield of the precursor for 3, as well as the low radiochemical yield for 3, limits the availability of [18F](2S,4R)-4-(3-fluoropropyl)glutamine, 3. We, therefore, focused on [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4. The in vitro cell uptake studies suggested that the new probe, [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4, is most sensitive to the LAT transport system, followed by System N and ASC transporters. A dual-isotope experiment using l-[3H]glutamine and the new probe showed that the uptake of [3H]Gln into 9L cells was highly associated with macromolecules (>90%), whereas the [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4, was not (<10%). This suggests a different mechanism of retention. In vivo PET imaging studies demonstrated tumor-specific uptake in rats bearing 9L xenographs with an excellent tumor to muscle ratio (maximum of ∼8 at 40 min). [18F](2S,4S)-4-(3-fluoropropyl)glutamine, 4, may be useful for testing tumors that may metabolize glutamine related amino acids.

Multidentate 18F-Polypegylated Styrylpyridines As Imaging Agents for Aβ Plaques in Cerebral Amyloid Angiopathy (CAA)

β-Amyloid plaques (Aβ plaques) in the brain are associated with cerebral amyloid angiopathy (CAA). Imaging agents that could target the Aβ plaques in the living human brain would be potentially valuable as biomarkers in patients with CAA. A new series of (18)F styrylpyridine derivatives with high molecular weights for selectively targeting Aβ plaques in the blood vessels of the brain but excluded from the brain parenchyma is reported. The styrylpyridine derivatives, 8a-c, display high binding affinities and specificity to Aβ plaques (K(i) = 2.87, 3.24, and 7.71 nM, respectively). In vitro autoradiography of [(18)F]8a shows labeling of β-amyloid plaques associated with blood vessel walls in human brain sections of subjects with CAA and also in the tissue of AD brain sections. The results suggest that [(18)F]8a may be a useful PET imaging agent for selectively detecting Aβ plaques associated with cerebral vessels in the living human brain.

68Ga-Bivalent Polypegylated Styrylpyridine Conjugates for Imaging Aβ Plaques in Cerebral Amyloid Angiopathy

Aβ plaques deposited on blood vessels are associated with cerebral amyloid angiopathy (CAA). In an effort to selectively map these Aβ plaques, we are reporting a new series of (68)Ga labeled styrylpyridine derivatives with high molecular weights. In vitro binding to Aβ plaques in post-mortem Alzheimers disease (AD) brain tissue showed that these (68)Ga labeled bivalent styrylpyridines displayed good affinities and specificity (Ki < 30 nM). In vitro autoradiography using post-mortem AD brain sections showed specific binding of these (68)Ga complexes to Aβ plaques. Biodistribution studies in normal mice showed very low initial brain uptakes (<0.3% dose/g) indicating a low blood-brain barrier (BBB) penetration. The preliminary results suggest that (68)Ga labeled bivalent styrylpyridines may be promising candidates as PET imaging radiotracers for detecting CAA.

In vitro and in vivo properties of murine monoclonal antibody for a novel immature thymocyte-differentiated antigen, JL1

JL1 is a novel thymocyte-differentiated antigen strictly confined to stage II immature cortical thymocytes. It is expressed in several types of leukemias and lymphomas. Murine anti-JL1 monoclonal antibody labeled with 131I and 99mTc showed 60-70% of immunoreactivity and 1.4-1.9 x 10(9) L/mol of affinity constant. The incubation of the radiolabeled antibody with Molt-4 cells showed no evidence of modulation or shedding. Localization indices increased from day 3 to day 5 in SCID mice bearing Molt-4 cells.