c-Met Activation Research Articles

A viral microRNA downregulates metastasis suppressor CD82 and induces cell invasion and angiogenesis by activating the c-Met signaling.

Kaposi’s sarcoma (KS) is the most common AIDS-associated malignancy etiologically caused by Kaposi’s sarcoma-associated herpesvirus (KSHV). KS is a highly disseminated and vascularized tumor comprised of poorly differentiated spindle-shaped endothelial cells. KSHV encodes 12 pre-microRNAs (pre-miRNAs) that yield 25 mature miRNAs, but their roles in KSHV-induced tumor dissemination and angiogenesis remain largely unknown. KSHV-encoded miR-K12-6 (miR-K6) can produce two mature miRNAs, miR-K6-3p and miR-K6-5p. Recently, we have shown that miR-K6-3p promoted cell migration and angiogenesis by directly targeting SH3 domain binding glutamate-rich protein (SH3BGR) (PLoS Pathog. 2016;12(4):e1005605). Here, by using mass spectrometry, bioinformatics analysis and luciferase reporter assay, we showed that miR-K6-5p directly targeted the coding sequence (CDS) of CD82 molecule (CD82), a metastasis suppressor. Ectopic expression of miR-K6-5p specifically inhibited the expression of endogenous CD82 and strongly promoted endothelial cells invasion in vitro and angiogenesis in vivo. Overexpression of CD82 significantly inhibited cell invasion and angiogenesis induced by miR-K6-5p. Mechanistically, CD82 directly interacted with c-Met to inhibit its activation. MiR-K6-5p directly repressed CD82, relieving its inhibition on c-Met activation and inducing cell invasion and angiogenesis. Deletion of miR-K6 from KSHV genome abrogated KSHV suppression of CD82 resulting in compromised KSHV activation of c-Met pathway, and KSHV-induced invasion and angiogenesis. In conclusion, these results show that by inhibiting CD82, KSHV miR-K6-5p promotes cell invasion and angiogenesis by activating the c-Met pathway. Our findings illustrate that KSHV miRNAs may play an essential role in the dissemination and angiogenesis of KSHV-induced malignancies.

Phase I study of the anti-HGF monoclonal antibody (mAb), ficlatuzumab, and cetuximab in cetuximab-resistant, recurrent/metastatic (R/M) head and neck squamous cell carcinoma (HNSCC).

6038 Background: Cetuximab, an anti-EGFR mAb, is approved for R/M HNSCC but only a minority benefits. Activation of cMet, the receptor for hepatocyte growth factor (HGF), overcomes EGFR inhibition in preclinical models and high serum HGF is associated with resistance in patients (pts). We conducted a phase I trial evaluating the combination of cetuximab and ficlatuzumab, an IgG1 anti-HGF mAb, in pts with cetuximab-resistant, R/M HNSCC. Methods: In this Narayana k-in-a-row phase I design, cetuximab 500 mg/m2 was administered every 2 weeks. Ficlatuzumab dose tiers were 10 mg/kg (tier 1) or 20 mg/kg IV every 2 weeks (tier 2), with inter-patient escalation or de-escalation based on cumulative dose-limiting toxicities (DLT). The recommended phase II dose (RP2D) was set at tier 2 if no DLTs were observed after 8 enrolled pts; expansion continued to n = 12. Key eligibility criteria: R/M HNSCC; recurrence within 6 months of cetuximab-radiation or progression during/within 6 months of palliative cetuximab; ECOG PS 0-1. Candidate biomarkers included serum Veristrat, a proteomic classifier in lung cancer where “good” predicts benefit from anti-EGFR therapy and “poor” indicates resistance and poor prognosis. Results: From Sept 2015–June 2016, 12 pts were enrolled and treated. Primary site: 1 oral cavity; 3 oropharynx (1 p16+); 2 hypopharynx; 5 larynx; 1 external auditory canal. Platinum-refractory: 11/12. Veristrat: 8 poor; 4 good. Three pts were treated at tier 1 and 9 at tier 2. No DLTs were observed. Grade 3 adverse events included: edema (1), hypoalbuminemia (1), infection (2), thromboembolic event (2). Median PFS and OS at RP2D were 6.0 mos (90% CI = 2.0 mos–not reached) and 8.2 mos (90% CI = 2.7 mos–not reached), respectively. Response rate was 17% (90% CI = 0–28%): 2/12 partial response (PR); 1/3 at 10 mg/kg; 1/9 at 20mg/kg. Clinical benefit rate (PR + stable disease) was 67%. Veristrat was not associated with PFS. Conclusions: The RP2D is cetuximab 500 mg/m2 and ficlatuzumab 20 mg/kg every 2 weeks. This well-tolerated combination demonstrated promising activity in pts with poor prognosis, cetuximab-resistant R/M HNSCC. Phase II testing is justified. Clinical trial information: NCT02277197.

