Pharmacological characterization of mutant huntingtin aggregate-directed PET imaging tracer candidates

Abstract

Huntington’s disease (HD) is caused by a CAG trinucleotide repeat expansion in the first exon of the huntingtin (HTT) gene coding for the huntingtin (HTT) protein. The misfolding and consequential aggregation of CAG-expanded mutant HTT (mHTT) underpin HD pathology. Our interest in the life cycle of HTT led us to consider the development of high-affinity small-molecule binders of HTT oligomerized/amyloid-containing species that could serve as either cellular and in vivo imaging tools or potential therapeutic agents. We recently reported the development of PET tracers CHDI-180 and CHDI-626 as suitable for imaging mHTT aggregates, and here we present an in-depth pharmacological investigation of their binding characteristics. We have implemented an array of in vitro and ex vivo radiometric binding assays using recombinant HTT, brain homogenate-derived HTT aggregates, and brain sections from mouse HD models and humans post-mortem to investigate binding affinities and selectivity against other pathological proteins from indications such as Alzheimer’s disease and spinocerebellar ataxia 1. Radioligand binding assays and autoradiography studies using brain homogenates and tissue sections from HD mouse models showed that CHDI-180 and CHDI-626 specifically bind mHTT aggregates that accumulate with age and disease progression. Finally, we characterized CHDI-180 and CHDI-626 regarding their off-target selectivity and binding affinity to beta amyloid plaques in brain sections and homogenates from Alzheimer’s disease patients.

Longitudinal preclinical evaluation of the novel radioligand [11C]CHDI-626 for PET imaging of mutant huntingtin aggregates in Huntington’s disease

“…This observation could indicate that in vivo PET imaging provides higher sensitivity than in vitro autoradiography in detecting differences between genotypes at 3 M, a relevant aspect for translation to clinical settings. Indeed, the in vitro profile of CHDI-626 suggests its potential as an mHTT PET radioligand in humans [ 16 ], particularly due to its lack of affinity towards amyloid plaques and/or tau tangles, as previously observed in human tissue with other mHTT-directed radioligands [ 15 , 17 ]. However, the rapid clearance and kinetics observed represent a limitation for [ 11 C]CHDI-626 PET imaging in detecting more subtle changes in mHTT load in mice, exemplified by the in vivo vs in vitro dichotomy between 9 and 13 M in HET mice.…”

“…A notable property of the mHTT PET radioligands currently under investigation is affinity towards amyloid plaques in post-mortem human tissue that might reduce their specificity [ 15 ], and efforts to overcome this issue are currently focused on the development of radioligands with high affinity and selectivity towards mHTT aggregates suitable for clinical applications. We have recently reported the development of a novel high-affinity radioligand, CHDI-626, that targets mHTT and lacks significant affinity towards amyloid plaques, representing a potential candidate for mHTT PET imaging in the clinic [ 16 , 17 ].…”

A Positron Emission Tomography Ligand for Mutant Huntingtin Sheds Light on Disease

“…Although HTT levels measured in cerebrospinal fluid correlate well with brain expression, visualization of mutant HTT in patients, particularly those entering trials of HTT‐lowering therapeutics, would provide insight into the regional distribution of pathology, assess target engagement, and permit in vivo evaluation of regional durability of therapeutic effects. A significant step forward is the development of a positron emission tomography (PET) imaging radioligand with high affinity and selectivity for mutant HTT 1,2 . The radioligand described by Bertoglio and colleagues, 1 dubbed CHDI‐180, binds mutant, but not wild‐type, HTT with low nanomolar affinity and recognizes a nonmonomeric species distinct from intranuclear inclusion bodies.…”

“…As with other neurodegenenerative proteinopathies, the identity of the relevant toxic protein species is unclear 3 . Further studies are needed to define the precise HTT species recognized by CHDI‐180, to determine whether this is the optimal target for PET imaging and to establish whether reported binding of CHDI‐180 to ß‐amyloid 2 confounds results in patients. ß‐Amyloid binding may not be an issue in this largely younger to middle‐aged population.…”

“…A significant step forward is the development of a positron emission tomography (PET) imaging radioligand with high affinity and selectivity for mutant HTT. 1,2 The radioligand described by Bertoglio and colleagues, 1 dubbed CHDI-180, binds mutant, but not wild-type, HTT with low nanomolar affinity and recognizes a nonmonomeric species distinct from intranuclear inclusion bodies. CHDI-180 identified ageand region-specific pathology in three mouse models and detected pharmacological effects of two intervention paradigms: direct striatal delivery of adeno-associated virus (AAV)-expressing zinc-finger protein repressors of mutant HTT and genetic suppression of mutant HTT in a regulatable mouse model.…”