HDAC6 Brain Mapping with [¹⁸F]Bavarostat Enabled by a Ru-Mediated Deoxyfluorination

Abstract

Histone deacetylase 6 (HDAC6) function and dysregulation have been implicated in the etiology of certain cancers and more recently in central nervous system (CNS) disorders including Rett syndrome, Alzheimer’s and Parkinson’s diseases, and major depressive disorder. HDAC6-selective inhibitors have therapeutic potential, but in the CNS drug space the development of highly brain penetrant HDAC inhibitors has been a persistent challenge. Moreover, no tool exists to directly characterize HDAC6 and its related biology in the living human brain. Here, we report a highly brain penetrant HDAC6 inhibitor, Bavarostat, that exhibits excellent HDAC6 selectivity (>80-fold over all other Zn-containing HDAC paralogues), modulates tubulin acetylation selectively over histone acetylation, and has excellent brain penetrance. We further demonstrate that Bavarostat can be radiolabeled with 18F by deoxyfluorination through in situ formation of a ruthenium π-complex of the corresponding phenol precursor: the only method currently suitable for synthesis of [18F]Bavarostat. Finally, by using [18F]Bavarostat in a series of rodent and nonhuman primate imaging experiments, we demonstrate its utility for mapping HDAC6 in the living brain, which sets the stage for first-in-human neurochemical imaging of this important target.

Radiosynthesis and preliminary evaluation of an18F‐labeled tubastatin A analog for PET imaging of histone deacetylase 6

“…Regarding the development of HDAC6-selective radioligands, Lu et al reported on 11 C-carbonylation methods for labeling tubastatin A, an HDAC6-selective inhibitor (1 in Figure 1), 36 that employed [ 11 C]carbon monoxide. 37 Radiosynthesis of the tubastatin A analog [ 11 C]KB631 ([ 11 C]2 in Figure 1) with [ 11 C]methyl iodide was also reported by Lu et al, 38 and its preclinical evaluation for visualization of B16F10 melanoma has recently been reported by Vermeulen et al 39 Additionally, [ 18 F] Bavarostat was developed by Strebl et al 40 as an adamantane-conjugated HDAC6-selective radioligand. Although [ 18 F]Bavarostat needs a unique rutheniummediated 18 F-deoxyfluorination 41 for its radiosynthesis, its BBB permeability and specific binding in the brain was confirmed by in vivo blocking studies in nonhuman primates.…”

“…36,42 The chemical structure of 1 consists of three motifs: a hydroxamic acid zinc-binding group that localizes a zinc ion within the HDAC active site, a carboline-based cap group that interacts with the protein surface of the binding pocket, and a benzyl linker group that bridges the F I G U R E 1 Chemical structures of tubastatin A and its radiolabeled analogs two groups. We intended to perform this radiosynthesis without intermediary solid-phase extraction or high-performance liquid chromatography (HPLC) purification steps, which have been incorporated in other hydroxamic acid-based radioligands, 32,40,43 because these processes often complicate automated radiosynthesis. Kalin et al demonstrated that a substituent could be introduced at the nitrogen atom of tetrahydropyridine of the β-carboline regioisomer of 1 without seriously disrupting HDAC6 selectivity.…”

“…Radiosynthesis of [ 18 F]3 was achieved using a two-step reaction composed of 18 F-fluorination by general nucleophilic substitution and conversion of a methylester group into a hydroxamic acid group. We intended to perform this radiosynthesis without intermediary solid-phase extraction or high-performance liquid chromatography (HPLC) purification steps, which have been incorporated in other hydroxamic acid-based radioligands, 32,40,43 because these processes often complicate automated radiosynthesis. In addition, preclinical evaluations such as binding selectivity assays and biodistribution assays were performed to assess the potential utility of [ 18 F]3 for visualization of HDAC6.…”

[18F]Atorvastatin: synthesis of a potential molecular imaging tool for the assessment of statin-related mechanisms of action

“…For the radiofluorination of atorvastatin, the benzyl ether pyrrole intermediate 6 was synthesized and subsequently converted to the corresponding phenol 7 by palladium on carbon (Pd/C)-catalyzed hydrogenolysis (Scheme 2). Coordination of 7 to ruthenium (II) decreases its π-electron density, providing 8, and activates the precursor towards radiodeoxyfluorination [58][59][60].…”

“…However, in the case of atorvastatin, the 18 F-fluorination of electron-rich arenes is a classical struggle in radiochemistry and has challenges associated with synthesis automation [57]. The fluorine-containing aromatic ring within the atorvastatin structure is electron-rich, and therefore, we use the ruthenium-mediated radiodeoxyfluorination strategy [58][59][60] to synthesize [ 18 F]atorvastatin ([ 18 F]12). This radiotracer may have the potential to become an attractive tool for statin-related research, enabling the understanding of cellular and subcellular mechanisms, the identification of off-target activity and, ultimately, allowing to select between statin-resistant and non-resistant patients for targeted therapy.…”