BF 227

The Binding of BF-227-Like Benzoxazoles to Human α-Synuclein and Amyloid β Peptide Fibrils

Development of an α-synuclein (α-Syn) positron emission tomography agent for the diagnosis and evaluation of Parkinson disease therapy is a key goal of neurodegenerative disease research. BF-227 has been described as an α-Syn binder and hence was employed as a lead to generate a library of α-Syn-binding compounds. [3H]BF-227 bound to α-Syn and amyloid β peptide (Aβ) fibrils with affinities (KD) of 46.0 nM and 15.7 nM, respectively. Affinities of BF-227-like compounds (expressed as Ki) for α-Syn and Aβ fibrils were determined, along with 5 reference compounds (flutafuranol, flutemetamol, florbetapir, BF-227, and PiB). Selectivity for α-Syn binding, defined as the Ki(Aβ)/Ki(α-Syn) ratio, was 0.23 for BF-227. A similar or lower ratio was measured for analogues decorated with alkyl or oxyethylene chains attached to the oxygen at the 6 position of BF-227, suggesting a lack of involvement of the side chain in fibril binding. BF-227-like iodobenzoxazoles had lower affinities and poor α-Syn selectivity. However, BF-227-like fluorobenzoxazoles had improved α-Syn selectively having Ki(Aβ)/Ki(α-Syn) ranging from 2.2 to 5.1 with appreciable fibril affinity, although not sufficient to warrant further investigation. Compounds based on fluorobenzoxazoles might offer an approach to obtaining an α-Syn imaging agent with an appropriate affinity and selectivity.

[11C]2-(2-[2-Dimethylaminothiazol-5-yl]ethenyl)-6-(2-[fluoro]ethoxy)benzoxazole

Alzheimer’s disease (AD) is a major neurodegenerative disease associated with an irreversible decline of mental functions and with cognitive impairment (1). It is characterized pathologically by neuronal loss with the presence in the brain of senile plaques of β-amyloid (Aβ) peptides and intracellular neurofibrillary tangles of filaments that contain the hyperphosphorylated protein tau (2, 3). Accelerated deposition of Aβ deposits seems to be a key risk factor associated with AD.

Early diagnosis of AD is important for treatment consideration and disease management (4). Several radioligands for positron emission tomography (PET) have been developed (5-7) and tested in humans as in vivo diagnostic tools for imaging and measuring the formation of Aβ deposits (7). The first agent successfully used in human studies was [¹⁸F]FDDNP (8), a malonitrile derivative found to bind to both neurofibrillary tangles and Aβ plaques. The second successful attempt was made with [¹¹C]PIB (9), also known as Pittsburgh Compound B or [¹¹C]6-OH-BTA-1, which showed marked retention in areas of the cortex known to contain substantial amounts of Aβ deposits. The third PET radioligand successfully tested in humans was [¹¹C]4-N-methylamino-4?-hydroxystilbene ([¹¹C]SB-13), a stilbene derivative that exhibits good binding affinities for Aβ aggregates in vitro, moderate lipophilicity, high initial brain uptake in the normal rat cortex, and a rapid washout (10).

Benzoxazole derivatives have shown very good binding affinities for Aβ aggregates and Aβ plaques, as well as good brain penetration (11). An optimized derivative, [¹¹C]2-(2-[2-dimethylaminothiazol-5-yl]ethenyl)-6-(2-[fluoro]ethoxy)benzoxazole ([¹¹C]BF-227), has been synthesized and is currently being studied as a PET radioligand for Aβ plaques (12).

