Inhibition of the histone demethylase KDM4B leads to activation of KDM1A, attenuates bacterial-induced pro-inflammatory cytokine release, and reduces osteoclastogenesis

Abstract

Periodontal disease (PD) afflicts 46% of Americans with no effective adjunctive therapies available. While most pharmacotherapy for PD targets bacteria, the host immune response is responsible for driving tissue damage and bone loss in severe disease. Herein, we establish that the histone demethylase KDM4B is a potential drug target for the treatment of PD. Immunohistochemical staining of diseased periodontal epithelium revealed an increased abundance of KDM4B that correlates with inflammation. In murine calvarial sections exposed to Aggregatibacter actinomycetemcomitans lipopolysaccharide (Aa-LPS), immunohistochemical staining revealed a significant increase in KDM4B protein expression. The 8-hydroxyquinoline ML324 is known to inhibit the related demethylase KDM4E in vitro, but has not been evaluated against any other targets. Our studies indicate that ML324 also inhibits KDM4B (IC50: 4.9 μM), and decreases the pro-inflammatory cytokine response to an Aa-LPS challenge in vitro. Our results suggest that KDM4B inhibition-induced immunosuppression works indirectly, requiring new protein synthesis. In addition, fluorescence-stained macrophages exhibited a significant decrease in global monomethyl histone 3 lysine 4 (H3K4me) levels following an Aa-LPS challenge that was prevented by KDM4B inhibition, suggesting this effect is produced through KDM1A-mediated demethylation of H3K4. Finally, ML324 inhibition of KDM4B in osteoclast progenitors produced a significant reduction in Aa-LPS-induced osteoclastogenesis. These data link histone methylation with host immune response to bacterial pathogens in PD, and suggest a previously unreported, alternative mechanism for epigenetic control of the host inflammatory environment. As such, KDM4B represents a new therapeutic target for treating hyper-inflammatory diseases that result in bone destruction.

Epigenetic Regulators Involved in Osteoclast Differentiation

“…The lysine demethylase (KDM) 4 family of protein enzymes differ from FAD+ dependent protein enzymes in that they contain a JmjC-domain that requires Fe + , O 2 and 2-oxogluterate as cofactors to properly function [ 28 ]. This family of proteins have the ability to demethylate di- and tri- methyl histone 3 lysine 9 and 36 (H3K9 and H3K36, respectively) as well as histone 1.4 lysine 26 (H1.4K26) and trimethyl histone 3 lysine 56 (H3K56) [ 28 ]. It has been demonstrated that large quantities of KDM4B protein in areas of inflammatory infiltrate increases the number of osteoclasts present [ 28 ].…”

“…This family of proteins have the ability to demethylate di- and tri- methyl histone 3 lysine 9 and 36 (H3K9 and H3K36, respectively) as well as histone 1.4 lysine 26 (H1.4K26) and trimethyl histone 3 lysine 56 (H3K56) [ 28 ]. It has been demonstrated that large quantities of KDM4B protein in areas of inflammatory infiltrate increases the number of osteoclasts present [ 28 ]. Additionally, in osteoclasts, it has been shown that the inhibition of KDM4B in pre-osteoclasts results in reduced osteoclastogenesis [ 28 ].…”

“…It has been demonstrated that large quantities of KDM4B protein in areas of inflammatory infiltrate increases the number of osteoclasts present [ 28 ]. Additionally, in osteoclasts, it has been shown that the inhibition of KDM4B in pre-osteoclasts results in reduced osteoclastogenesis [ 28 ]. Furthermore, with the inhibition of KDM4B, gene transcription of pro-inflammatory cytokines is also reduced [ 28 ].…”

“…Additionally, in osteoclasts, it has been shown that the inhibition of KDM4B in pre-osteoclasts results in reduced osteoclastogenesis [ 28 ]. Furthermore, with the inhibition of KDM4B, gene transcription of pro-inflammatory cytokines is also reduced [ 28 ]. Therefore, this data demonstrates that KDM4B is a transcriptional activator of osteoclast differentiation [ 28 ].…”

Inhibition of microRNA‐27b‐3p relieves osteoarthritis pain via regulation of KDM4B‐dependent DLX5

“…13,14 High expression of KDM4B has been reported to be correlated with inflammation and the inhibition of KDM4B, highlighting its potential as a novel therapeutic target for hyper-inflammatory diseases that gave rise to bone destruction. 15 A previous report indicated that KDM4B plays a vital role in lipolysis in adipose tissues. 16 DLX5 is an osteogenic gene and a negative regulator of adipogenesis.…”