Comparison of the c-MET gene amplification between primary tumor and metastatic lymph nodes in non-small cell lung cancer.

e23138 Background: Activation of c-MET lead to a wide range of biological activities. MET has recently been identified as a novel promising target in NSCLC, some c-MET inhibitors have been developed. The primary aim of this study was to evaluate the c-MET amplification status in advanced NSCLC and compare the consistency of c-MET amplification analyses in metastatic lymph nodes and tumor tissue.Methods: Real-time fluorescent quantitative polymerase chain reaction was used to test the tumor tissues in 368 patients of NSCLCs and 178 paired metastatic lymph nodes samples and compared the amplifications consistency inmetastatic lymph nodesand tissue samples, and analyzed the correlation between c-MET gene amplification and clinical characteristics of patients. Another 5 cases of normal lung tissue were taken as negative control.Results: The c-MET gene amplification rate was 8.97%(33/368) in tumor tissues . Of the178 paired cases, c-MET gene amplification was positive in 7.95%(15/178) cancerous tissuesand18.54%(33/178) in metastatic lymph nodes; Of these patients,13 were positive in the two kinds of samples. 20 cases in the paired group were detected positive c-MET in metastatic lymph nodes but was negative in cancerous tissues; 2 cases in the paired group were detected positive c-MET gene amplification in cancerous tissues but was negative in metastatic lymph nodes,there were statistically significant differences between the two samples (χ2= 45.536, P < 0.001). c-MET gene amplification was detected more in metastatic lymph nodes than the primary cancerous tissue. Consistency was evaluated by consistency test(Kappa = 0.482, P < 0.001).When lymph nodes were used as surrogate samples of primary cancerous tissues, sensitivity was86.67%, the specificity was 87.69%.Conclusions: More c-MET gene amplification positive were detected in metastatic lymph nodes compare with the primary cancerous tissue. c-MET gene amplification detected in lymph node metastases could screen more patients suitable for TKI therapy. Lymph node Metastasis can predict the c-MET gene amplification of primary tumor, and guide the clinical use of gene targeted drugs.

Phase Ib study of tepotinib in EGFR-mutant/c-Met-positive NSCLC: Final data and long-term responders.

8547 Background: Patients (pts) with NSCLC that is initially responsive to EGFR tyrosine kinase inhibitors (EGFR TKI) typically develop resistance, often associated with aberrant c-Met activity. Dual inhibition of EGFR and c-Met is therefore a rational option to treat c-Met+ EGFR TKI-resistant NSCLC. Tepotinib is a highly selective c-Met inhibitor with good tolerability and promising activity against solid tumors. We report final data from a phase Ib trial of tepotinib + gefitinib in pts with c-Met+/EGFR-mutant NSCLC conducted in Asia. Methods: Eligible pts were adults with locally advanced/metastatic NSCLC and ECOG PS 0–1. Tumors had to express EGFR with an activating mutation be resistant to EGFR TKI therapy and be c-Met+ by IHC. Pts received tepotinib 300 or 500 mg QD combined with gefitinib 250 mg QD (T300G250 or T500G250). The primary objective was to determine the recommended phase II dose (RP2D) of tepotinib in combination with gefitinib; secondary objectives included pharmacokinetics (PK), safety, and antitumor activity. Results: 18 pts were enrolled (median age 65 [41–78]; 8 male); 6 received T300G250, 12 T500G250. No dose-limiting toxicities were observed, and tepotinib 500 mg QD was confirmed as the RP2D. 17 pts experienced treatment-related treatment-emergent adverse events (TRTEAEs), mostly grade ≤2 and most commonly diarrhea (12), rash (8), and amylase increase (6). Grade ≥3 TRTEAEs were increased amylase (n = 4), increased lipase (3), neutropenia (1), and hyperglycemia (1). The best overall response was partial response in 6 pts; 4/7 pts with IHC 3+ tumors responded (all treated with T500G250) vs 2/11 with IHC 2+ tumors. Response durations of pts with PR were 4.2–12.5 months. 4/18 pts (IHC 2+, n = 3) had stable disease. 8 pts experienced progression free survival > 5 months, 3 pts > 10 months. PK were as expected from previous studies. Conclusions: Tepotinib in combination with gefitinib was well tolerated. The RP2D of tepotinib for use in combination with gefitinib in NSCLC is 500 mg QD. T500G250 showed signs of activity against c-Met+ tumors. A phase II trial is randomizing ≈156 pts with c-Met+/T790M– tumors who have failed first-line EFGR TKI 2:1 to tepotinib + gefitinib or pemetrexed + cisplatin/carboplatin. Clinical trial information: NCT02864992.