In vivo detection of prion amyloid plaques using [(11)C]BF-227 PET

Purpose: In vivo detection of pathological prion protein (PrP) in the brain is potentially useful for the diagnosis of transmissible spongiform encephalopathies (TSEs). However, there are no non-invasive ante-mortem means for detection of pathological PrP deposition in the brain. The purpose of this study is to evaluate the amyloid imaging tracer BF-227 with positron emission tomography (PET) for the non-invasive detection of PrP amyloid in the brain.
Methods: The binding ability of BF-227 to PrP amyloid was investigated using autoradiography and fluorescence microscopy. Five patients with TSEs, including three patients with Gerstmann-Str?ussler-Scheinker disease (GSS) and two patients with sporadic Creutzfeldt-Jakob disease (CJD), underwent [(11)C]BF-227 PET scans. Results were compared with data from 10 normal controls and 17 patients with Alzheimer's disease (AD). The regional to pons standardized uptake value ratio was calculated as an index of BF-227 retention.
Results: Binding of BF-227 to PrP plaques was confirmed using brain samples from autopsy-confirmed GSS cases. In clinical PET study, significantly higher retention of BF-227 was detected in the cerebellum, thalamus and lateral temporal cortex of GSS patients compared to that in the corresponding tissues of normal controls. GSS patients also showed higher retention of BF-227 in the cerebellum, thalamus and medial temporal cortex compared to AD patients. In contrast, the two CJD patients showed no obvious retention of BF-227 in the brain.
Conclusion: Although [(11)C]BF-227 is a non-specific imaging marker of cerebral amyloidosis, it is useful for in vivo detection of PrP plaques in the human brain in GSS, based on the regional distribution of the tracer. PET amyloid imaging might provide a means for both early diagnosis and non-invasive disease monitoring of certain forms of TSEs.

Amyloid-Beta Radiotracer [18F]BF-227 Does Not Bind to Cytoplasmic Glial Inclusions of Postmortem Multiple System Atrophy Brain Tissue

The accumulation of aggregated alpha-synuclein (α-syn) in multiple brain regions is a neuropathological hallmark of synucleinopathies. Multiple system atrophy (MSA) is a synucleinopathy characterized by the predominant cerebral accumulation of aggregated α-syn as cytoplasmic glial inclusions (CGI). A premortem diagnosis tool would improve early diagnosis and help monitoring disease progression and therapeutic efficacy. One Positron Emission Tomography (PET) study suggested [¹¹C]BF-227 as a promising radiotracer for monitoring intracellular α-syn deposition in MSA patients. We sought to confirm the binding of this radiotracer to α-syn using state-of-the-art autoradiography. Medulla sections were obtained from 9 MSA patients and 9 controls (London Neurodegenerative Diseases Brain Bank). [¹⁸F]BF-227, chemically identical to [¹¹C]BF-227, was used at nanomolar concentrations to perform in vitro autoradiography assays. Autoradiograms were superimposed on fluorescent staining from the conformational anti-α-syn antibody 5G4 and quantified after immunofluorescence-driven definition of regions of interest. Autoradiography showed no specific signals in MSA patients in comparison to controls despite widespread pathology detected by immunofluorescence. Autoradiography does not support a significant binding of [¹⁸F]BF-227 to CGI at concentrations typically achieved in PET experiments.

Amyloid PET in mild cognitive impairment and Alzheimer's disease with BF-227: comparison to FDG-PET

We recently developed a novel PET tracer, (11)C-labeled 2-(2-[2-dimethylaminothiazol-5-yl]ethenyl)-6-(2-[fluoro]ethoxy)benzoxazole ([(11)C]BF-227), and had success with in vivo detection of amyloid plaques in Alzheimer's disease (AD) brains (Kudo et al. in J Nucl Med 8:553-561, 2007). We applied this tracer to subjects with mild cognitive impairment (MCI) and AD in order to elucidate the status of amyloid plaque deposition in MCI and compared the diagnostic performance of BF-227-PET with that of FDG-PET in AD cases. We studied 12 aged normal (AN) subjects, 15 MCIs and 15 ADs with PET using [(11)C]BF-227. PET images were obtained after administration of BF-227 and the regional standardized uptake value (SUV) and the ratio of regional to cerebellar SUV were calculated as an index of BF-227 binding. AD patients showed increased uptake of [(11)C]BF-227 in the neocortical areas and striatum as well as decreased glucose metabolism in temporoparietal, posterior cingulate and medial temporal areas. MCI subjects showed a significant increase in BF-227 uptake in the neocortical areas similar to AD, and the most significant difference of BF-227 retention was observed in the parietal lobe if its retentions for MCI were compared to those for AD and AN. On the other hand, glucose hypometabolism in MCI was confined to cingulate and medial temporal cortices. Neocortical BF-227 uptake negatively correlated with glucose metabolism. Receiver operating characteristic (ROC) analysis indicated higher specificity and sensitivity with BF-227-PET than those with FDG-PET for differential diagnosis between AD and normal control. We conclude that [(11)C]BF-227-PET has a possibility to be a useful technology for early detection of AD pathology and also even in the MCI stage.