A selective small molecule inhibitor of c-Met, PHA665752, inhibits tumorigenicity and angiogenesis in mouse lung cancer xenografts

The c-Met receptor tyrosine kinase is emerging as a novel target in many solid tumors, including lung cancer. PHA-665752 was identified as a small molecule, ATP competitive inhibitor of the catalytic activity of the c-Met kinase. Here, we show that treatment with PHA665752 reduced NCI-H69 (small cell lung cancer) and NCI-H441 (non-small cell lung cancer) tumorigenicity in mouse xenografts by 99% and 75%, respectively. Reduction in tumor size was also observed by magnetic resonance imaging of tumors in mice. PHA665752 inhibited c-Met phosphorylation at the autophosphorylation and c-Cbl binding sites in mouse xenografts derived from non-small cell lung cancer cell lines (NCI-H441 and A549) and small cell lung cancer cell line (NCI-H69). PHA665752 also inhibited angiogenesis by >85% in all the abovementioned cell lines and caused an angiogenic switch which resulted in a decreased production of vascular endothelial growth factor and an increase in the production of the angiogenesis inhibitor thrombospondin-1. These studies show the feasibility of selectively targeting c-Met with ATP competitive small molecule inhibitors and suggest that PHA665752 may provide a novel therapeutic approach to lung cancer.     

An orally available small-molecule inhibitor of c-Met, PF-2341066, exhibits cytoreductive antitumor efficacy through antiproliferative and antiangiogenic mechanisms

The c-Met receptor tyrosine kinase and its ligand, hepatocyte growth factor (HGF), have been implicated in the progression of several human cancers and are attractive therapeutic targets. PF-2341066 was identified as a potent, orally bioavailable, ATP-competitive small-molecule inhibitor of the catalytic activity of c-Met kinase. PF-2341066 was selective for c-Met (and anaplastic lymphoma kinase) compared with a panel of >120 diverse tyrosine and serine-threonine kinases. PF-2341066 potently inhibited c-Met phosphorylation and c-Met-dependent proliferation, migration, or invasion of human tumor cells in vitro (IC(50) values, 5-20 nmol/L). In addition, PF-2341066 potently inhibited HGF-stimulated endothelial cell survival or invasion and serum-stimulated tubulogenesis in vitro, suggesting that this agent also exhibits antiangiogenic properties. PF-2341066 showed efficacy at well-tolerated doses, including marked cytoreductive antitumor activity, in several tumor models that expressed activated c-Met. The antitumor efficacy of PF-2341066 was dose dependent and showed a strong correlation to inhibition of c-Met phosphorylation in vivo. Near-maximal inhibition of c-Met activity for the full dosing interval was necessary to maximize the efficacy of PF-2341066. Additional mechanism-of-action studies showed dose-dependent inhibition of c-Met-dependent signal transduction, tumor cell proliferation (Ki67), induction of apoptosis (caspase-3), and reduction of microvessel density (CD31). These results indicated that the antitumor activity of PF-2341066 may be mediated by direct effects on tumor cell growth or survival as well as antiangiogenic mechanisms. Collectively, these results show the therapeutic potential of targeting c-Met with selective small-molecule inhibitors for the treatment of human cancers.     

Selective in vivo and in vitro effects of a small molecule inhibitor of cyclin-dependent kinase 4

BACKGROUND: Cyclin-dependent kinase 4 (Cdk4) represents a prime target for the treatment of cancer because most human cancers are characterized by overexpression of its activating partner cyclin D1, loss of the natural Cdk4-specific inhibitor p16, or mutation(s) in Cdk4's catalytic subunit. All of these can cause deregulated cell growth, resulting in tumor formation. We sought to identify a small molecule that could inhibit the kinase activity of Cdk4 in vitro and to then ascertain the effects of that inhibitor on cell growth and tumor volume in vivo. Methods: A triaminopyrimidine derivative, CINK4 (a chemical inhibitor of Cdk4), was identified by screening for compounds that could inhibit Cdk4 enzyme activity in vitro. Kinase assays were performed on diverse human Cdks and on other kinases that were expressed in and purified from insect cells to determine the specificity of CINK4. Cell cycle effects of CINK4 on tumor and normal cells were studied by flow cytometry, and changes in phosphorylation of the retinoblastoma protein (pRb), a substrate of Cdk4, were determined by western blotting. The effect of the inhibitor on tumor growth in vivo was studied by use of tumors established through xenografts of HCT116 colon carcinoma cells in mice. Statistical tests were two-sided. RESULTS: CINK4 specifically inhibited Cdk4/cyclin D1 in vitro. It caused growth arrest in tumor cells and in normal cells and prevented pRb phosphorylation. CINK4 treatment resulted in statistically significantly (P: =.031) smaller mean tumor volumes in a mouse xenograft model. CONCLUSIONS: Like p16, the natural inhibitor of Cdk4, CINK4 inhibits Cdk4 activity in vitro and slows tumor growth in vivo. The specificity of CINK4 for Cdk4 raises the possibility that this small molecule or one with a similar structure could have therapeutic value.     

