Abstract 1135: Chemical Proteomics effort identifies PKN1 as a key player in the canonical NF-κB signaling pathway.
Authors: Tang Katherine , Lampa Michael , Le Tieu-Binh , Barrague Matthieu , Tomlinson Ronald , Scott Brittain , Majid Tahir , Dorsch Marion , Cheng Hong , Lengauer Christoph , Garcia-Echeverria Carlos , Adrian Francisco , Levit Mikhail , Zhang Balin , Yu Kin , Cornella-Taracido Ivan , Srinivasan Lakshmi 
Published: August 2014

Deregulation of NF-κB signaling pathway has been identified as one of the key drivers of many hematological malignancies. Activation of the NF-κB pathway in these cells can happen either through activating mutations within the pathway, loss of function mutations of inhibitory molecules or through interaction with the stroma that secrete specific cytokines. The strategies for blocking the NF-κB pathway activation have so far focused on NF-κB pathway kinase inhibitors or the more general proteasome inhibitors. However, the adverse events associated with available NF-κB pathway modulators in pre-clinical and clinical settings have limited their general therapeutic use. To discover new targets and modulators of this key survival pathway, we performed a cellular phenotypic screen that led to the identification of a novel class of small molecules that inhibit the activation of the canonical NF-κB pathway. The compounds showed potent inhibition of NF-κB activation by multiple receptors including the TNF, antigen and BAFF/ APRIL receptors. Structure activity relationship analysis for this novel class of molecules identified pairs of stereoisomers wherein the cis isoform showed greater than 100 fold activity than the trans isoform. Detailed interrogation of the mechanism of action of the active cis isomers using biochemical and cellular pathway analysis revealed that the compounds are potent inhibitors of RHO associated kinases, only weakly inhibit IKK1 and IKK2 and strongly inhibit the induction of phosphorylated IkBα upon stimulation. However, the differential activity of the stereoisomers was not seen for the inhibition of the RHO associated kinases, suggesting the inhibition of NF-κB pathway was due to modulation of other targets. This raises the possibility of identifying novel targets associated with this compound. Using in-lysate affinity chemical proteomics with the active and in-active enantiomer, we have identified PKN1 as a key efficacy target for the inhibitors. Knockdown of PKN1 using genetic tools was sufficient to abrogate the response of cells to TNFα stimulation implicating a critical role for PKN1 in the activation of NF-κB pathway. Our findings therefore present an opportunity to develop novel NF-κB inhibitors for the tumors that depend on this key pathway for survival.

Citation Format: Katherine Tang, Michael Lampa, Tieu-Binh Le, Matthieu Barrague, Ronald Tomlinson, Brittain Scott, Tahir Majid, Marion Dorsch, Hong Cheng, Christoph Lengauer, Carlos Garcia-Echeverria, Francisco Adrian, Mikhail Levit, Balin Zhang, Kin Yu, Ivan Cornella-Taracido, Lakshmi Srinivasan. Chemical Proteomics effort identifies PKN1 as a key player in the canonical NF-κB signaling pathway. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1135. doi:10.1158/1538-7445.AM2013-1135