Abstract 3228: Characterization of a novel, orally bioavailable, potent and highly selective small molecule inhibitor of PERK: A tool to probe the biphasic concentration-dependent induction of ER stress in models of multiple myeloma and B-cell lymphoma
Authors: Stansfield Ian , Ligny Yannick , Simonnet Yvan , Demestre Christophe , Amblard Nathalie , Meyer Christophe , Geerts Tamara , Ven Jeroen Van de , Wyngaert Ilse Van den , Vermeulen Peter , Beerden Inge , Peeters Danielle , Liebregts Johnny , Baelen Kurt Van , Simillion Cedric , Janssen Boudewijn , Verhulst Tinne , Esser Norbert , Bischoff James , Meerpoel Lieven , Versele Matthias 
Published: May 2015

The unfolded protein response (UPR) is a signal transduction pathway that coordinates cellular adaptation to microenvironmental stress and the accumulation of malfolded proteins in the endoplasmic reticulum (ER). In highly secretory malignancies, such as multiple myeloma (MM), characteristically there is both an extensively developed ER and a markedly elevated UPR to ensure a homeostatic balance between ER burden and ER capacity - thus making components of the UPR attractive targets for therapeutic intervention. One such target is PERK (PKR-like endoplasmic reticulum kinase), a member of the eukaryotic initiation factor (eIF)2α kinase family, that is activated during the UPR to chronic ER stress.

Here, we describe the identification and characterization of compound (1) - a novel, orally bioavailable, potent and highly selective small molecule inhibitor of PERK. (1) is a sub-nM inhibitor of PERK, with a &gt;100-fold window against other kinases (including other eIF2α kinases) - targeting only 6 out of a 400 kinase panel with sub-μM IC50's. It inhibits phosphorylation of eIF2α at 10 nM (IC50) in HEK293 cells (incubated with the ER stressor tunicamycin) and proves to be selectively anti-proliferative in an ER-stressed epithelial cancer model (A549 cells with tunicamycin) at nM concentrations, but to a lesser extent in the absence of ER stress. Furthermore, in the absence of an exogenous ER stressor, (1) induced excessive ER stress (eg, as evidenced by induction of the pro-apoptotic CHOP gene), and decreased cell viability selectively in MM cell lines and certain B-cell lymphoma lines at low nM concentrations, but not in normal or malignant epithelial cells. However, despite a sustained impact on eIF2α phosphorylation, interestingly compound (1) elicited a biphasic concentration dependent induction of ER stress (and consequent inhibition of cell viability) - with ER stress induction maximal at inhibitor concentrations corresponding to circa 50-75 % inhibition of PERK, returning progressively to baseline with concentrations rising to &gt; IC90. Whole-genome expression profiling revealed that all significant changes seen at 10-100 nM of (1), returned to baseline levels at 1000 nM, suggesting a compensatory mechanism very proximal to phosphorylation of eIF2α. In summary, the highly selective PERK inhibitor, (1), validated the unique sensitivity of subsets of multiple myeloma and B-cell lymphoma to disruption of the UPR, but unexpectedly revealed a compensatory mechanism associated with potent PERK inhibition.

Citation Format: Ian Stansfield, Yannick Ligny, Yvan Simonnet, Christophe Demestre, Nathalie Amblard, Christophe Meyer, Tamara Geerts, Jeroen Van de Ven, Ilse Van den Wyngaert, Peter Vermeulen, Inge Beerden, Danielle Peeters, Johnny Liebregts, Kurt Van Baelen, Cedric Simillion, Boudewijn Janssen, Tinne Verhulst, Norbert Esser, James Bischoff, Lieven Meerpoel, Matthias Versele. Characterization of a novel, orally bioavailable, potent and highly selective small molecule inhibitor of PERK: A tool to probe the biphasic concentration-dependent induction of ER stress in models of multiple myeloma and B-cell lymphoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3228. doi:10.1158/1538-7445.AM2014-3228