CEP-751 inhibits trk receptor tyrosine kinase activity in vitro and exhibits anti-tumor activity

The present report describes the in vitro and in vivo profile of CEP-751, a novel receptor tyrosine kinase inhibitor. CEP-751 at 100 nM inhibits the receptor tyrosine kinase activity of the neurotrophin receptors trkA, trkB and trkC. CEP-751 has no effect on activity of receptors for EGF, IGF-I, insulin or on erbB2; inhibition of receptors for PDGF and bFGF was observed but occurred with lesser potency than inhibition of trk. CEP-751 exhibited anti-tumor efficacy against tumors derived from NIH3T3 cells transfected with trkA. Inhibition of trk phosphorylation could also be measured in these tumors, suggesting that anti-tumor efficacy of CEP-751 is related to inhibition of trk receptor tyrosine kinase activity. CEP-751 was found to be without effect when administered to nude mice bearing SK-OV-3 tumors, which overexpress erbB2 receptors, providing further evidence that inhibition of tumor growth may be related to inhibition of trk receptor tyrosine kinase activity. Our data indicate that CEP-751 is a potent trk inhibitor which possesses anti-tumor activity. Int. J. Cancer 72:673–679, 1997. © 1997 Wiley-Liss, Inc.     

Enhancement of antitumor activity of mitomycin C in vitro and in vivo by UCN-01, a selective inhibitor of protein kinase C

UCN-01 (7-hydroxy-staurosporine) is a potent and selective inhibitor of protein kinase C (PKC), one of several protein kinases examined. UCN-01 itself was shown to exhibit antitumor activity in vitro and in vivo in oncogene-activated human and murine tumor cell lines. Since the mechanism(s) of action of UCN-01 is thought to be different from those of alkylating agents, including mitomycin C (MMC), we tested the combined effect of UCN-01 with MMC on human epidermoid carcinoma A431 cells. UCN-01 potentiated the antiproliferative activity of MMC and yet it did not affect the growth of the cells in vitro. However, other nonselective protein kinase inhibitors, such as staurosporine, K-252a, KT6124 (a derivative of K-252a) and H7, did not enhance the activity of MMC. Isobologram analysis revealed that the interaction of UCN-01 with MMC was synergistic in its antiproliferative activity. A DNA histogram of A431 cells treated with both UCN-01 and MMC showed a block in the cell cycle at the G₁/S phase. However, a histogram of cells treated with UCN-01 or MMC alone showed a G₁ or a G₂M block, respectively. The combined effect of UCN-01 with MMC was further examined in vivo in xenografted A431 cells in nude mice. The combination of both drugs in a single i.v. injection exhibited greater antitumor activity than MMC and UCN-01 alone (P<0.01). This synergistic antitumor effect was also confirmed in two other solid tumor cell lines, i.e. human xenografted colon carcinoma Co-3 and murine sarcoma 180. The same was observed in the i.v.-inoculated P388 leukemia model, in which we saw an increased lifespan of mice when UCN-01 was combined with MMC. These results suggests the feasibility of using UCN-01 in clinical oncology, especially in combination with alkylating agents such as MMC. In addition, this combination therapy might be a novel chemotherapeutic approach to MMC-insensitive tumors in clinical trials.     

