Small Molecule c-Met Research Articles

Small molecule c-Met kinase inhibitors: a review of recent patents

Importance of the field: c-Met kinase is the receptor for hepatocyte growth factor. Primarily expressed on epithelial and mesenchymal cells its normal function is associated with wound healing, liver regeneration and embryo development. However, dysregulation of c-Met through overexpression, gene amplification, mutation or a ligand-dependent autocrine/paracrine loop is associated with tumorigenesis. c-Met dysregulation in human cancer patients is typically associated with a poor prognosis, aggressive disease, increased metastasis and shortened patient survival. Targeting the hepatocyte growth factor/c-Met signalling pathway as a means of cancer therapy has, therefore, become increasingly popular with a number of different therapeutic approaches undergoing clinical trials.Areas covered by this review: This review covers the patent applications for small molecule c-Met kinase inhibitors since 2007, attempts to place them in context from a structural point of view and examines their potential applications in cancer therapy.What the reader will gain: Readers will gain an overview of the structural types of c-Met inhibitors, the major players in the field and an insight into what is progressing into the clinic.Take home message: This area is developing rapidly and the results of the various ongoing clinical trials will generate an increased understanding of the potential benefits and pitfalls of c-Met inhibitors as therapeutic agents.

Discovery of Small Molecule c-Met Inhibitors: Evolution and Profiles of Clinical Candidates

The scatter factor/hepatocyte growth factor (HGF)-c-Met axis is involved in the malignant phenotype of various tumor types via activation of a wide range of autocrine and paracrine processes. Autocrine activation of tumor cell c-Met receptors enhances tumor cell proliferation, angiogenesis, invasion/metastasis and resistance to apoptosis and cytotoxic therapies. In addition, tumor and stroma cell-derived HGF functions as a potent angiogenic factor. Therefore, the HGF-c-Met axis is critically involved in multiple facets of normal cellular growth and homeostasis and activated in a dysregulated manner in a variety of cancers. Consequently, inhibiting the HGF-c-Met axis would be anticipated to have potent anti-tumor effects in many cancers through multiple complimentary mechanisms including increased sensitivity to current cytotoxic chemo-and radiotherapies. The acceptance of c-Met as a tractable target for cancer therapy has fostered intensive drug discovery efforts across the pharmaceutical industry. This research has led to 20 published crystal structures (with and without ligands) that revealed two distinct binding modes for ATP-competitive inhibitors: Type I ligands which assumes a U shape geometry through interactions with both hinge and activation loop residue Y1230, and Type II ligands which adopt a more extended orientation, either binding a conventional DFG-out conformation or protein conformations with varying degrees of 'DFG-out' character. Nearly a dozen small molecule c-Met inhibitors have entered human clinical trials ranging from Type I inhibitors solely selective for c-Met to Type I inhibitors with broader kinase activities to Type II inhibitors with "spectrum selective" kinase activity. The identification, profiles and properties of these clinical candidates are summarized in this review.

Abstract C190: BMS-777607, a potent small molecule c-MET inhibitor, inhibits prostate cancer progression in vitro

Abstract Overexpression of c-MET is highly associated with prostate cancer progression and therapy resistance. BMS-777607 is a selective and potent c-MET inhibitor that was advanced to clinical evaluation. Herein we report the effects of BMS-777607 in androgen-independent prostate cancer cells (PC-3, DU145). When assessed using an MTT assay, BMS-777607 treatment was found to inhibit cell growth in the absence of hepatocyte growth factor (HGF), with an IC50 >5.0 uM for both cell lines. In a wound-healing assay, BMS-777607 suppressed PC-3 and HGF-induced DU145 cell motility in a dose-dependent fashion (IC50 values of 3.0 and 0.5 uM, respectively). Moreover, BMS-777607 efficiently blocked HGF-triggered DU145 cell scattering in a cell scatter assay; with 40% inhibition at 0.2 uM. Using a Matrigel-based Transwell assay, BMS-777607 also inhibited PC-3 cell invasion in a dose-dependent manner, with 60% inhibition occurring at 1 uM. Western blot analysis demonstrated that BMS-777607 blocked c-MET autophosphorylation, and downregulated multiple downstream kinases involved in c-MET signaling pathway, including Src, Fak and Akt. These data indicate that BMS-777607 inhibited prostate cancer progression in vitro including proliferation, motility, scattering and invasion. Furthermore, the finding of more potent effects on cell migration and invasion than cell proliferation suggests that this small molecule c-MET inhibitor could have utility as an anti-metastatic agent for the treatment of advanced prostate cancer. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C190.

