RESUMEN
Chromosomal instability (CIN) is the persistent reshuffling of cancer karyotypes via chromosome mis-segregation during cell division. In cancer, CIN exists at varying levels that have differential effects on tumor progression. However, mis-segregation rates remain challenging to assess in human cancer despite an array of available measures. We evaluated measures of CIN by comparing quantitative methods using specific, inducible phenotypic CIN models of chromosome bridges, pseudobipolar spindles, multipolar spindles, and polar chromosomes. For each, we measured CIN fixed and timelapse fluorescence microscopy, chromosome spreads, six-centromere FISH, bulk transcriptomics, and single-cell DNA sequencing (scDNAseq). As expected, microscopy of tumor cells in live and fixed samples significantly correlated (R = 0.72; P < 0.001) and sensitively detect CIN. Cytogenetics approaches include chromosome spreads and 6-centromere FISH, which also significantly correlate (R = 0.76; P < 0.001) but had limited sensitivity for lower rates of CIN. Bulk genomic DNA signatures and bulk transcriptomic scores, CIN70 and HET70, did not detect CIN. By contrast, scDNAseq detects CIN with high sensitivity, and significantly correlates with imaging methods (R = 0.82; P < 0.001). In summary, single-cell methods such as imaging, cytogenetics, and scDNAseq can measure CIN, with the latter being the most comprehensive method accessible to clinical samples. To facilitate the comparison of CIN rates between phenotypes and methods, we propose a standardized unit of CIN: Mis-segregations per Diploid Division. This systematic analysis of common CIN measures highlights the superiority of single-cell methods and provides guidance for measuring CIN in the clinical setting.
Asunto(s)
Inestabilidad Cromosómica , Neoplasias , Humanos , Línea Celular Tumoral , Inestabilidad Cromosómica/genética , Centrómero , Cariotipificación , Perfilación de la Expresión Génica , Segregación Cromosómica , AneuploidiaRESUMEN
Gene fusions involving EWSR1 or FUS as the 5' partner have been reported in a diverse array of sarcomas. Here, we characterize the histopathology and genomics of six tumors harboring a gene fusion between EWSR1 or FUS and POU2AF3, an understudied, putative colorectal cancer predisposition gene. Striking morphologic features reminiscent of synovial sarcoma were observed including a biphasic appearance with variable fusiform to epithelioid cytomorphology and staghorn-type vasculature. RNA sequencing demonstrated variable breakpoints in EWSR1/FUS along with similar breakpoints in POU2AF3 that encompassed a 3' portion of this gene. For cases in which additional information was available, the behavior of these neoplasms was aggressive with local spread and/or distant metastases. Although further studies are needed to confirm the functional significance of our findings, POU2AF3 fusions to EWSR1 or FUS may define a novel type of POU2AF3-rearranged sarcomas with aggressive, malignant behavior.
Asunto(s)
Sarcoma Sinovial , Sarcoma , Neoplasias de los Tejidos Blandos , Humanos , Proteína EWS de Unión a ARN/genética , Sarcoma/genética , Neoplasias de los Tejidos Blandos/genética , Fusión Génica , Hibridación Fluorescente in Situ , Biomarcadores de Tumor/genética , Proteínas de Fusión Oncogénica/genética , Proteínas de Neoplasias/genética , Proteína FUS de Unión a ARN/genéticaRESUMEN
The ability to distinguish endometrial serous carcinoma (SC) from high-grade endometrioid adenocarcinoma is of great importance given their differences in prognosis and management. In practice, this distinction typically relies upon the use of a focused immunohistochemical panel including p53, p16, and mismatch repair proteins. The expression of p16 is characteristically strong and diffuse in SCs, and weak and/or patchy in many high-grade endometrioid adenocarcinomas. Here, we report a subset of SCs that are entirely negative for p16 immunostaining, a pattern we refer to as "p16 null." This pattern was identified in 2 of 63 cases of SC diagnosed at our institution-1 with histologically classic features and 1 with ambiguous high-grade histologic features. These tumors otherwise showed a SC signature by immunohistochemical and demonstrated an SC pattern of genetic mutations. No mutation in the gene for p16, cyclin-dependent kinase inhibitor 2A (CDKN2A), was identified in either case. However, molecular correlates for the absent p16 expression were present, including homozygous deletion of CDKN2A in one case and hemizygous deletion of CDKN2A with promotor hypermethylation of the remaining allele in the other case. To our knowledge, this constitutes the first report conclusively demonstrating the existence of a small subset of SCs that are completely negative by p16 immunohistochemistry, and the molecular lesions responsible for this pattern. In the context of an otherwise clinically and histologically classic example of SC, we endorse this "null" p16 staining pattern as an alternative aberrant staining pattern that should not deter one from committing to this diagnosis.