In vivo antitumor activity of NVP-AEW541-A novel, potent, and selective inhibitor of the IGF-IR kinase

IGF-IR-mediated signaling promotes survival, anchorage-independent growth, and oncogenic transformation, as well as tumor growth and metastasis formation in vivo. NVP-AEW541 is a pyrrolo[2,3-d]pyrimidine derivative small molecular weight kinase inhibitor of the IGF-IR, capable of distinguishing between the IGF-IR (IC50 = 0.086 microM) and the closely related InsR (IC50 = 2.3 microM) in cells. As expected for a specific IGF-IR kinase inhibitor, NVP-AEW541 abrogates IGF-I-mediated survival and colony formation in soft agar at concentrations that are consistent with inhibition of IGF-IR autophosphorylation. In vivo, this orally bioavailable compound inhibits IGF-IR signaling in tumor xenografts and significantly reduces the growth of IGF-IR-driven fibrosarcomas. Thus, NVP-AEW541 represents a class of selective, small molecule IGF-IR kinase inhibitors with proven in vivo antitumor activity and potential therapeutic application.     

Antiangiogenic and antitumor activity of a selective PDGFR tyrosine kinase inhibitor, CP-673,451

CP-673,451 is a potent inhibitor of platelet-derived growth factor beta-receptor (PDGFR-beta) kinase- and PDGF-BB-stimulated autophosphorylation of PDGFR-beta in cells (IC(50) = 1 nmol/L) being more than 450-fold selective for PDGFR-beta versus other angiogenic receptors (e.g., vascular endothelial growth factor receptor 2, TIE-2, and fibroblast growth factor receptor 2). Multiple models have been used to evaluate in vivo activity of CP-673,451 and to understand the pharmacology of PDGFR-beta inhibition and the effect on tumor growth. These models include an ex vivo measure of PDGFR-beta phosphorylation in glioblastoma tumors, a sponge model to measure inhibition of angiogenesis, and multiple models of tumor growth inhibition. Inhibition of PDGFR-beta phosphorylation in tumors correlates with plasma and tumor levels of CP-673,451. A dose of 33 mg/kg was adequate to provide >50% inhibition of receptor for 4 hours corresponding to an EC(50) of 120 ng/mL in plasma at C(max). In a sponge angiogenesis model, CP-673,451 inhibited 70% of PDGF-BB-stimulated angiogenesis at a dose of 3 mg/kg (q.d. x 5, p.o., corresponding to 5.5 ng/mL at C(max)). The compound did not inhibit vascular endothelial growth factor- or basic fibroblast growth factor-induced angiogenesis at concentrations which inhibited tumor growth. The antitumor efficacy of CP-673,451 was evaluated in a number of human tumor xenografts grown s.c. in athymic mice, including H460 human lung carcinoma, Colo205 and LS174T human colon carcinomas, and U87MG human glioblastoma multiforme. Once-daily p.o. x 10 days dosing routinely inhibited tumor growth (ED(50) < or = 33 mg/kg). These data show that CP-673,451 is a pharmacologically selective PDGFR inhibitor, inhibits tumor PDGFR-beta phosphorylation, selectively inhibits PDGF-BB-stimulated angiogenesis in vivo, and causes significant tumor growth inhibition in multiple human xenograft models.     

A small molecule inhibitor of atypical protein kinase C signaling inhibits pancreatic cancer cell transformed growth and invasion

Pancreatic cancer is highly resistant to current chemotherapies. Identification of the critical signaling pathways that mediate pancreatic cancer transformed growth is necessary for the development of more effective therapeutic treatments. Recently, we demonstrated that protein kinase C iota (PKCι) and zeta (PKCζ) promote pancreatic cancer transformed growth and invasion, by activating Rac1→ERK and STAT3 signaling pathways, respectively. However, a key question is whether PKCι and PKCζ play redundant (or non-redundant) roles in pancreatic cancer cell transformed growth. Here we describe the novel observations that 1) PKCι and PKCζ are non-redundant in the context of the transformed growth of pancreatic cancer cells; 2) a gold-containing small molecule known to disrupt the PKCι/Par6 interaction, aurothiomalate, also disrupts PKCζ/Par6 interaction; 3) aurothiomalate inhibits downstream signaling of both PKCι and PKCζ, and blocks transformed growth of pancreatic cancer cells in vitro; and 4) aurothiomalate inhibits pancreatic cancer tumor growth and metastasis in vivo. Taken together, these data provide convincing evidence that an inhibitor of atypical PKC signaling inhibits two key oncogenic signaling pathways, driven non-redundantly by PKCι and PKCζ, to significantly reduce tumor growth and metastasis. Our results demonstrate that inhibition of atypical PKC signaling is a promising therapeutic strategy to treat pancreatic cancer.     

The selective tyrosine kinase inhibitor STI571 inhibits small cell lung cancer growth.