The c-Met receptor tyrosine kinase inhibitor MP470 radiosensitizes glioblastoma cells

PurposeGlioblastoma multiforme (GBM) is resistant to current cytotoxic therapies, in part because of enhanced DNA repair. Activation of the receptor tyrosine kinase c-Met has been shown to protect cancer cells from DNA damage. We hypothesized that inhibiting c-Met would decrease this protection and thus sensitize resistant tumor cells to the effects of radiation therapy.Materials and methodsEight human GBM cell lines were screened for radiosensitivity to the small-molecule c-Met inhibitor MP470 with colony-count assays. Double-strand (ds) DNA breaks was quantified by using antibodies to gamma H2AX. Western blotting demonstrate expression of RAD51, glycogen synthase kinase (GSK)-3β, and other proteins. A murine xenograft tumor flank model was used for in vivo radiosensitization studies.ResultsMP470 reduced c-Met phosphorylation and enhanced radiation-induced cell kill by 0.4 logs in SF767 cells. Cells pretreated with MP470 had more ds DNA damage than cells treated with radiation alone. Mechanistically, MP470 was shown to inhibit dsDNA break repair and increase apoptosis. MP470 influences various survival and DNA repair related proteins such as pAKT, RAD51 and GSK3β. In vivo, the addition of MP470 to radiation resulted in a tumor-growth-delay enhancement ratio of 2.9 over radiation alone and extended survival time.ConclusionsGBM is a disease site where radiation is often used to address both macroscopic and microscopic disease. Despite attempts at dose escalation outcomes remain poor. MP470, a potent small-molecule tyrosine kinase inhibitor of c-Met, radiosensitized several GBM cell lines both in vitro and in vivo, and may help to improve outcomes for patients with GBM.

Bone marrow mesenchymal stem cells undergo nemosis and induce keratinocyte wound healing utilizing the HGF/c‐Met/PI3K pathway

We previously showed cell-cell contacts of human dermal fibroblasts to induce expression of the hepatocyte growth factor/scatter factor (HGF) in a process designated as nemosis. Now we report on nemosis initiation in bone marrow mesenchymal stem cells (BMSCs). Because BMSCs are being used increasingly in cell transplantation therapy we aimed to demonstrate a functional effect and benefit of BMSC nemosis for wound healing. Nemotic and monolayer cells were used to stimulate HaCaT keratinocyte migration in a scratch-wound healing assay. Both indicators of nemosis, HGF production and cyclooxygenase-2 expression, were induced in BMSC spheroids. When compared with a similar amount of cells as monolayer, nemotic cells induced keratinocyte in vitro scratch-wound healing in a concentration-dependent manner. The HGF receptor, c-Met, was rapidly phosphorylated in the nemosis-stimulated keratinocytes. Nemosis-induced in vitro scratch-wound healing was inhibited by an HGF-neutralizing antibody as well as the small molecule c-Met inhibitor, SU11274. HGF-induced in vitro scratch-wound healing was inhibited by PI3K inhibitors, wortmannin and LY294002, while LY303511, an inactive structural analogue of LY294002, had no effect. Inhibitors of the mitogen-activated protein kinases MEK/ERK1/2 (PD98059 and U0126), and p38 (SB203580) attenuated HGF-induced keratinocyte in vitro scratch-wound healing. We conclude that nemosis of BMSCs can induce keratinocyte in vitro scratch-wound healing, and that in this effect signaling via HGF/c-Met is involved.

