Specific Inhibition of Histone Deacetylase 8 Reduces Gene Expression and Production of Proinflammatory Cytokines in Vitro and in Vivo

ITF2357 (generic givinostat) is an orally active, hydroxamic-containing histone deacetylase (HDAC) inhibitor with broad anti-inflammatory properties, which has been used to treat children with systemic juvenile idiopathic arthritis. ITF2357 inhibits both Class I and II HDACs and reduces caspase-1 activity in human peripheral blood mononuclear cells and the secretion of IL-1 beta and other cytokines at 25-100 nM; at concentrations >200 nM, ITF2357 is toxic in vitro. ITF3056, an analog of ITF2357, inhibits only HDAC8 (IC50 of 285 nM). Here we compared the production of IL-1 beta, IL-1 alpha, TNF alpha, and IL-6 by ITF2357 with that of ITF3056 in peripheral blood mononuclear cells stimulated with lipopolysaccharide (LPS), heat-killed Candida albicans, or anti-CD3/anti-CD28 antibodies. ITF3056 reduced LPS-induced cytokines from 100 to 1000 nM; at 1000 nM, the secretion of IL-1 beta was reduced by 76%, secretion of TNF alpha was reduced by 88%, and secretion of IL-6 was reduced by 61%. The intracellular levels of IL-1 alpha were 30% lower. There was no evidence of cell toxicity at ITF3056 concentrations of 100-1000 nM. Gene expression of TNF alpha was markedly reduced (80%), whereas IL-6 gene expression was 40% lower. Although anti-CD3/28 and Candida stimulation of IL-1 beta and TNF alpha was modestly reduced, IFN gamma production was 75% lower. Mechanistically, ITF3056 reduced the secretion of processed IL-1 beta independent of inhibition of caspase-1 activity; however, synthesis of the IL-1 beta precursor was reduced by 40% without significant decrease in IL-1 beta mRNA levels. In mice, ITF3056 reduced LPS-induced serum TNF alpha by 85% and reduced IL-1 beta by 88%. These data suggest that specific inhibition of HDAC8 results in reduced inflammation without cell toxicity.