At least 70% of small cell lung cancers express the Kit receptor tyrosine kinase and its ligand, stem cell factor (SCF). Numerous lines of evidence have demonstrated that this coexpression constitutes a functional autocrine loop, suggesting that inhibitors of Kit tyrosine kinase activity could have therapeutic efficacy in this disease. STI571, formerly known as CGP 57148B, is a p.o. bioavailable 2-phenylaminopyrimide derivative that was designed as an Abl tyrosine kinase inhibitor, but also has efficacy against the platelet-derived growth factor receptor and Kit in vitro. Pretreatment of the H526 small cell lung cancer (SCLC) cell line with STI571 inhibited SCF-mediated Kit activation with an IC50 of 0.1 microM as measured by inhibition of receptor tyrosine phosphorylation and 0.2 microM as measured by immune complex kinase assay. This paralleled the inhibition of SCF-mediated growth by STI571, which had an IC50 of approximately 0.3 microM. Growth inhibition in SCF-containing medium was accompanied by induction of apoptosis. STI571 efficiently blocked SCF-mediated activation of mitogen-activated protein kinase and Akt, but did not affect insulin-like growth factor-1 or serum-mediated mitogen-activated protein kinase or Akt activation. Growth of five of six SCLC cell lines in medium containing 10% FCS was inhibited by STI571 with an IC50 of approximately 5 microM. Growth inhibition in serum-containing medium appeared to be cytostatic in nature because no increase in apoptosis was observed. Despite this growth inhibition, STI571 failed to enhance the cytotoxicity of either carboplatinum or etoposide when coadministered. However, taken together with the minimal toxicity that this compound has shown in preclinical studies, these data suggest that STI571 could have a role in the treatment of SCLC, possibly to block or slow recurrence after chemotherapy-induced remissions.     

D-1553: A novel KRASG12C inhibitor with potent and selective cellular and in vivo antitumor activity

D-1553 is a small molecule inhibitor selectively targeting KRAS^G12C and currently in phase II clinical trials. Here, we report the preclinical data demonstrating antitumor activity of D-1553. Potency and specificity of D-1553 in inhibiting GDP-bound KRAS^G12C mutation were determined by thermal shift assay and KRAS^G12C-coupled nucleotide exchange assay. In vitro and in vivo antitumor activity of D-1553 alone or in combination with other therapies were evaluated in KRAS^G12C mutated cancer cells and xenograft models. D-1553 showed selective and potent activity against mutated GDP-bound KRAS^G12C protein. D-1553 selectively inhibited ERK phosphorylation in NCI-H358 cells harboring KRAS^G12C mutation. Compared to the KRAS WT and KRAS^G12D cell lines, D-1553 selectively inhibited cell viability in multiple KRAS^G12C cell lines, and the potency was slightly superior to sotorasib and adagrasib. In a panel of xenograft tumor models, D-1553, given orally, showed partial or complete tumor regression. The combination of D-1553 with chemotherapy, MEK inhibitor, or SHP2 inhibitor showed stronger potency on tumor growth inhibition or regression compared to D-1553 alone. These findings support the clinical evaluation of D-1553 as an efficacious drug candidate, both as a single agent or in combination, for patients with solid tumors harboring KRAS^G12C mutation.     

BRD-glucan exhibits potent immunochemotherapeutic activity in vitro and in vivo

We carried out in vitro and in vivo assays to investigate the immunomodulatory and immunochemotherapeutic action mechanism of BRD-glucan, a high molecular weight ( approximately 3,500 kDa) polysaccharide isolated from Aureobasidium sp, and assessed the efficacy of BRD-glucan/adriamycin co-treatment of animal cancer models. RT-PCR and suspension hemolytic, plaque forming, wounding, invasion and cell proliferation assays were utilized to investigate the in vitro immunochemotherapeutic effects of BRD-glucan. In vivo, the effects of BRD-glucan and BRD-glucan/adriamycin co-treatment were tested in a B16 melanoma initiation model and in C57BL/6 mice. In vitro, BRD-glucan did not affect the cellular wounding response or invasion activity; treatment with BRD-glucan led to increase proliferation of B cells, natural killer cells and macrophages, but not T cells. In addition, we found that the BRD-glucan activation of B cells and macrophages was dependent on Toll-like receptor2 (TLR2) and TLR4, which play important roles in innate and adaptive immunity. In vivo, BRD-glucan/adriamycin co-treatment effectively reduced the number and size of metastatic colonies. Based on the results of our in vitro and in vivo toxicity, safety and immunochemotherapy assays, we propose that BRD-glucan is a promising immunochemotherapeutic anti-tumor agent.     