Targeting the c-MET signaling pathway for cancer therapy

Background: In many human cancers, c-MET is activated via receptor overexpression, amplification, mutation and/or a ligand-dependent autocrine/paracrine loop. These biochemical and genetic abnormalities have been correlated with poor clinical outcomes and drug resistance in cancer patients. Preclinical studies suggest that targeting aberrant c-MET signaling could be an attractive therapy in cancer, but this notion has only recently been tested in the clinic. Objectives: To describe the biological aspects of the c-MET signaling pathway and to discuss recent progress and possible future trends in the development of agents that target the c-MET pathway, with an emphasis on small-molecule c-MET kinase inhibitors. Method: A review of relevant publications, including published articles in literature, reports at scientific meetings, and information available through the Internet. Results/conclusion: The dysregulated c-MET pathway represents a promising target for cancer drug development. The agents that target the c-MET pathway have demonstrated impressive evidence of early clinical activity and may have a significant therapeutic potential.

Small molecule c‐MET inhibitor PHA665752: Effect on cell growth and motility in papillary thyroid carcinoma

c-Met is upregulated in papillary thyroid carcinoma (PTC) and can be an attractive therapeutic target. We tested the effects of the small molecule c-met inhibitor PHA665752 in blocking c-met-dependent phenotypic effects in PTC cell lines. PTC patient tissues and cell lines were evaluated for c-met expression. The effect of PHA665752 on c-met phosphorylation, downstream signaling, hepatocyte growth factor (HGF)-dependent cell growth, and induction of apoptosis was studied. The IC(50) of PHA665752 in c-met-expressing PTC cells was determined, and growth curves at 0.1x, 1x, and 10x IC(50) concentrations were obtained. Poly(ADP-ribose) polymerase (PARP) and caspase-9-processing post-PHA665752 treatment were studied as markers of apoptosis, and assays analyzing HGF-dependent cell invasion and migration in the presence and absence of PHA665752 were done. c-Met was upregulated in most of the patient tissues with PTC and in many PTC cell lines. PHA665752 specifically inhibited c-met phosphorylation, c-met-dependent cell growth, signal transduction, cell survival, cell invasion, and migration in PTC cells with high c-met. PHA665752 is an effective and specific inhibitor of c-met in PTC cells with high levels of c-met expression.

Inhibitors targeting hepatocyte growth factor receptor and their potential therapeutic applications

The hepatocyte growth factor (HGF)/c-Met signaling pathway is important in mediating a wide range of biological activities, including embryological development, wound healing, tissue regeneration, angiogenesis, proliferation, survival, scattering, motility, invasion and morphogenic differentiation. HGF and/or c-Met are expressed at abnormally high levels in a large variety of solid tumors. Various c-Met mutations have been described in many solid tumors and some hematologic malignancies. Therefore, inhibition of the HGF/c-Met signaling pathway has great potential in molecular targeted cancer therapy. Many patent applications associated with inhibition of the HGF/c-Met axis have been published in the past few years. There are four small molecule c-Met inhibitors presently in clinical trials. This review focusses on recent patent applications for small molecule c-Met inhibitors and their potential applications in cancer therapy.

Activation of HGF/c-Met pathway contributes to the reactive oxygen species generation and motility of small cell lung cancer cells

Small cell lung cancer (SCLC) is a difficult disease to treat and sometimes has overexpression or mutation of c-Met receptor tyrosine kinase. The effects of c-Met/hepatocyte growth factor (c-Met/HGF, ligand for c-Met) on activation of reactive oxygen species (ROS) was determined. HGF stimulation of c-Met-overexpressing H69 SCLC cells (40 ng/ml, 15 min) resulted in an increase of ROS, measured with fluorescent probe 2'-7'-dichlorofluorescein diacetate (DCFH-DA) or dihydroethidine (DHE) but not in c-Met-null H446 cells. ROS was increased in juxtamembrane (JM)-mutated variants (R988C and T1010I) of c-Met compared with wild-type c-Met-expressing cells. ROS was significantly inhibited by preincubation of SCLC cells with pyrrolidine dithiocarbamate (PDTC, 100 microM) and/or SU11274 (small molecule c-Met tyrosine kinase inhibitor, 2 microM) for 3 h. PDTC and SU11274 also abrogated the HGF proliferative signal and cell motility in a cooperative fashion. H(2)O(2) treatment of SCLC cells (over 15 min) led to phosphorylation of c-Met receptor tyrosine kinase and further upregulated downstream phosphorylation of phospho-AKT, ERK1/2, and paxillin in a dose-dependent manner (125 microM to 500 microM). c-Met is an important target in lung cancer, and the pathways responsible for ROS generation together may provide novel therapeutic intervention.

