Novel first-in-class selective IKKα inhibitors abrogate cancer cell NFκB signalling, survival and proliferation.

Introduction- Up-regulation of the key transcription factor pathway, nuclear factor kappa B (NFκB), has been strongly implicated in cancer(Gamble et al., 2012). Two main pathways operate to regulate the activation of different NFκB species; the canonical or classical NFκB (p65) pathway regulated exclusively by inhibitory kappa B kinase beta (IKKβ) and the non-canonical NFκB pathway driven by IKKα. To date the majority of studies have focussed on IKKβ inhibition in tackling cancer progression, however evidence showed inhibition of IKKβ dependent canonical NFκB pathway also leads to inhibition of apoptosis in human prostate cancer cells(Parrondo et al., 2010). We have therefore examined the potential for selective inhibition of IKKα as a novel therapeutic target. To this end we have developed the first selective IKKα inhibitor (SU1261) and have tested it on various cancer cells.

Methods- U2OS, PC3m and LNCAP AI cells(passage 3-35) were used for the study. Western blotting was used to assess pathway activation (non-canonical; p-p100, p52 formation, canonical; IκBα degradation, p-p65), cell cycle progression was assessed using FACS analysis, and cell viability assessed using alamar blue and MTT assays. In addition, clogenic survival was assessed by examining the cell survival fraction. IKKα-selective small molecule kinase inhibitors SU1261 (Ki α vs. β; 10 vs 680nM) and SU1266 (Ki α vs. β; 2 vs. 77nM) were synthesised in-house as part of a CRUK small molecule drug discovery programme (PI Prof. S. MacKay).

Results- U2OS cells and prostate cancer cells (PC3m and LNCAP AI)were treated with FCS (10%) or lymphotoxinα1β2 respectively and increased the phosphorylation of p100 and the formation of p52 NFκB, as two non-canonical NFκB pathway markers. These responses were also inhibited in a concentration dependent manner by SU1261 (IC50 = 2μM) and also the non-selective inhibitor SU1266 (IC501.87μM). However, SU1266 reversed two responses indicative of IKKβ inhibition; the degradation of IκBα and the phosphorylation of p65NFκB (Ser536). In contrast, SU1261 was without effect on these two markers at concentrations up to 30μM, indicating selectivity for IKKα. Although, SU1261 showed no effect on cell cycle profile in both cancer cell types, the clonogenic assay showed SU1261 inhibited both U2OS (IC501.58 +/- 0.1μM) and PC3m (IC502.10 +/- 0.46μM)survival and proliferation.

Conclusions/Implications- These studies reveal for the first time the potential of IKKα inhibition as a new therapeutic target in osteosarcoma and prostate cancers. Compounds of this nature will circumvent the limits imposed by selective IKKβ inhibition and may have future potential use as chemotherapeutics.

Acknowledgements- This work was sponsored by CRUK, PCUK and MRC

Gamble C, McIntosh K, Scott R, Ho KH, Plevin R, Paul A. 2012Br J Pharmacol. 2012 Feb;165(4): 802-19. doi: 10.1111/j.1476-5381.2011.01608.x.

Parrondo R, de las Pozas A, Reiner T, Rai P, Perez-Stable C. 2010Mol Cancer. 2010 Jul 9;9: 182. doi: 10.1186/1476-4598-9-182.