Characterization of the antitumor effects of the selective farnesyl protein transferase inhibitor R115777 in vivo and in vitro

R115777 [(B)-6-[amino(4-chlorophenyl)(1-methyl-1H-imidazol-5-yl)-methyl]-4-(3-chlorophenyl)-1-methyl-2(1H)-quinolinone] is a potent and selective inhibitor of farnesyl protein transferase with significant antitumor effects in vivo subsequent to oral administration in mice. In vitro, using isolated human farnesyl protein transferase, R115777 competitively inhibited the farnesylation of lamin B and K-RasB peptide substrates, with IC50s of 0.86 nM and 7.9 nM, respectively. In a panel of 53 human tumor cell lines tested for growth inhibition, approximately 75% were found to be sensitive to R115777. The majority of sensitive cell lines had a wild-type ras gene. Tumor cell lines bearing H-ras or N-ras mutations were among the most sensitive of the cell lines tested, with responses observed at nanomolar concentrations of R115777. Tumor cell lines bearing mutant K-ras genes required higher concentrations for inhibition of cell growth, with 50% of the cell lines resistant to R115777 up to concentrations of 500 nM. Inhibition of H-Ras, N-Ras, and lamin B protein processing was observed at concentrations of R115777 that inhibited cell proliferation. However, inhibition of K-RasB protein-processing could not be detected. Oral administration b.i.d. of R115777 to nude mice bearing s.c. tumors at doses ranging from 6.25-100 mg/kg inhibited the growth of tumors bearing mutant H-ras, mutant K-ras, and wild-type ras genes. Histological evaluations revealed heterogeneity in tumor responses to R115777. In LoVo human colon tumors, treatment with R115777 produced a prominent antiangiogenic response. In CAPAN-2 human pancreatic tumors, an antiproilferative response predominated, whereas in C32 human melanoma, marked induction of apoptosis was observed. The heterogeneity of histological changes associated with antitumor effects suggested that R115777, and possibly farnesyl protein transferase inhibitors as a class, alter processes of transformation related to tumor-host interactions in addition to inhibiting tumor-cell proliferation.     

In vitro and in vivo antitumor properties of the cyclin dependent kinase inhibitor CYC202 (R-roscovitine)

CDK2 inhibitors have been proposed as effective anti-cancer therapeutics. We show here that CYC202 (R-roscovitine) is a potent inhibitor of recombinant CDK2/cyclin E kinase activity (IC(50) = 0.10 microM) with an average cytotoxic IC(50) of 15.2 microM in a panel of 19 human tumour cell lines, and we also demonstrate selectivity for rapidly proliferating cells over non-proliferating cells. A study of the cell cycle effects of CYC202 in Lovo colorectal carcinoma cells showed that the major effect was not the predicted arrest in one part of the cycle, but rather an induction of cell death from all compartments of the cell cycle. The maximum tolerated dose given intravenously to mice was in excess of 20 mg/kg. Doses up to 2,000 mg/kg were tolerated when administered orally in mice. Following repeated intraperitoneal administration (3 times daily for 5 days) of 100 mg/kg to nude mice bearing the Lovo human colorectal tumour, CYC202 induced a significant antitumour effect with a 45% reduction in tumour growth compared to controls. A second experiment using the human uterine xenograft MESSA-DX5 treated with orally administered CYC202 (500 mg/kg 3 times daily for 4 days) also exhibited a significant reduction in the rate of growth of the tumour (62%). These data, showing enzyme and cellular potency together with antitumour activity, confirm the potential of CDK2 inhibitors such as CYC202 as anticancer drugs.     