Clinical impact of high serum hepatocyte growth factor in advanced non-small cell lung cancer.

Activation of c-MET through hepatocyte growth factor (HGF) increases tumorigenesis, induces resistance, and is associated with poor prognosis in various solid tumors. However, the clinical value of serum HGF (sHGF) in patients with advanced non-small cell lung cancer (NSCLC), especially those receiving cytotoxic chemotherapy, remains unknown. Here, we show that sHGF may be useful to predict tumor response and progression-free survival (PFS) in patients with advanced NSCLC. A total of 81 patients with NSCLC were investigated. sHGF levels were evaluated using ELISA at 4 time-points: at pre-treatment, at response-evaluation (1–2 months after treatment initiation), at the best tumor response, and at disease progression. As a control biomarker, CEA was also evaluated in lung adenocarcinoma. Positive-sHGF at response-evaluation predicted poor PFS compared with Negative-sHGF in both first-line (median, 153.5 vs. 288.0; P < 0.05) and second-line treatment (87.0 vs. 219.5; P = 0.01). In 55 patients that received cytotoxic chemotherapy, multiple Cox proportional hazards models showed significant independent associations between poor PFS and Positive-sHGF at response-evaluation (hazard ratio, 4.24; 95% CI, 2.05 to 9.46; P < 0.01). Lung adenocarcinoma subgroup analysis showed that in patients receiving second cytotoxic chemotherapy, there were no significant differences in PFS between patients with low-CEA compared with those with high-CEA, but Positive-sHGF at pre-treatment or at response-evaluation predicted poor PFS (35.0 vs. 132.0; P < 0.01, 50.0 vs. 215.0; P < 0.01, respectively). These findings give a rationale for future research investigating the merit of sHGF as a potential clinical biomarker to evaluate HGF/c-MET activity, which would be useful to indicate administration of c-MET inhibitors.

Development of an everolimus-resistant BON1 cell line with altered cell cycle and c-Met activity

Development of an everolimus-resistant BON1 cell line with altered cell cycle and c-Met activity

Migration of oral squamous cell carcinoma cells are induced by HGF/c-Met signalling via lamellipodia and filopodia formation.

The activation of receptor tyrosine kinases (RTKs) results in cellular effects including cell proliferation, survival, migration and invasion; RTKs also play an important role in tumourigenesis. It has been reported that EGFR signalling controls the migration of oral squamous cell carcinoma (OSCC) SAS and HSC3 cells but not of HSC4 cells, although the proliferation of HSC4 cells is regulated by EGF/EGFR. In the present study, we investigated the roles of EGFR and the c-Met signalling pathway in cell migration via filopodia and lamellipodia formation, which may be prerequisites for migration. To explore the role of c-Met in cell migration, we inhibited c-Met RTK activity using the c-Met inhibitor SU11274 and activated c-Met using hepatocyte growth factor (HGF) in three OSCC cell lines HSC4, SAS and Ca9-22 and investigated migration potency using a wound healing assay. We showed that inhibition of c-Met significantly suppressed, and activation of c-Met significantly promoted, the migration of OSCC cells. Additionally, the migration of SAS and Ca9-22 cells was inhibited by the EGFR inhibitors AG1478 and cetuximab and promoted by EGF treatment. Moreover, migration potency was correlated with lamellipodia formation. Furthermore, western blot analyses demonstrated that SU11274 decreased and HGF increased lamellipodin protein levels as well as phosphorylated c-Met levels. Collectively, we demonstrated that c-Met signalling induced lamellipodia formation by upregulating lamellipodin, thereby promoting the migration of OSCC cells.

