Evaluation of Tau Radiotracers in Chronic Traumatic Encephalopathy

Introduction: Chronic traumatic encephalopathy (CTE) is a neurological disorder associated with head injuries. We recently revealed the variability in neuroinflammatory pathology following brain injuries (Brain Commun. in press). Diagnosis of CTE occurs upon autopsy and is based on the perivascular accumulation of hyperphosphorylated tau protein. Tau-PET radiopharmaceuticals that target neurofibrillary tangles (NFTs) are under evaluation in populations who have experienced brain injuries. Identifying appropriate radiotracers to successfully image CTE-tau in vivo could enable ante-mortem diagnosis of CTE for the first time and provides opportunities for therapeutic interventions following brain injuries. The goal of the present study is to conduct a head-to-head evaluation of five leading tritium-labeled tau-PET radiotracers in 12 cases of pathologically diagnosed subjects with CTE. In vitro radioligand binding assays were carried out to assess the utility of tau-PET radiotracers to image CTE-specific tau inclusions.

Methods: Thin-section autoradiography was employed to assess specific binding and distribution of [³H]flortaucipir (a.k.a. Tauvid^TM, AV-1451, T807), [³H]MK-6240 (a.k.a. florquinitau), [³H]PI-2620, [³H]APN-1607 (a.k.a. PM-PBB3) and [³H]CBD-2115 in fresh-frozen human post-mortem CTE frontal cortex. Radiotracer binding was quantified by digital autoradiography and non-specific binding was defined by homologous or heterologous blockade. Immunohistochemistry validation was performed for phospho-tau with AT8, 3R tau with RD3, 4R tau with RD4 and amyloid-β with 6F/3D antibodies. Tau target density (B_max) in postmortem tissue sections was quantified by saturation analysis.

Results: [³H]Flortaucipir demonstrated positive signal in all CTE cases examined, with a varying degree of specific binding (68.7 ± 10.5%; n=12) defined by homologous blockade and to a lesser extent by heterologous blockade with cold MK-6240 (27.3 ± 13.6%; n=12). The [³H]flortaucipir signal was also displaced by the monoamine oxidase (MAO)-A inhibitor clorgyline (43.9 ± 4.6%; n = 3), however off-target binding to MAO-B was not observed as defined by blockade with lazabemide. [³H]APN-1607 was moderately displaced in self-blocking studies and had high variability between CTE samples. [³H]APN-1607 was not displaced by cold flortaucipir, however, substantial displacement was observed when blocking with the β-amyloid targeting compound NAV-4694, indicating off-target binding of this radiotracer and lack of selectivity for tau. Target validation was performed by immunostaining for amyloid-β, and positive staining was observed in the cases where NAV-4694 displaced the [³H]APN-1607 signal. [³H]MK-6240 and [³H]PI-2620 displayed negligible binding in 10/12 CTE cases. In the singular CTE case with the most robust [³H]MK-6240 and [³H]PI-2620 signal, immunostaining revealed this individual was amyloid positive, and may be attributed to mixed AD/CTE pathology. [³H]CBD-2115 showed moderate binding that was self-blocked and aligned with 4R-tau immunostaining.

Conclusions: This head-to-head in vitro evaluation of tau-PET radiotracers to assess their ability for imaging CTE reveals the caveats of off-target binding to MAO-A and β-amyloid with [³H]flortaucipir and [³H]APN-1607, respectively. Potential use for [³H]MK-6240 and [³H]PI-2620 in mixed pathology or severe CTE cases was shown. [³H]CBD-2115 will continue to serve as an in vitro screening tool for our next-generation radiotracers to image the mixed 3R/4R tau pathology of CTE.

Acknowledgments. C.V. thanks the Canadian Institutes of Health Research (CIHR) for the receipt of the Canada Graduate Scholarship (Doctoral). We thank the VA-BU-CLF brain bank (funded by the NIH: P30AG072978, U54NS115266, R01AG062348 and RF1AG057902) for providing the CTE tissues.