Dual tyrosine kinase inhibitor for focal adhesion kinase and insulin-like growth factor-I receptor exhibits anticancer effect in esophageal adenocarcinoma in vitro and in vivo

PURPOSE: Focal adhesion kinase (FAK) regulates integrin and growth factor-mediated signaling pathways to enhance cell migration, proliferation, and survival, and its up-regulation correlates malignant grade and poor outcome in several types of cancer. In this study, we aimed to raise a potential therapeutic strategy using a FAK inhibitor for Barrett's esophageal adenocarcinoma. EXPERIMENTAL DESIGN: The expression status of FAK in clinical Barrett's esophageal adenocarcinoma tissues was determined by immunohistochemistry. Cultured esophageal adenocarcinoma cells were treated with TAE226, a specific FAK inhibitor with an additional effect of inhibiting insulin-like growth factor-I receptor (IGF-IR), to assess its anticancer effect in vitro. Western blot was carried out to explore a participating signaling pathway for TAE226-induced cell death. Furthermore, TAE226 was orally administered to s.c. xenograft animals to investigate its anticancer effect in vivo. RESULTS: Strong expression of FAK was found in 94.0% of Barrett's esophageal adenocarcinoma compared with 17.9% of Barrett's epithelia, suggesting that FAK might play a critical role in the progression of Barrett's esophageal adenocarcinoma. When esophageal adenocarcinoma cells were treated with TAE226, cell proliferation and migration were greatly inhibited with an apparent structural change of actin fiber and a loss of cell adhesion. The activities of FAK, IGF-IR, and AKT were suppressed by TAE226 and subsequent dephosphorylation of BAD at Ser(136) occurred, resulting in caspase-mediated apoptosis. In vivo tumor volume was significantly reduced by oral administration of TAE226. CONCLUSIONS: These results suggest that TAE226, a dual tyrosine kinase inhibitor for FAK and IGF-IR, could become a new remedy for Barrett's esophageal adenocarcinoma.     

The pan-erbB tyrosine kinase inhibitor CI-1033 inhibits human esophageal cancer cells in vitro and in vivo

The epidermal growth factor receptor (EGFR) is a member of the EGFR family of receptors. EGFR and other members of the EGFR family have been shown to play significant roles in human cancer cell proliferation and therefore present important molecular targets for the treatment of cancer. The purpose of this study was to examine the effect of the pan-erbB tyrosine kinase inhibitor CI-1033 against esophageal squamous cell carcinoma in vitro and in vivo. We selected 4 human esophageal squamous cell carcinoma cell lines (TT, TE2, TE6, and TE10), and determined their expression of EGFR and HER2. We examined the ability of CI-1033 to inhibit cell growth in vitro and in vivo. EGFR and HER2 were overexpressed in all 4 esophageal cancer cells. We found that CI-1033 could inhibit the growth of esophageal cancer cell lines in a dose-dependent manner with the inhibition of phosphorylation of both MAPK and AKT. The oral administration of CI-1033 exerted a significant antitumor effect on esophageal cancer tumors in athymic nude mice. Our results suggest that CI-1033 effectively inhibits the growth of esophageal squamous cell carcinoma which co-expresses both EGFR and HER2 with the inhibition of phosphorylation of both MAPK and AKT. Furthermore, in vivo animal studies of CI-1033 suggest that CI-1033 holds significant clinical potential in esophageal cancer.     

Identification of a novel recepteur d'origine nantais/c-met small-molecule kinase inhibitor with antitumor activity in vivo

Recepteur d'origine nantais (RON) is a receptor tyrosine kinase closely related to c-Met. Both receptors are involved in cell proliferation, migration, and invasion, and there is evidence that both are deregulated in cancer. Receptor overexpression has been most frequently described, but other mechanisms can lead to the oncogenic activation of RON and c-Met. They include activating mutations or gene amplification for c-Met and constitutively active splicing variants for RON. We identified a novel inhibitor of RON and c-Met, compound I, and characterized its in vitro and in vivo activities. Compound I selectively and potently inhibited the kinase activity of RON and c-Met with IC(50)s of 9 and 4 nmol/L, respectively. Compound I inhibited hepatocyte growth factor-mediated and macrophage-stimulating protein-mediated signaling and cell migration in a dose-dependent manner. Compound I was tested in vivo in xenograft models that either were dependent on c-Met or expressed a constitutively active form of RON (RONDelta160 in HT-29). Compound I caused complete tumor growth inhibition in NIH3T3 TPR-Met and U-87 MG xenografts but showed only partial inhibition in HT-29 xenografts. The effect of compound I in HT-29 xenografts is consistent with the expression of the activating b-Raf V600E mutation, which activates the mitogen-activated protein kinase pathway downstream of RON. Importantly, tumor growth inhibition correlated with the inhibition of c-Met-dependent and RON-dependent signaling in tumors. Taken together, our results suggest that a small-molecule dual inhibitor of RON/c-Met has the potential to inhibit tumor growth and could therefore be useful for the treatment of patients with cancers where RON and/or c-Met are activated.