ABSTRACT
Chronic myeloid leukemia (CML) is a myeloproliferative disease caused by bcr-abl1, a constitutively active tyrosine kinase fusion gene responsible for an abnormal proliferation of leukemic stem cells (LSCs). Inhibition of BCR-ABL1 kinase activity offers long-term relief to CML patients. However, for a proportion of them, BCR-ABL1 inhibition will become ineffective at treating the disease, and CML will progress to blast crisis (BC) CML with poor prognosis. BC-CML is often associated with excessive phosphorylated eukaryotic translation initiation factor 4E (eIF4E), which renders LSCs capable of proliferating via self-renewal, oblivious to BCR-ABL1 inhibition. In vivo, eIF4E is exclusively phosphorylated on Ser209 by MNK1/2. Consequently, a selective inhibitor of MNK1/2 should reduce the level of phosphorylated eIF4E and re-sensitize LSCs to BCR-ABL1 inhibition, thus hindering the proliferation of BC LSCs. We report herein the structure-activity relationships and pharmacokinetic properties of a selective MNK1/2 inhibitor clinical candidate, ETC-206, which in combination with dasatinib prevents BC-CML LSC self-renewal in vitro and enhances dasatinib antitumor activity in vivo.
Subject(s)
Blast Crisis/drug therapy , Cell Proliferation , Intracellular Signaling Peptides and Proteins/antagonists & inhibitors , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Animals , Blast Crisis/pathology , Female , Humans , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/pathology , Mice , Mice, SCID , Models, Molecular , Molecular Structure , Protein Conformation , Protein Kinase Inhibitors/chemistry , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
Current JAK2 inhibitors used for myeloproliferative neoplasms (MPN) treatment are not specific enough to selectively suppress aberrant JAK2 signalling and preserve physiological JAK2 signalling. We tested whether combining a JAK2 inhibitor with a series of serine threonine kinase inhibitors, targeting nine signalling pathways and already used in clinical trials, synergized in inhibiting growth of haematopoietic cells expressing mutant and wild-type forms of JAK2 (V617F) or thrombopoietin receptor (W515L). Out of 15 kinase inhibitors, the ZSTK474 phosphatydylinositol-3'-kinase (PI3K) inhibitor molecule showed strong synergic inhibition by Chou and Talalay analysis with JAK2 and JAK2/JAK1 inhibitors. Other pan-class I, but not gamma or delta specific PI3K inhibitors, also synergized with JAK2 inhibitors. Synergy was not observed in Bcr-Abl transformed cells. The best JAK2/JAK1 and PI3K inhibitor combination pair (ruxolitinib and GDC0941) reduces spleen weight in nude mice inoculated with Ba/F3 cells expressing TpoR and JAK2 V617F. It also exerted strong inhibitory effects on erythropoietin-independent erythroid colonies from MPN patients and JAK2 V617F knock-in mice, where at certain doses, a preferential inhibition of JAK2 V617F mutated progenitors was detected. Our data support the use of a combination of JAK2 and pan-class I PI3K inhibitors in the treatment of MPNs.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Janus Kinase 2/antagonists & inhibitors , Myeloproliferative Disorders/drug therapy , Phosphoinositide-3 Kinase Inhibitors , Animals , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Drug Screening Assays, Antitumor , Drug Synergism , Female , Gene Knock-In Techniques , Hematologic Neoplasms/drug therapy , Hematologic Neoplasms/enzymology , Humans , Janus Kinase 2/genetics , Janus Kinase 2/metabolism , Mice , Mice, Nude , Mice, Transgenic , Mutation, Missense , Myeloproliferative Disorders/enzymology , Neoplasm Transplantation , Nitriles , Phosphatidylinositol 3-Kinases/metabolism , Pyrazoles/administration & dosage , Pyrimidines , Pyrrolidines/administration & dosage , Signal Transduction/drug effects , Sulfonamides/administration & dosage , Triazines/administration & dosage , Tumor Cells, CulturedABSTRACT
Tankyrases constitute potential drug targets for cancer and myelin-degrading diseases. We have applied a structure- and biophysics-driven fragment-based ligand design strategy to discover a novel family of potent inhibitors for human tankyrases. Biophysical screening based on a thermal shift assay identified highly efficient fragments binding in the nicotinamide-binding site, a local hot spot for fragment binding. Evolution of the fragment hit 4-methyl-1,2-dihydroquinolin-2-one (2) along its 7-vector yields dramatic affinity improvements in the first cycle of expansion. A crystal structure of 7-(2-fluorophenyl)-4-methylquinolin-2(1H)-one (11) reveals that the nonplanar compound extends with its fluorine atom into a pocket, which coincides with a region of the active site where structural differences are seen between tankyrases and other poly(ADP-ribose) polymerase (PARP) family members. A further cycle of optimization yielded compounds with affinities and IC50 values in the low nanomolar range and with good solubility, PARP selectivity, and ligand efficiency.