Functional Analysis of c-Met/Hepatocyte Growth Factor Pathway in Malignant Pleural Mesothelioma

c-Met receptor tyrosine kinase (RTK) has not been extensively studied in malignant pleural mesothelioma (MPM). In this study, c-Met was overexpressed and activated in most of the mesothelioma cell lines tested. Expression in MPM tissues by immunohistochemistry was increased (82%) in MPM in general compared with normal. c-Met was internalized with its ligand hepatocyte growth factor (HGF) in H28 MPM cells, with robust expression of c-Met. Serum circulating HGF was twice as high in mesothelioma patients as in healthy controls. There was a differential growth response and activation of AKT and extracellular signal-regulated kinase 1/2 in response to HGF for the various cell lines. Dose-dependent inhibition (IC50 < 2.5 micromol/L) of cell growth in mesothelioma cell lines, but not in H2052, H2452, and nonmalignant MeT-5A (IC50 > 10 micromol/L), was observed with the small-molecule c-Met inhibitor SU11274. Furthermore, migration of H28 cells was blocked with both SU11274 and c-Met small interfering RNA. Abrogation of HGF-induced c-Met and downstream signaling was seen in mesothelioma cells. Of the 43 MPM tissues and 7 cell lines, we have identified mutations within the semaphorin domain (N375S, M431V, and N454I), the juxtamembrane domain (T1010I and G1085X), and an alternative spliced product with deletion of the exon 10 of c-Met in some of the samples. Interestingly, we observed that the cell lines H513 and H2596 harboring the T1010I mutation exhibited the most dramatic reduction of cell growth with SU11274 when compared with wild-type H28 and nonmalignant MeT-5A cells. Ultimately, c-Met would be an important target for therapy against MPM.

A Selective Small Molecule c-MET Inhibitor, PHA665752, Cooperates with Rapamycin

c-MET is believed to be an attractive receptor target for molecular therapeutic inhibition. TPR-MET, a constitutively active oncogenic variant of MET, serves as excellent model for testing c-MET inhibitors. Here, we characterized a small molecule c-MET inhibitor, PHA665752, and tested its cooperation with the mammalian target of rapamycin inhibitor as potential targeted therapy. The effect of PHA665752 treatment was determined on cell growth, motility and migration, apoptosis, and cell-cycle arrest of TPR-MET-transformed cells. Moreover, the effect of PHA665752 on the phosphorylation on MET, as well as its downstream effectors, p-AKT and p-S6K, was also determined. Finally, growth of TPR-MET-transformed cells was tested in the presence of PHA665752 and rapamycin. H441 non-small cell lung cancer (NSCLC) cells (with activated c-Met) were also tested against both PHA665752 and rapamycin. PHA665752 specifically inhibited cell growth in BaF3. TPR-MET cells (IC(50) < 0.06 micromol/L), induced apoptosis and cell cycle arrest. Constitutive cell motility and migration of the BaF3. TPR-MET cells was also inhibited. PHA665752 inhibited specific phosphorylation of TPR-MET as well as phosphorylation of downstream targets of the mammalian target of rapamycin pathway. When combined with PHA665752, rapamycin showed cooperative inhibition to reduce growth of BaF3. TPR-MET- and c-MET-expressing H441 NSCLC cells. PHA665752 is a potent small molecule-selective c-MET inhibitor and is highly active against TPR-MET-transformed cells both biologically and biochemically. PHA665752 is also active against H441 NSCLC cells. The c-MET inhibitor can cooperate with rapamycin in therapeutic inhibition of NSCLC, and in vivo studies of this combination against c-MET expressing cancers would be merited.