Discovery of 1H‑Pyrazol-3(2H)‑ones as Potent and Selective Inhibitors of Protein Kinase R‑like Endoplasmic Reticulum Kinase (PERK) Adrian L. Smith,* ,† Kristin L. Andrews, ‡ Holger Beckmann, ¶ Steven F. Bellon, ⊥ Pedro J. Beltran, § Shon Booker, † Hao Chen, # Young-Ah Chung, § Noel D. D’Angelo, † Jennifer Dao, ‡ Kenneth R. Dellamaggiore, § Peter Jaeckel, ¶ Richard Kendall, § Katja Labitzke, ¶ Alexander M. Long, ⊥ Silvia Materna-Reichelt, ¶ Petia Mitchell, § Mark H. Norman, † David Powers, ‡ Mark Rose, ∥ Paul L. Shaffer, ⊥ Michelle M. Wu, § and J. Russell Lipford* ,§ Departments of † Medicinal Chemistry, ‡ Molecular Structure and Characterization, § Oncology Research, and ∥ Pharmacokinetics and Drug Metabolism, Amgen Inc., One Amgen Center Drive, Thousand Oaks, California 91320-1799, United States Departments of ⊥ Molecular Structure and Characterization and # Protein Technologies, Amgen Inc., 360 Binney Street, Cambridge, Massachusetts 02142, United States ¶ Amgen Research GmbH, Josef-Engert-Straße 11, D-93053 Regensburg, Germany * S Supporting Information ABSTRACT: The structure-based design and optimization of a novel series of selective PERK inhibitors are described resulting in the identification of 44 as a potent, highly selective, and orally active tool compound suitable for PERK pathway biology exploration both in vitro and in vivo. ■ INTRODUCTION The unfolded protein response (UPR) is an evolutionarily conserved mechanism in mammals by which cells respond to endoplasmic reticulum (ER) stress. 1 Cellular ER stress is characterized by an accumulation of misfolded proteins in the ER lumen resulting from an inability of the cell to adequately process protein production in the ER, for example, under nutrient deprivation conditions (hypoxia, glucose starvation) or where a high secretory load exists. There are three sensors of misfolded proteins that are known to mediate the UPR through complementary pathways: protein kinase R-like endoplasmic reticulum kinase (PERK), inositol-requiring enzyme 1 (IRE1), and activating transcription factor 6 (ATF6). 2 PERK, IRE1, and ATF6 are all located in the ER membrane and sense misfolded proteins in the ER lumen through the molecular chaperone BiP/Grp78 and/or direct binding of unfolded proteins to the sensor domains. 3 The UPR serves as a mechanism for cellular survival whereby cells are able to adapt to cope with ER stress, but under extreme stress the UPR switches the cellular machinery toward apoptosis. 4−6 The UPR is known to be active in highly secretory tissues such as the pancreas, 7 but tumors are also thought to utilize the UPR for survival under stressed conditions such as nutrient deprivation or chemo- therapeutic insult. 8 This suggests that modulators of the UPR may be useful for the treatment of cancer 9−11 either as a sole agent or in combination with other anticancer treatments, thus spurring recent interest in the identification of UPR pathway inhibitors. PERK is an ER transmembrane protein with a stress-sensing domain inside the ER lumen and a cytosolic kinase domain. 10 Upon sensing misfolded proteins, PERK is activated by autophosphorylation and oligomerization through release of BiP/Grp78 from the stress-sensing domain. Activated PERK phosphorylates and activates its downstream substrate, eukaryotic initiation factor 2α (eIF2α), which inhibits the ribosome translation initiation complex in order to attenuate protein synthesis. This serves to prevent exacerbation of ER stress by preventing the accumulation of additional misfolded proteins. Although it inhibits general protein synthesis, activated eIF2α causes the translation of specific mRNAs involved in restoring ER homeostasis including activating transcription factor 4 (ATF4). ATF4 mediates the transcription of certain UPR target genes including those for the endoplasmic-reticulum-associated protein degradation (ERAD) pathway proteins which target misfolded proteins Received: November 11, 2014 Article pubs.acs.org/jmc © XXXX American Chemical Society A DOI: 10.1021/jm5017494 J. Med. Chem. XXXX, XXX, XXX−XXX