Huntington's disease (HD) is caused by an expanded CAG trinucleotide repeat in exon 1 of the huntingtin (HTT) gene. We report the design of a series of HTT pre-mRNA splicing modulators that lower huntingtin (HTT) protein, including the toxic mutant huntingtin (mHTT), by promoting insertion of a pseudoexon containing a premature termination codon at the exon 49-50 junction. The resulting transcript undergoes nonsense-mediated decay, leading to a reduction of HTT mRNA transcripts and protein levels. The starting benzamide core was modified to pyrazine amide and further optimized to give a potent, CNS-penetrant, and orally bioavailable HTT-splicing modulator 27. This compound reduced canonical splicing of the HTT RNA exon 49-50 and demonstrated significant HTT-lowering in both human HD stem cells and mouse BACHD models. Compound 27 is a structurally diverse HTT-splicing modulator that may help understand the mechanism of adverse effects such as peripheral neuropathy associated with branaplam.

Huntington’s disease (HD) is a neurodegenerative genetic disorder that affects muscle coordination and leads to cognitive decline. HD is caused by a trinucleotide repeat expansion in the huntingtin (HTT) gene, specifically a CAG (polyglutamine) expansion in exon 1 which results in an abnormal mutant protein. HD therapeutic discovery is currently biased towards HTT lowering agents. Encouragingly, a clinical pipeline for DNA/RNA targeted HTT-lowering agents now exists; however, most of these approaches use biological agents, such as ASOs, RNAi and ZFTRs that require invasive administration and have relatively limited biodistribution. To overcome these challenges, we seek to identify novel brain penetrant small molecules with suitable oral dosing that selectively lower mutant HTT protein. To do this, we developed an unbiased phenotypic assay in HD-patient derived, polyQ48 embryonic stem cells (ESC) and screened AstraZeneca’s industry-leading 250k EPEC compound library through the Open Innovation Partnership Scheme. Progressed HTT-lowering hits showed good translation in IC50 format; counter-screen and orthogonal assays were then applied to define specificity, selectivity, and putative mechanism of action. One compound was identified for further progression as it displayed a robust and specific HTT-lowering profile. Around 30 close analogues of this hit were synthesized to reveal an active series that was shown to work via a unique HTT RNA-lowering mechanism. A review of this screening campaign and its output will be presented along with the characterisation studies aimed at understanding the mechanism of action of the hit series.

The Rho kinase (ROCK) pathway is implicated in the pathogenesis of several conditions, including neurological diseases. In Huntington's disease (HD), ROCK is implicated in mutant huntingtin (HTT) aggregation and neurotoxicity, and members of the ROCK pathway are increased in HD mouse models and patients. To validate this mode of action as a potential treatment for HD, we sought a potent, selective, central nervous system (CNS)-penetrant ROCK inhibitor. Identifying a compound that could be dosed orally in mice with selectivity against other AGC kinases, including protein kinase G (PKG), whose inhibition could potentially activate the ROCK pathway, was paramount for the program. We describe the optimization of published ligands to identify a novel series of ROCK inhibitors based on a piperazine core. Morphing of the early series developed in-house by scaffold hopping enabled the identification of a compound exhibiting high potency and desired selectivity and demonstrating a robust pharmacodynamic (PD) effect by the inhibition of ROCK-mediated substrate (MYPT1) phosphorylation after oral dosing.

Huntington's disease (HD) is a devastating, genetic neurodegenerative disease caused by a tri-nucleotide expansion in exon 1 of the huntingtin gene. HD is clinically characterized by chorea, emotional and psychiatric disturbances and cognitive deficits with later symptoms including rigidity and dementia. Pathologically, the cortico-striatal pathway is severely dysfunctional as reflected by striatal and cortical atrophy in late-stage disease. Brain-derived neurotrophic factor (BDNF) is a neuroprotective, secreted protein that binds with high affinity to the extracellular domain of the tropomyosin-receptor kinase B (TrkB) receptor promoting neuronal cell survival by activating the receptor and down-stream signaling proteins. Reduced cortical BDNF production and transport to the striatum have been implicated in HD pathogenesis; the ability to enhance TrkB signaling using a BDNF mimetic might be beneficial in disease progression, so we explored this as a therapeutic strategy for HD. Using recombinant and native assay formats, we report here the evaluation of TrkB antibodies and a panel of reported small molecule TrkB agonists, and identify the best candidate, from those tested, for in vivo proof of concept studies in transgenic HD models.