Epigenetic Regulation of Wnt Signaling by Carboxamide-Substituted Benzhydryl Amines that Function as Histone Demethylase Inhibitors

Abstract

Summary Aberrant activation of Wnt signaling triggered by mutations in either Adenomatous Polyposis Coli ( APC ) or CTNNB1 (β-catenin) is a hallmark of colorectal cancers (CRC). As part of a program to develop epigenetic regulators for cancer therapy, we developed carboxamide-substituted benzhydryl amines (CBAs) bearing either aryl or heteroaryl groups that selectively targeted histone lysine demethylases (KDMs) and functioned as inhibitors of the Wnt pathway. A biotinylated variant of N -((5-chloro-8-hydroxyquinolin-7-yl) (4-(diethylamino)phenyl)-methyl)butyramide (CBA-1) identified KDM3A as a binding partner. KDM3A is a Jumonji (JmjC) domain-containing demethylase that is significantly upregulated in CRC. KDM3A regulates the demethylation of histone H3's lysine 9 (H3K9Me 2 ), a repressive marker for transcription. Inhibiting KDM3 increased H3K9Me 2 levels, repressed Wnt target genes, and curtailed in vitro CRC cell proliferation. CBA-1 also exhibited in vivo inhibition of Wnt signaling in a zebrafish model without displaying in vivo toxicity.

Chemical Inhibitors Targeting the Histone Lysine Demethylase Families with Potential for Drug Discovery

“…Two members of the KDM3 group, KDM3A and KDM3C, represent the most promising therapeutic targets for colorectal cancers (CRC) [ 71 , 72 ] and a subtype of acute leukemia, respectively [ 73 ]. KDM3A has been implicated in CRC progression via Wnt signaling where KDM3A coactivates downstream Wnt target genes including c- Myc and cyclin D1 [ 71 , 72 ].…”

Protocol for rapid assessment of the efficacy of novel Wnt inhibitors using zebrafish models

“…A pigmented strain is ideal for easier visualization of eye formation. Note: This protocol utilizes the Wnt inhibitor N -((5-chloro-8-hydroxyquinolin-7-yl)(4-(diethylamino)phenyl)-methyl)butyramide (CBA-1) ( Zhang et al, 2020 ) but can be used to examine Wnt inhibition and in vivo toxicity of any compound. Doses given to animals may need to be adjusted based on potency and toxicity of compounds.…”

“…These experiments are completed within 1 week to rapidly narrow drug candidates before moving to more expensive pre-clinical testing. For complete details on the use and execution of this protocol, please refer to Zhang et al. (2020) .…”

“…For complete details on the use and execution of this protocol, please refer to Zhang et al. (2020) .…”

Genetic Mutations and Non-Coding RNA-Based Epigenetic Alterations Mediating the Warburg Effect in Colorectal Carcinogenesis

“…Epigenetics has been reported to either restore or inhibit the activity of APC. A biotinylated variant of N -((5-chloro-8-hydroxyquinoline-7-yl) (4-(diethylamino)phenyl)-methyl) butyramide (CBA-1) has been shown to epigenetically promote the upregulation of the Wnt pathway through its partner KDM3A [ 71 ]. The latter is a demethylase enzyme that promotes the demethylation of the transcription marker histone H3′s lysine 9 (H3K9Me2).…”