Asunto(s)
Carcinoma Endometrioide , Cistadenocarcinoma Seroso , Neoplasias Endometriales , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Carcinoma Endometrioide/diagnóstico , Carcinoma Endometrioide/genética , Carcinoma Endometrioide/metabolismo , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Cistadenocarcinoma Seroso/genética , Neoplasias Endometriales/diagnóstico , Neoplasias Endometriales/genética , Neoplasias Endometriales/metabolismo , Femenino , Homocigoto , Humanos , Eliminación de Secuencia , Coloración y EtiquetadoRESUMEN
BACKGROUND: Trastuzumab has shown a survival benefit in cases of Her2-positive gastroesophageal cancer (GEC). Immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) currently determine eligibility for trastuzumab-based therapy. However, these low-throughput assays often produce discordant or equivocal results. METHODS: We developed a targeted proteomic assay based on selected reaction monitoring mass spectrometry (SRM-MS) and quantified levels (amol/µg) of Her2-SRM protein in cell lines (n = 27) and GEC tissues (n = 139). We compared Her2-SRM protein expression with IHC/FISH, seeking to determine optimal SRM protein expression cutoffs in order to identify HER2 gene amplification. RESULTS: After demonstrating assay development, precision, and stability, Her2-SRM protein measurement was observed to be highly concordant with the HER2/CEP17 ratio, particularly in a multivariate regression model adjusted for SRM expression of the covariates Met, Egfr, Her3, and HER2 heterogeneity, as well as their interactions (cell lines r (2) = 0.9842; FFPE r (2) = 0.7643). In GEC tissues, Her2-SRM protein was detected at any level in 71.2 % of cases. ROC curves demonstrated that Her2-SRM protein levels have a high specificity (100 %) at an upper-level cutoff of >750 amol/µg and sensitivity of 75 % at a lower-level cutoff of <450 amol/µg for identifying HER2 FISH-amplified tumors. An "equivocal zone" of 450-750 amol/µg of Her2-SRM protein was analogous to IHC2+ but represented fewer cases (9-16 % of cases versus 36-41 %). CONCLUSIONS: Compared to IHC, targeted SRM-Her2 proteomics provided more objective and quantitative Her2 expression with excellent HER2/CEP17 FISH correlation and fewer equivocal cases. Along with its multiplex capability for other relevant oncoproteins, these results demonstrate a refined HER2 protein expression assay for clinical application.
Asunto(s)
Hibridación Fluorescente in Situ/métodos , Proteómica/métodos , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Amplificación de Genes , Humanos , Técnicas para Inmunoenzimas , Neoplasias Gástricas/patologíaRESUMEN
Chromosomal instability (CIN) is the persistent reshuffling of cancer karyotypes via chromosome mis-segregation during cell division. In cancer, CIN exists at varying levels that have differential effects on tumor progression. However, mis-segregation rates remain challenging to assess in human cancer despite an array of available measures. We evaluated measures of CIN by comparing quantitative methods using specific, inducible phenotypic CIN models of chromosome bridges, pseudobipolar spindles, multipolar spindles, and polar chromosomes. For each, we measured CIN fixed and timelapse fluorescence microscopy, chromosome spreads, 6-centromere FISH, bulk transcriptomics, and single cell DNA sequencing (scDNAseq). As expected, microscopy of tumor cells in live and fixed samples correlated well (R=0.77; p<0.01) and sensitively detect CIN. Cytogenetics approaches include chromosome spreads and 6-centromere FISH, which also correlate well (R=0.77; p<0.01) but had limited sensitivity for lower rates of CIN. Bulk genomic DNA signatures and bulk transcriptomic scores, CIN70 and HET70, did not detect CIN. By contrast, single-cell DNA sequencing (scDNAseq) detects CIN with high sensitivity, and correlates very well with imaging methods (R=0.83; p<0.01). In summary, single-cell methods such as imaging, cytogenetics, and scDNAseq can measure CIN, with the latter being the most comprehensive method accessible to clinical samples. To facilitate comparison of CIN rates between phenotypes and methods, we propose a standardized unit of CIN: Mis-segregations per Diploid Division (MDD). This systematic analysis of common CIN measures highlights the superiority of single-cell methods and provides guidance for measuring CIN in the clinical setting.
