The Histone Deacetylase (HDAC) Inhibitor PCI-24781 Decreases Pro-Inflammatory Cytokine Secretion In Vitro and In Vivo and Protects Against Endotoxemia In a Sepsis Model

Abstract 3914

Inhibitors of histone deacetylases (HDACs) are currently in clinical testing for treating various cancers, and two have been recently approved by the US FDA for treating cutaneous T-cell lymphoma. Here we describe novel anti-inflammatory properties of the HDAC inhibitor PCI-24781 which is in clinical trials for multiple indications including lymphoma (Evens et al., Blood 114: 2726, ASH 2009 Annual Meeting Abstracts). Cytokines such as interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-a) have been shown to be involved in human inflammatory disorders, and an anti-IL-6 treatment was recently approved for rheumatoid arthritis (RA). Therefore, the effect of PCI-24781 on cytokine production by lipopolysaccharide (LPS)-stimulated human peripheral mononuclear blood cells (PBMC) as well as isolated monocytes was studied at the RNA expression level by microarrays and Taqman, and at the protein level by ELISA. PCI-24781 potently inhibits the production and secretion of several pro-inflammatory cytokines, including IL-6, TNF-a and interleukin-1beta (IL-1b), at both RNA and protein levels. In murine RAW macrophages as well, PCI-24781 inhibited LPS-stimulated IL-6 secretion at 20nM. PCI-24781 was most effective when given with or before LPS, but was still effective when given an hour after LPS. Similarly, PCI-24781 greatly attenuated in vivo pro-inflammatory cytokine production in LPS-treated Balb/c mice; the IC50 for IL-6 inhibition was < 5 mg/kg. Both the in vitro and in vivo IC50s for IL-6 inhibition are considerably less than the concentrations required to inhibit growth and induce apoptosis in tumor cells (0.2-0.5mM) and in xenograft models (60-80 mg/kg). The mechanism by which these cytokines are controlled involves attenuation of the LPS receptor TLR4 signaling at multiple levels, including acetylation of targets such as MKP-1 and NF-kB subunit p65 in the downstream MAPK and NF-kB pathways; other factors include reduced expression of proteasome, IKK and other NF-kB subunits. Interestingly, we observed a large reduction in levels of NOS2, which causes hypotension during sepsis by producing the inflammatory mediator nitric oxide (NO). Therefore the activity of PCI-24781 was tested in a model of sepsis where mice were treated with a lethal dose of 100 mg/kg LPS, an endotoxin known to be a major mediator of sepsis in humans. PCI-24781 was injected twice, first 16 h before LPS and then 2 h before LPS, in groups of 10 mice each. Control mice that did not receive any PCI-24781 all died within 2 days after LPS (mortality 100%). Pretreatment with PCI-24781 led to dose-dependent increase in survival with 60% of the mice surviving past 6 days with 2 doses of 50mg/kg PCI-24781. These data show that the HDAC inhibitor PCI-24781 protects mice from lethal endotoxemia. Thus, taken together, our data suggest that PCI-24781 has potent anti-inflammatory activities and may be useful to treat inflammatory disorders including RA and sepsis in humans.