Acute myelogenous leukemia (AML), a disease of the blood and bone marrow, is characterized by the inability of myeloblasts to differentiate into mature cell types. Dihydroorotate dehydrogenase (DHODH) is an enzyme well-known in the pyrimidine biosynthesis pathway; however, small molecule DHODH inhibitors were recently shown to induce differentiation in multiple AML subtypes. Using virtual screening and structure-based drug design approaches, a new series of N-heterocyclic 3-pyridyl carboxamide DHODH inhibitors were discovered. Two lead compounds, 19 and 29, have potent biochemical and cellular DHODH activity, favorable physicochemical properties, and efficacy in a preclinical model of AML.

Enhanced NF-kB signaling is a hallmark of aggressive lymphoid malignancies, including multiple myeloma (MM), mantle cell lymphoma (MCL), and diffuse large B-cell lymphoma (DLBCL). Non-canonical NF-kB signaling involves NIK-dependent activation of IKKα, which triggers nuclear accumulation of p52/RelB heterodimers. NIK is a highly unstable protein and degradation is mediated by a ubiquitin ligase complex consisting of TRAF2, TRAF3 and c-IAP1/2 (encoded by BIRC2/3). In a subset of MM, NIK is stabilized by mutations in NIK, TRAF2/3 or BIRC2/3 (Annunziata et al./Keats et al., Canc. Cell 2007). Similar activating mutations in the non-canonical NF-kB signaling pathway were recently found in ibrutinib-refractory MCL cell lines (Rahal et al., Nat. Med. 2014), and in DLBCL (Zhang et al., Cell Rep. 2015). In many other cases of B-cell malignancies, NIK is stabilized by high level expression of the upstream TNF receptors (BAFFR, CD40, LTβR) or high abundance of their ligands in the bone marrow or the lymph nodes. To date, bioavailable NIK inhibitors have not been reported, and it has remained unclear whether NIK inhibitors are effective and tolerated in mouse models of B-cell malignancies associated with activation of NIK. Here, we report for the first time on a potent orally bioavailable NIK kinase inhibitor, TRC694. TRC694 selectively inhibits NIK enzymatic activity, translating into inhibition of phospho-IKKα in NIK and in TRAF3 mutant cell lines with single digit nM IC50. TRC694 prevents nuclear accumulation of p52/RelB (but not canonical NF-kB) and represses the associated NF-kB gene program selectively in MM cell lines with genetic activation of the non-canonical NF-kB pathway. Proliferation of NIK translocated, TRAF3 or BIRC3 mutant MM cell lines is inhibited by low nM concentrations of TRC694, whereas MM cell lines which lack genetic activation of non-canonical NF-kB are much less sensitive to TRC694. Consistently, elevated expression of a previously described 11-gene NFkB signature in MM (Annunziata et al., Canc. Cell 2007) is predictive of sensitivity to TRC694 in a 21-MM cell line panel. A single, oral dose of 10 to 40 mg/kg of TRC694 to mice bearing a NIK-translocated MM tumor (JJN-3), was sufficient to inhibit phospho-IKKα and repress P52-mediated transcription of NFkB regulated genes in the tumors. Consistently, once-daily, oral dosing of TRC694 to mice bearing subcutaneous NIK translocated (JJN-3) or TRAF3 (RPMI-8226, MM.1S) mutant MM tumors, completely inhibits growth of these tumors at doses of 10 to 40 mg/kg, with no signs of toxicities. In conclusion, TRC694 is a first-in-class orally bioavailable NIK kinase inhibitor, and provides the first opportunity to test the clinical relevance of non-canonical NF-kB inhibition in aggressive lymphoid malignancies. Citation Format: Matthias Versele, Lut Janssen, Tamara Geerts, Wim Floren, Boudewijn Janssens, Hillary Millar, Edgar Jacoby, Gerhard Gross, Yannick Ligny, Yvan Simonnet, Nathalie Amblard, Olivier Querolle, Imre Csoka, Virginie Poncelet, Virginie Tronel, Sophie Nocquet-Thibault, Lieven Meerpoel, James Edwards, Marc Salvati, Sriram Balasubramanian, Laurie Lenox, Charles Theuer, Ricardo Attar, Ian Stansfield. Inhibition of NF-kB inducing kinase (NIK) selectively abrogates NIK and TRAF3 mutant multiple myeloma tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4199. doi:10.1158/1538-7445.AM2017-4199

Abstract 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 >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 > 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

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Fibroblast growth factors (FGFs) and their receptors (FGFR1 through 4) regulate a variety of key cellular processes, including proliferation, migration, survival, and differentiationa. Aberrant activation of FGF/FGFR is strongly implicated in oncogenic signalling in many tumor types. This has stimulated the development of a number of FGFR inhibitors, with diverse kinase inhibition and pharmacological profiles that are currently being evaluated in clinical studies. We conducted a fragment screening campaign and this resulted in identification of a 6-aminoquinoxalinyl fragment with a binding affinity in the micromolar range. Structure-guided medicinal chemistry led to the identification of a novel quinoxaline-based chemical series with nanomolar affinity for FGFR1, 2, 3, and 4, activity in cells, and selectivity with respect to VEGFR-2. Further optimisation resulted in the generation of JNJ-42756493, a compound with favourable drug-like properties that demonstrated strong anti-tumoral activity in a FGFR2-dependent SNU-16 human gastric carcinoma xenograft model. This report represents the first disclosure of the structure-activity relationships as well as the chemical synthesis pathway of the JNJ-42756493 series and illustrates how a fragment-based drug discovery approach has been efficiently used to discover FGFR1-4 inhibitors with nanomolar affinity. aTurner, N. and Grose, R. Nat. Rev. Cancer, 2010, 10, 116-129. Citation Format: Patrick R. Angibaud, Laurence Mevellec, Gordon Saxty, Christophe Adelinet, Rhalid Akkari, Valerio Berdini, Pascal Bonnet, Marine Bourgeois, Xavier Bourdrez, Anne Cleasby, Helene Colombel, Imre Csoka, Werner Embrechts, Eddy Freyne, Ronaldus Gilissen, Eleonora Jovcheva, Peter King, Jean Lacrampe, Delphine Lardeau, Yannick Ligny, Steve Mcclue, Lieven Meerpoel, David R. Newell, Martin Page, Alexandra Papanikos, Elisabeth Pasquier, Isabelle Pilatte, Virginie Poncelet, Olivier Querolle, David C. Rees, Sharna Rich, Bruno Roux, Elodie Sement, Yvan Simonnet, Matthew Squires, Virginie Tronel, Tinne Verhulst, Jorge Vialard, Marc Willems, Steven J. Woodhead, Berthold Wroblowski, Christopher W. Murray, Timothy Perera. Discovery of JNJ-42756493, a potent fibroblast growth factor receptor (FGFR) inhibitor using a fragment based approach. [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 4748. doi:10.1158/1538-7445.AM2014-4748

We describe the discovery and the structure-activity relationship of a new series of quinoline derivatives acting as selective and highly potent noncompetitive mGlu1 antagonists. We first identified cis-10 as a fairly potent mGlu1 antagonist (IC(50) = 20 nM) in a cell-based signal transduction assay on the rat mGlu1 receptor expressed in CHO-K1 cells, and then we were able to design and synthesize highly potent compounds on both rat and human mGlu1 receptors as exemplified by compound cis-64a, which has an antagonist potency of 0.5 nM for the human mGlu1 receptor. We briefly present and discuss the in vitro metabolic stability of the compounds in human liver microsomes. We finally report the pharmacokinetic properties of our lead compound cis-64a.