Natural variants of Listeria monocytogenes internalin B with different ability to stimulate cell proliferation and cytoskeleton rearrangement in HEp-2 cells

The Gram-positive bacterium Listeria monocytogenes is a facultative intracellular pathogen. The InlB protein is required for L. monocytogenes active invasion into epithelial cells. InlB interacts with the surface tyrosine kinase receptor c-Met. Activation of c-Met by its natural ligand, hepatocyte growth factor, induces intracellular signaling pathways and causes cell proliferation and/or migration. Interactions of InlB with c-Met in the course of infection activates the receptor and results in cytoskeleton rearrangement and bacterial uptake. In this study, we investigated the potential of the purified InlB protein as the c-Met ligand causing cell proliferation. Three distinct InlB variants have been compared. These variants were previously described in L. monocytogenes strains isolated from different sources. The variants were shown to be different in the induction of HEp-2 cell proliferation. In addition, the InlB variants differed in their potential to induce cytoskeleton rearrangements.

MP39-05 TRANSPOSON MUTAGENESIS DRIVES RENAL CYST FORMATION IN VIVO WHEN COMBINED WITH C-MET HYPERACTIVATION: IMPLICATIONS FOR ACQUIRED RENAL CYSTIC DISEASE

You have accessJournal of UrologyKidney Cancer: Basic Research & Pathophysiology I1 Apr 2017MP39-05 TRANSPOSON MUTAGENESIS DRIVES RENAL CYST FORMATION IN VIVO WHEN COMBINED WITH C-MET HYPERACTIVATION: IMPLICATIONS FOR ACQUIRED RENAL CYSTIC DISEASE Jason Scovell, Juan Hernandez, Adam Hollander, and Richard Link Jason ScovellJason Scovell More articles by this author , Juan HernandezJuan Hernandez More articles by this author , Adam HollanderAdam Hollander More articles by this author , and Richard LinkRichard Link More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.1179AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Acquired renal cystic disease (ARCD) imparts a high risk for renal cell carcinoma (8%) in patients with end-stage renal disease. The molecular mechanism of cyst formation in ARCD remains unknown, although increased hepatocyte growth factor / C-MET signaling have been implicated in cyst formation. To explore molecular mechanisms of renal cyst development relevant to ARCD, we developed a murine model system based on tissue-selectable C-Met activation and transposon-mediated mutagenesis. METHODS To allow conditional activation of C-Met signaling, we engineered a tandem duplicated and mutated human C-MET coding sequence. Cre recombinase catalyzes exchange of the wild type C-Met locus with a constitutively active variant (M1248T). C-met+/M1248T mice were crossed with mice carrying the mutagenic Sleeping Beauty (Onc2) transposable element activated by a cre-dependent transposase (trpase). Localization to the renal epithelium was achieved by ggt-cre. Mice with activated C-Met and transposon mutagenesis (1: c-met+/M1248T; Onc2+/-, trpase+/-; ggt-cre+/-) were compared to mice with activated C-Met (2: c-met+/M1248T; ggt-cre+/-) or activated transposon mutagenesis (3: Onc2+/-, trpase+/-; ggt-cre+/-) alone. Mice were serial imaged by ultrasound and MRI. All kidneys were then examined grossly and histologically at necropsy after aging. RESULTS All mice (N=5) with both C-Met hyperactivation and transposon activity (1) developed large renal cysts by 6 months. No mice with only C-Met hyper-activation (2; N=10) or transposon activity (3; N=10) developed cysts by 6 months. Histological analysis demonstrated fluid filled and hemorrhagic renal cysts without evidence for malignancy. CONCLUSIONS C-Met hyperactivation is insufficient to drive renal cyst formation in isolation. Combining M1248T with transposon mutagenesis drove renal cyst formation with 100% penetrance. By mapping transposon insertion sites in cyst-lining epithelial cells in this model, we can now begin to isolate these interacting genes, which may improve our understanding of the molecular mechanisms underlying ARCD. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e494-e495 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Jason Scovell More articles by this author Juan Hernandez More articles by this author Adam Hollander More articles by this author Richard Link More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