RESUMEN
Variation in genes underlying host immunity can lead to marked differences in susceptibility to HIV infection among humans. Despite heavy reliance on non-human primates as models for HIV/AIDS, little is known about which host factors are shared and which are unique to a given primate lineage. Here, we investigate whether copy number variation (CNV) at CCL3-like genes (CCL3L), a key genetic host factor for HIV/AIDS susceptibility and cell-mediated immune response in humans, is also a determinant of time until onset of simian-AIDS in rhesus macaques. Using a retrospective study of 57 rhesus macaques experimentally infected with SIVmac, we find that CCL3L CNV explains approximately 18% of the variance in time to simian-AIDS (p<0.001) with lower CCL3L copy number associating with more rapid disease course. We also find that CCL3L copy number varies significantly (p<10(-6)) among rhesus subpopulations, with Indian-origin macaques having, on average, half as many CCL3L gene copies as Chinese-origin macaques. Lastly, we confirm that CCL3L shows variable copy number in humans and chimpanzees and report on CCL3L CNV within and among three additional primate species. On the basis of our findings we suggest that (1) the difference in population level copy number may explain previously reported observations of longer post-infection survivorship of Chinese-origin rhesus macaques, (2) stratification by CCL3L copy number in rhesus SIV vaccine trials will increase power and reduce noise due to non-vaccine-related differences in survival, and (3) CCL3L CNV is an ancestral component of the primate immune response and, therefore, copy number variation has not been driven by HIV or SIV per se.
Asunto(s)
Quimiocina CCL3/genética , Dosificación de Gen , Síndrome de Inmunodeficiencia Adquirida del Simio/genética , Síndrome de Inmunodeficiencia Adquirida del Simio/fisiopatología , Animales , Calibración , Cartilla de ADN/química , Progresión de la Enfermedad , Genética de Población , Sistema Inmunológico , Funciones de Verosimilitud , Macaca mulatta , Repeticiones de Microsatélite , Modelos Estadísticos , Modelos de Riesgos Proporcionales , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
In the Supplementary Information originally published with this article, a lane was missing in the ß-actin blot in Supplementary Fig. 2. The lane has been added. The error has been corrected in the Supplementary Information associated with this article.
RESUMEN
The role of KRAS, when activated through canonical mutations, has been well established in cancer1. Here we explore a secondary means of KRAS activation in cancer: focal high-level amplification of the KRAS gene in the absence of coding mutations. These amplifications occur most commonly in esophageal, gastric and ovarian adenocarcinomas2-4. KRAS-amplified gastric cancer models show marked overexpression of the KRAS protein and are insensitive to MAPK blockade owing to their capacity to adaptively respond by rapidly increasing KRAS-GTP levels. Here we demonstrate that inhibition of the guanine-exchange factors SOS1 and SOS2 or the protein tyrosine phosphatase SHP2 can attenuate this adaptive process and that targeting these factors, both genetically and pharmacologically, can enhance the sensitivity of KRAS-amplified models to MEK inhibition in both in vitro and in vivo settings. These data demonstrate the relevance of copy-number amplification as a mechanism of KRAS activation, and uncover the therapeutic potential for targeting of these tumors through combined SHP2 and MEK inhibition.
Asunto(s)
Neoplasias Esofágicas/genética , Amplificación de Genes , Quinasas de Proteína Quinasa Activadas por Mitógenos/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 11/antagonistas & inhibidores , Proteínas Proto-Oncogénicas p21(ras)/genética , Neoplasias Gástricas/genética , Animales , Línea Celular Tumoral , Modelos Animales de Enfermedad , Neoplasias Esofágicas/patología , Humanos , Ratones , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Piperidinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Proteína Tirosina Fosfatasa no Receptora Tipo 11/metabolismo , Piridonas/farmacología , Pirimidinas/farmacología , Pirimidinonas/farmacología , Neoplasias Gástricas/patologíaRESUMEN
PURPOSE: Sonic hedgehog (SHH), an activating ligand of smoothened (SMO), is overexpressed in > 70% of pancreatic cancers (PCs). We investigated the impact of vismodegib, an SHH antagonist, plus gemcitabine (GV) or gemcitabine plus placebo (GP) in a multicenter phase Ib/randomized phase II trial and preclinical PC models. PATIENTS AND METHODS: Patients with PC not amenable to curative therapy who had received no prior therapy for metastatic disease and had Karnofsky performance score ≥ 80 were enrolled. Patients were randomly assigned in a one-to-one ratio to GV or GP. The primary end point was progression-free-survival (PFS). Exploratory correlative studies included serial SHH serum levels and contrast perfusion computed tomography imaging. To further investigate putative biologic mechanisms of SMO inhibition, two autochthonous pancreatic cancer models (Kras(G12D); p16/p19(fl/fl); Pdx1-Cre and Kras(G12D); p53(R270H/wt); Pdx1-Cre) were studied. RESULTS: No safety issues were identified in the phase Ib portion (n = 7), and the phase II study enrolled 106 evaluable patients (n = 53 in each arm). Median PFS was 4.0 and 2.5 months for GV and GP arms, respectively (95% CI, 2.5 to 5.3 and 1.9 to 3.8, respectively; adjusted hazard ratio, 0.81; 95% CI, 0.54 to 1.21; P = .30). Median overall survival (OS) was 6.9 and 6.1 months for GV and GP arms, respectively (95% CI, 5.8 to 8.0 and 5.0 to 8.0, respectively; adjusted hazard ratio, 1.04; 95% CI, 0.69 to 1.58; P = .84). Response rates were not significantly different. There were no significant associations between correlative markers and overall response rate, PFS, or OS. Preclinical trials revealed no significant differences with vismodegib in drug delivery, tumor growth rate, or OS in either model. CONCLUSION: The addition of vismodegib to gemcitabine in an unselected cohort did not improve overall response rate, PFS, or OS in patients with metastatic PC. Our preclinical and clinical results revealed no statistically significant differences with respect to drug delivery or treatment efficacy using vismodegib.
