Visual Abstract
Abstract
Heat
shock protein 90 (Hsp90) is essential for maintaining cellular
proteostasis and may play an important role in the development of
neurodegenerative proteinopathies. Therefore, we aimed to develop an
Hsp90-specific PET brain tracer to quantify Hsp90 expression in the
brain in vivo in order to explore its potential as a biomarker for
neurodegenerative disease characterization and to support Hsp90-targeted
drug development. Methods: We developed the radiosynthesis of (R)-2-amino-7-(4-fluoro-2-(6-(methoxy-11C)pyridin-2-yl)phenyl)-4-methyl-7,8-dihydropyrido[4,3-d]pyrimidin-5(6H)-one, [11C]HSP990,
and validated the tracer using in vitro autoradiography, in vitro brain
homogenate saturation binding, ex vivo biodistribution, and in vivo PET
imaging in rodent models of Alzheimer disease (AD) and Parkinson
disease versus healthy age-matched and young controls. Human brain
samples from AD patients and healthy subjects were included in our
in vitro binding studies. A nonhuman primate PET brain study with
arterial blood sampling was conducted under baseline and blocking
conditions. Results: In vitro and in vivo [11C]HSP990
studies in rodents and a nonhuman primate revealed saturable Hsp90
binding pools in natural killer lymphocytes, bone marrow, and notably
the brain, where the highest binding was observed, particularly in gray
matter. Blocking studies indicated that saturable Hsp90 in natural
killer lymphocytes considerably influences the pharmacokinetics of
Hsp90-targeting probes, which is critical for Hsp90 drug development. In
vitro [3H]HSP990 brain homogenate saturation binding assays
suggested that the tracer binds a distinct subfraction of the total
Hsp90 pool, which is significantly diminished in both rodent and human
AD brain tissue compared with age-matched controls. In vivo PET imaging
confirmed reduced [11C]HSP990 brain binding on aging and an
even stronger decrease in AD mice, suggesting that Hsp90 depletion may
impair protein quality control and accelerate proteinopathies. Conclusion: [11C]HSP990
is a promising Hsp90-specific tracer and reveals strong Hsp90 binding
in the brain. Uniformly reduced tracer binding was observed in AD brain
tissue compared with age-matched controls. [11C]HSP990 holds
potential as a biomarker for neurodegenerative disease characterization
and progression, and it may aid in patient stratification and therapy
monitoring. Human [11C]HSP990 PET neuroimaging studies are under way to investigate whether these findings translate to humans.
Heat
shock protein 90 (Hsp90), present in 4 isoforms, is a key chaperone in
the protein quality control system, maintaining cellular proteostasis by
stabilizing, folding and refolding, and regulating many client proteins
(1).
Accordingly, Hsp90 function is implicated in diseases associated with
proteotoxic stress, such as neurodegenerative disorders and cancer (2,3).
The role of Hsp90 in neurodegenerative diseases, including Alzheimer
disease (AD), Parkinson disease (PD), amyotrophic lateral sclerosis, and
Huntington disease, remains controversial in the literature and has
been linked to both protective functions and pathologic roles.
Proteins involved in aggregation, such as β-amyloid, phosphorylated tau, and α-synuclein (αSyn), are reported Hsp90 clients (4–6).
Some studies have shown that upregulated Hsp90 colocalized with these
aggregates in a neurodegenerative brain, potentially contributing to
disease pathology by exacerbating aggregate accumulation and hindering
misfolded protein degradation (2,6).
This has sparked interest in Hsp90 inhibitors, to suppress its aberrant
neuronal activity, as potential treatments for neurodegenerative
disorders (7).
Conversely, reduced Hsp90 levels have been linked to neuronal cell
death. In this context, induction of the heat shock response through
Hsp90 inhibition has been explored to upregulate chaperone function,
thereby reducing protein aggregation and supporting cytoprotection (1,7–11). For instance, (S)-2-fluoro-6-((tetrahydrofuran-3-yl)amino)-4-(3,6,6-trimethyl-4-oxo-4,5,6,7-tetrahydro-1H-indol-1-yl)benzamide) (SNX-0723) and 9-(3-(tert-butylamino)propyl)-8-((6-iodobenzo[d][1,3]dioxol-5-yl)thio)-9H-purin-6-amine (PU-AD) (Fig. 1)
have shown promise in preclinical studies by preventing αSyn
oligomerization and rescuing striatal dopamine levels in a PD rat model
and by inducing the degradation of misfolded proteins and restoring
memory in an AD mouse model, respectively (2,12).
PU-AD was evaluated in clinical trials for AD, amyotrophic lateral
sclerosis, and glioblastoma, but these studies were withdrawn or
terminated and results have yet to be published (13–15). The compound (R)-2-amino-7-(4-fluoro-2-(6-methoxypyridin-2-yl)phenyl)-4-methyl-7,8-dihydropyrido[4,3-d]pyrimidin-5(6H)-one (HSP990) (Fig. 1)
has also shown therapeutic potential, improving cognitive function in
AD mouse models and reducing huntingtin aggregation in Huntington
disease, but exhibited neurotoxicity in a phase I trial for solid tumors
(16–18).
Chemical structure of Hsp90 inhibitors and PET brain tracers.
The
potential clinical efficacy of Hsp90 inhibitors depends on their
ability to exert therapeutic effects at safe doses. Accurate measurement
of Hsp90 occupancy in the brain using PET can significantly contribute
to determining optimal dosing regimens and avoiding toxic effects (19). Furthermore, Hsp90 PET brain imaging may provide insights into the role of Hsp90 in neurologic diseases.
Only a few Hsp90-targeting PET probes that can permeate the blood–brain barrier have been developed so far (2,20,21). [124I]PU-AD (Fig. 1)
showed higher hippocampal retention in human AD brain than in controls
on static delayed (3 h after injection) PET. However, absolute brain [124I]PU-AD concentrations were very low, and 124I produces low-quality PET images and exposes patients to high radiation because of its long half-life (2,22). More recently, 6-chloro-9-((4-(methoxy-11C)-3,5-dimethyl-2-pyridinyl)methyl]-9H-purin-2-amine ([11C]BIIB021) (Fig. 1)
was developed, exhibiting Hsp90-specific binding in the rat brain but
showing the presence of brain radiometabolites, which may complicate PET
image quantification (21). Our group previously developed (R)-2-amino-7-(2-(6-(methoxy-11C)pyridin-2-yl)phenyl)-4-methyl-7,8-dihydropyrido[4,3-d]pyrimidin-5(6H)-one ([11C]YC-72-AB85) (Fig. 1), demonstrating reversible Hsp90-specific brain binding in healthy rodents and a nonhuman primate (NHP) (20),
but it has not been explored in neurodegeneration. Given that HSP990
has already been used in a clinical trial, this study sought to develop
and evaluate [11C]HSP990 (Fig. 1)
as a Hsp90 PET tracer to elucidate saturable Hsp90 binding and its role
in health and neurodegenerative disease and to advance clinical
translation.
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