P08.47 Crosstalk of the HGF/c-MET and TGF-β pathways in glioblastoma

AbstractMultiple target inhibition has gained considerable interest in combating drug resistance in glioblastoma, however, understanding the molecular mechanisms of crosstalk between signaling pathways and predicting responses of cancer cells to targeted interventions has remained challenging. Despite the significant role attributed to transforming growth factor (TGF)-β family and hepatocyte growth factor (HGF)/c-MET signaling in glioblastoma pathogenesis, their functional interactions have not been well characterized. Using genetic and pharmacological approaches to stimulate or antagonize the TGF-β-pathway in human glioma-initiating cells (GIC), we observed that TGF-β exerts an inhibitory effect on c-MET phosphorylation through the activation of non-canonical, SMAD-independent pathways, including mitogen-activated protein kinase (MAPK)/ extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB / AKT) pathways. Moreover, a comparison of c-MET-driven and c-MET independent GIC models revealed that TGF-β inhibits stemness in GIC at least in part via its negative regulation of c-MET activity, suggesting that stem cell maintenance may be controlled by the balance between these two oncogenic pathways. Importantly, immunohistochemical analyses of TGF-β and p-c-MET in human glioblastoma specimens supports a concept of negative regulation between these pathways. These novel insights into the crosstalk of two major pathogenic pathways in glioblastoma may explain some of the disappointing results when targeting either pathway alone in human glioblastoma patients and inform on potential future designs on targeted pharmacological or genetic intervention.

Preclinical development of a humanized neutralizing antibody targeting HGF

Hepatocyte growth factor (HGF) and its receptor, cMET, play critical roles in cell proliferation, angiogenesis and invasion in a wide variety of cancers. We therefore examined the anti-tumor activity of the humanized monoclonal anti-HGF antibody, YYB-101, in nude mice bearing human glioblastoma xenografts as a single agent or in combination with temozolomide. HGF neutralization, The extracellular signal-related kinases 1 and 2 (ERK1/2) phosphorylation, and HGF-induced scattering were assessed in HGF-expressing cell lines treated with YYB-101. To support clinical development, we also evaluated the preclinical pharmacokinetics and toxicokinetics in cynomolgus monkeys, and human and cynomolgus monkey tissue was stained with YYB-101 to test tissue cross-reactivity. We found that YYB-101 inhibited cMET activation in vitro and suppressed tumor growth in the orthotopic mouse model of human glioblastoma. Combination treatment with YYB-101 and temozolomide decreased tumor growth and increased overall survival compared with the effects of either agent alone. Five cancer-related genes (TMEM119, FST, RSPO3, ROS1 and NBL1) were overexpressed in YYB-101-treated mice that showed tumor regrowth. In the tissue cross-reactivity assay, critical cross-reactivity was not observed. The terminal elimination half-life was 21.7 days. Taken together, the in vitro and in vivo data demonstrated the anti-tumor efficacy of YYB-101, which appeared to be mediated by blocking the HGF/cMET interaction. The preclinical pharmacokinetics, toxicokinetics and tissue cross-reactivity data support the clinical development of YYB-101 for advanced cancer.

Molecular mechanisms of activating c-MET in KSHV+ primary effusion lymphoma.

The oncogenic Kaposi's sarcoma–associated herpesvirus (KSHV) is a principal causative agent of primary effusion lymphoma (PEL), which is mostly seen in immunosuppressed patients. PEL is a rapidly progressing malignancy with a median survival time of approximately 6 months even under the conventional chemotherapy. We recently report that the hepatocyte growth factor (HGF)/c-MET pathway is highly activated in PEL cells and represents a promising therapeutic target (Blood. 2015;126(26):2821-31). However, the underlying mechanisms of c-MET activation within PEL cells remain largely unknown. To solve this puzzle, here we have utilized the next generation sequencing (NGS) based bioinformatics approach to investigate the genomic landscape of the c-MET gene and we found that there's no single nucleotide variations (SNVs) occurred in the c-MET genomic regions in a cohort of PEL samples. Consistently, Sanger sequencing analysis of frequently mutated exons such as exon 10, 14 and 19 shows no mutation of these c-MET exons in PEL cell-lines, which further supports the notion that mutations are not the major mechanism responsible for c-MET activation in PEL. Further, we found that a transmembrane receptor protein, Plexin-B1, is expressed in PEL cell-lines, which is required for c-MET-mediated PEL cell survival via direct protein interaction.

