Characterization of a selective Protein Kinase C theta (PKC[theta]) inhibitor for blocking T Cell responses

Abstract PKC theta (PKCθ) regulates T cell activation in lung inflammation and airway hyperresponsiveness, and in mouse models of multiple sclerosis, arthritis, and inflammatory bowel disease. We investigated inhibition of PKCθ enzyme activity by a small molecule as a strategy for modulating T cell mediated responses. Screening and lead optimization identified Compound A, a potent PKCθ inhibitor in both enzymatic and cell based assays, with low nM inhibitory activities. Compound A was highly selective (>1000 fold) over 50 kinases including representative serine/threonine, tyrosine, and phosphoinositol kinases, and the conventional and atypical PKCs, PKCβ and PKCξ. Compound A was 10 fold selective against PKCε, and 60 to 100 fold selectivity against PKCδand PKCη, respectively. We found that compound A strongly inhibited IL-2 production in antiCD3/anti-CD28 activated whole blood, using both human and mouse blood, with submicromolar potency. Based on these findings, we evaluated compound A- mediated inhibition of anti-CD3 induced IL-2 production in vivo. The results show a dose dependent inhibitory effect of compound A in this 4 hour cytokine production model. Additional cytokines inhibited by compound A in this model are IFNγ, IL-4, and TNF. IL-5 production was not reduced. We propose that compound A is a selective PKCθ inhibitor for further evaluating blockade of T cell responses in inflammation.