IN VITRO BINDING STUDIES TO EVALUATE THE NFT-SPECIFICITY OF [3H]MK-6240 AND [3H]AV-1451 BINDING IN SUBCORTICAL REGIONS OF HUMAN AD BRAIN

Abstract

[18F]MK-6240 is a promising PET tracer for the in vivo quantification of NFTs in Alzheimer's Disease (AD) patients1, 2. In clinical studies, [18F]MK-6240 shows high levels of retention in brain regions expected to contain NFT pathology in AD patients, including the amygdala region, but not in healthy control. To increase confidence that the in vivo retention of [18F]MK-6240 in these brain regions of AD patients is due to NFT binding, in vitro studies were carried out using human brain tissues. Similar studies were done with [3H]AV-1451 (T-807) to compare two tracer binding patterns in the same brain sample. Frozen, age-matched AD and non-AD brains were used in the study. [3H]MK-6240 and [3H]AV-1451 were synthesized in house. Autoradiography (ARG) was performed using frozen brain slices. Brain homogenates were used for filtration binding assays. Immunohistochemistry (IHC) was performed with brain slices adjacent to those of ARG study using antibodies made against amyloid plaque (6E10) and NFTs (AT8). In AD amygdala slices, both [3H]MK-6240 and [3H]AV-1451 bind to brain regions with AD tauopathy as shown by co-localization of tau tracer binding with positive AT8 IHC stain (NFTs & neurites) from adjacent slices. However, in non-AD brain samples with negative AT8 IHC stain, [3H]MK-6240 shows minimal or no displaceable binding, while [3H]AV-1451 exhibits displaceable binding of moderate density in these brain samples, including the amygdala, pineal gland, raphe nucleus, substantia nigra, choroid plexus, cerebellum, and dura mater. [3H]MK-6240 does not block MAO-B tracer binding in non-AD brain homogenates, but [3H]AV-1451 does (Ki = 60 nM), in addition to its binding to MAO-A as previously reported. These results validate the expectation that MK-6240 binding in the amygdala is due to binding NFT pathology, and confirms the specificity of [3H]MK-6240 binding to NFTs in human AD brains. In contrast, [3H]-AV-1451 shows dis-placeable binding in regions that do not correlate with AT8 IHC staining, consistent with previous reports1,3–5.