PAK4 interacts with p85 alpha: implications for pancreatic cancer cell migration

It has been reported that p21-activated kinase 4 (PAK4) is amplified in pancreatic cancer tissue. PAK4 is a member of the PAK family of serine/threonine kinases, which act as effectors for several small GTPases, and has been specifically identified to function downstream of HGF-mediated c-Met activation in a PI3K dependent manner. However, the functionality of PAK4 in pancreatic cancer and the contribution made by HGF signalling to pancreatic cancer cell motility remain to be elucidated. We now find that elevated PAK4 expression is coincident with increased expression levels of c-Met and the p85α subunit of PI3K. Furthermore, we demonstrate that pancreatic cancer cells have a specific motility response to HGF both in 2D and 3D physiomimetic organotypic assays; which can be suppressed by inhibition of PI3K. Significantly, we report a specific interaction between PAK4 and p85α and find that PAK4 deficient cells exhibit a reduction in Akt phosphorylation downstream of HGF signalling. These results implicate a novel role for PAK4 within the PI3K pathway via interaction with p85α. Thus, PAK4 could be an essential player in PDAC progression representing an interesting therapeutic opportunity.

ABBV-399, a c-Met Antibody-Drug Conjugate that Targets Both MET-Amplified and c-Met-Overexpressing Tumors, Irrespective of MET Pathway Dependence.

Purpose: Despite the importance of the MET oncogene in many malignancies, clinical strategies targeting c-Met have benefitted only small subsets of patients with tumors driven by signaling through the c-Met pathway, thereby necessitating selection of patients with MET amplification and/or c-Met activation most likely to respond. An ADC targeting c-Met could overcome these limitations with potential as a broad-acting therapeutic.Experimental Design: ADC ABBV-399 was generated with the c-Met-targeting antibody, ABT-700. Antitumor activity was evaluated in cancer cells with overexpressed c-Met or amplified MET and in xenografts including patient-derived xenograft (PDX) models and those refractory to other c-Met inhibitors. The correlation between c-Met expression and sensitivity to ABBV-399 in tumor and normal cell lines was assessed to evaluate the risk of on-target toxicity.Results: A threshold level of c-Met expressed by sensitive tumor but not normal cells is required for significant ABBV-399-mediated killing of tumor cells. Activity extends to c-Met or amplified MET cell line and PDX models where significant tumor growth inhibition and regressions are observed. ABBV-399 inhibits growth of xenograft tumors refractory to other c-Met inhibitors and provides significant therapeutic benefit in combination with standard-of-care chemotherapy.Conclusions: ABBV-399 represents a novel therapeutic strategy to deliver a potent cytotoxin to c-Met-overexpressing tumor cells enabling cell killing regardless of reliance on MET signaling. ABBV-399 has progressed to a phase I study where it has been well tolerated and has produced objective responses in c-Met-expressing non-small cell lung cancer (NSCLC) patients. Clin Cancer Res; 23(4); 992-1000. ©2016 AACR.

C-Met/miR-130b axis as novel mechanism and biomarker for castration resistance state acquisition.

Although a significant subset of prostate tumors remain indolent during the entire life, the advanced forms are still one of the leading cause of cancer-related death. There are not reliable markers distinguishing indolent from aggressive forms. Here we highlighted a new molecular circuitry involving microRNA and coding genes promoting cancer progression and castration resistance. Our preclinical and clinical data demonstrated that c-Met activation increases miR-130b levels, inhibits androgen receptor expression, promotes cancer spreading and resistance to hormone ablation therapy. The relevance of these findings was confirmed on patients' samples and by in silico analysis on an independent patient cohort from Taylor's platform. Data suggest c-Met/miR-130b axis as a new prognostic marker for patients' risk assessment and as an indicator of therapy resistance. Our results propose new biomarkers for therapy decision-making in all phases of the pathology. Data may help identify high-risk patients to be treated with adjuvant therapy together with alternative cure for castration-resistant forms while facilitating the identification of possible patients candidates for anti-Met therapy. In addition, we demonstrated that it is possible to evaluate Met/miR-130b axis expression in exosomes isolated from peripheral blood of surgery candidates and advanced patients offering a new non-invasive tool for active surveillance and therapy monitoring.

Phase II trial of cabozantinib in patients with carcinoid and pancreatic neuroendocrine tumors (pNET).

228 Background: Activation of VEGFR2 and c-MET has been implicated in driving growth of neuroendocrine tumors (NET); additionally, expression of c-MET has been associated with shorter survival. Cabozantinib inhibits VEGFR2 and c-MET. We performed a two-cohort phase II study to evaluate the efficacy of cabozantinib in patients (pts) with advanced carcinoid or pNET (NCT01466036). Methods: Pts with progressive, well differentiated, grade 1-2 carcinoid or pNET were enrolled in parallel cohorts and treated with cabozantinib 60 mg po qd . There was no limit to prior therapy. Pts were restaged every 2 mos for the first 6 mos, then every 3 mos. The primary endpoint was objective response rate as measured by RECIST 1.1. Enrollment of approximately 35 patients of each tumor type was planned. Results: 41 pts with carcinoid (median age 63 yrs, 44% male, % ECOG PS 0/1 = 51/49) were accrued. Accrual to the pNET cohort was halted due to investigator/sponsor decision after 20 pts (median age 55 yrs, 60% male, % ECOG PS 0/1 = 40/60). Carcinoid pts completed a median of 8 (range 0-44) 28-day treatment cycles; pNET pts completed a median of 10 (0-35) cycles. 14 pts remain on treatment. Reasons for discontinuation were progression or death (51%), withdrawal of consent or investigator decision (28%), adverse events (AE, 21%). 3/20 pts with pNET achieved PR (ORR 15%, 95% CI 5-36%); 15/20 had SD. 6/41 pts with carcinoid achieved PR (ORR 15%, 95% CI 7-28%); 26/41 had SD . Median PFS was 21.8 mo (95% CI, 8.5-32.0 mo) in pts with pNET and 31.4 mo (95% CI, 8.5 mo-NR) in pts with carcinoid. Gr 3/4 toxicity in ≥ 1 pt included hypertension (13%), hypophosphatemia (11%), diarrhea (10%), lymphopenia (7%), thrombocytopenia (5%), fatigue (5%), increased lipase or amylase (8%). Unexpected Gr 3/4 AEs included heart failure and autoimmune hemolytic anemia, each in 1 pt. 81% of 53 pts completing more than 1 cycle of treatment required dose reduction from the initial 60 mg dose. Conclusions: Treatment with cabozantinib was associated with objective tumor responses and encouraging PFS durations in patients with advanced carcinoid and advanced pNET. While dose reduction was common, treatment was tolerable. Further evaluation of cabozantinib is warranted in both NET subgroups. Clinical trial information: NCT01466036.

PHA665752 inhibits the HGF-stimulated migration and invasion of cells by blocking PI3K/AKT pathway in uveal melanoma.

HGF/c-MET is frequently associated with tumor metastasis in many cancers, including uveal melanoma (UM). PHA665752, a selective c-MET inhibitor, exhibits anticancer activity through inhibiting cell motility in some cancers. In this study, we investigated the effects of PHA665752 on UM cell lines M17 and SP6.5. Our data show that HGF stimulated the motility of UM cells, and induced the activation of both c-MET and PI3K/AKT, but not ERK1/2. Moreover, consistent with the amount of c-MET within the nucleus, PHA665752 significantly inhibited HGF-promoted cell motility and suppressed the phosphorylation of c-MET and PI3K/AKT, but not ERK1/2 induced by HGF. Additionally, the effects of PHA665752 on both the inhibition of HGF-induced cell motility and the suppression of active AKT are similar to those of PI3K inhibitor LY294002. In xenograft models, PHA665752 significantly inhibited tumor growth in nude mice and similarly suppressed the phosphorylation of c-MET and PI3K/AKT. Our current findings, combined with previous results, demonstrate that PHA665752 inhibits HGF-induced motility via the inhibition of PI3K/AKT. This study suggests that targeting HGF/c-MET could be a promising therapeutic strategy for UM by preventing cell motility.

Pharmacophore Mapping Approach for Drug Target Identification: A Chemical Synthesis and in Silico Study on Novel Thiadiazole Compounds.

Compounds containing thiadiazole moiety are cognized to possess with variety of clinical and therapeutic activity. Finding a suitable drug target for newly synthesized compounds remain a major bottle neck in current high throughout medicinal chemistry era. To effectively synthesize di substituted thiadiazole compounds and demonstrate drug target identification using an in silico pharmacophore probing approach. Moreover, we also aim to validate the suitability of identified drug target. A cost-effective and environmental friendly chemical synthesis scheme for production of di substituted thiadiazole compounds was employed. Target identification was conducted by Pharmmapper software. Validation was accomplished by performing molecular docking and further Molecular Hydrophobic Potential (MHP) analysis. Pharmacophore probing base approach identified hepatocyte growth factor receptor (c-Met) as a suitable biological target for newly synthesized compounds. Binding free energy values indicate that compound 4b, 4e, 4g and 4h has tremendous potential to be further used as lead compound to design selective inhibitors of c-Met receptor. MHP data from current study supports the possibility that hydrophobic contacts might act as major factor stabilizing thiadiazole- c-Met complex. Moreover, in silico observations of current study are in absolute accordance with previously described in vitro and crystallographic analysis. We demonstrate that thiadiazole compounds synthesized in current investigation has high potential to act in modulation of hepatocyte growth factor receptor (c-Met) activity and thereby act as putative therapeutic agent in cancer therapy.

The regulatory role of heparin on c-Met signaling in hepatocellular carcinoma cells.

The role of heparin as an anticoagulant is well defined; however, its role in tumorigenesis and tumor progression is not clear yet. Some studies have shown that anticoagulant treatment in cancer patients improve overall survival, however, recent clinical trials have not shown a survival benefit in cancer patients receiving heparin treatment. In our previous studies we have shown the inhibitory effects of heparin on Hepatocyte Growth Factor (HGF)-induced invasion and migration in hepatocellular carcinoma (HCC) cells. In this study, we showed the differential effects of heparin on the behaviors of HCC cells based on the presence or absence of HGF. In the absence of HGF, heparin activated HGF/c-Met signaling and promoted motility and invasion in HCC cells. Heparin treatment led to c-Met receptor dimerization and activated c-Met signaling in an HGF independent manner. Heparin-induced c-Met activation increased migration and invasion through ERK1/2, early growth response factor 1 (EGR1) and Matrix Metalloproteinases (MMP) axis. Interestingly, heparin modestly decreased the proliferation of HCC cells by inhibiting activatory phosphorylation of Akt. The inhibition of c-Met signaling reversed heparin-induced increase in motility and invasion and, proliferation inhibition. Our study provides a new perspective into the role of heparin on c-Met signaling in HCC.

Functional and therapeutic relevance of hepatocyte growth factor/c‐MET signaling in synovial sarcoma

Synovial sarcoma (SS) is an aggressive soft tissue sarcoma with a poor prognosis and, thus, novel therapeutic strategies for SS are urgently required. In the present study, we investigated the functional and therapeutic relevance of hepatocyte growth factor (HGF)/c‐MET signaling in SS. Both HGF and c‐MET were highly expressed in Yamato‐SS cells, resulting in activation of c‐MET and its downstream AKT and extracellular signal‐regulated kinase signaling pathways, whereas c‐MET was expressed but not activated in SYO‐1 or HS‐SY‐II cells. c‐MET‐activated Yamato‐SS cells showed higher anchorage‐independent growth ability and less sensitivity to chemotherapeutic agents than did c‐MET‐inactivated SYO‐1 or HS‐SY‐II cells. INC280, a selective c‐MET inhibitor, inhibited growth of Yamato‐SS cells both in vitro and in vivo but not that of SYO‐1 or HS‐SY‐II cells. INC280 induced cell cycle arrest and apoptosis, and blocked phosphorylation of c‐MET and its downstream effectors in Yamato‐SS cells. Co‐expression of HGF and c‐MET in SS clinical samples correlated with a poor prognosis in patients with SS. Taken together, activation of HGF/c‐MET signaling in an autocrine fashion leads to an aggressive phenotype in SS and targeting of this signaling exerts superior antitumor effects on c‐MET‐activated SS. HGF/c‐MET expression status is a potential biomarker for identification of SS patients with a worse prognosis who can benefit from c‐MET inhibitors.