Asunto(s)
Anilidas/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Proteínas Hedgehog/antagonistas & inhibidores , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/patología , Piridinas/uso terapéutico , Transducción de Señal/efectos de los fármacos , Adulto , Anciano , Anilidas/farmacología , Animales , Desoxicitidina/administración & dosificación , Desoxicitidina/análogos & derivados , Supervivencia sin Enfermedad , Femenino , Proteínas Hedgehog/metabolismo , Humanos , Estimación de Kaplan-Meier , Estado de Ejecución de Karnofsky , Masculino , Ratones , Persona de Mediana Edad , Estadificación de Neoplasias , Oportunidad Relativa , Neoplasias Pancreáticas/metabolismo , Piridinas/farmacología , Resultado del Tratamiento , GemcitabinaRESUMEN
BACKGROUND: Overexpression of Met tyrosine kinase receptor is associated with poor prognosis. Overexpression, and particularly MET amplification, are predictive of response to Met-specific therapy in preclinical models. Immunohistochemistry (IHC) of formalin-fixed paraffin-embedded (FFPE) tissues is currently used to select for 'high Met' expressing tumors for Met inhibitor trials. IHC suffers from antibody non-specificity, lack of quantitative resolution, and, when quantifying multiple proteins, inefficient use of scarce tissue. METHODS: After describing the development of the Liquid-Tissue-Selected Reaction Monitoring-mass spectrometry (LT-SRM-MS) Met assay, we evaluated the expression level of Met in 130 FFPE gastroesophageal cancer (GEC) tissues. We assessed the correlation of SRM Met expression to IHC and mean MET gene copy number (GCN)/nucleus or MET/CEP7 ratio by fluorescence in situ hybridization (FISH). RESULTS: Proteomic mapping of recombinant Met identified 418TEFTTALQR426 as the optimal SRM peptide. Limits of detection (LOD) and quantitation (LOQ) for this peptide were 150 and 200 amol/µg tumor protein, respectively. The assay demonstrated excellent precision and temporal stability of measurements in serial sections analyzed one year apart. Expression levels of 130 GEC tissues ranged (<150 amol/µg to 4669.5 amol/µg. High correlation was observed between SRM Met expression and both MET GCN and MET/CEP7 ratio as determined by FISH (nâ=â30; R2â=â0.898). IHC did not correlate well with SRM (nâ=â44; R2â=â0.537) nor FISH GCN (nâ=â31; R2â=â0.509). A Met SRM level of ≥1500 amol/µg was 100% sensitive (95% CI 0.69-1) and 100% specific (95% CI 0.92-1) for MET amplification. CONCLUSIONS: The Met SRM assay measured the absolute Met levels in clinical tissues with high precision. Compared to IHC, SRM provided a quantitative and linear measurement of Met expression, reliably distinguishing between non-amplified and amplified MET tumors. These results demonstrate a novel clinical tool for efficient tumor expression profiling, potentially leading to better informed therapeutic decisions for patients with GEC.
Asunto(s)
Neoplasias Esofágicas , Amplificación de Genes , Espectrometría de Masas/métodos , Proteínas Proto-Oncogénicas c-met , Neoplasias Gástricas , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/patología , Femenino , Humanos , Inmunohistoquímica/métodos , Masculino , Proteínas Proto-Oncogénicas c-met/genética , Proteínas Proto-Oncogénicas c-met/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologíaRESUMEN
BACKGROUND: Paxillin is a modular protein that localises to cell adhesion sites where it facilitates bidirectional communication between the intracellular actin cytoskeleton and the extracellular matrix. These complex and dynamic interactions are essential for cell adhesion, cell migration and cell survival. The authors have previously demonstrated that paxillin is overexpressed in lung cancer tissues and identified somatic paxillin mutations in 9% of lung cancers. A murine in vivo xenograft model of the most common paxillin mutation (A127T) showed increased cell proliferation and invasive tumour growth, establishing an important role for paxillin in the development of lung cancer. METHODS: The authors analysed 279 bronchoscopy-aided biopsy specimens from 92 high-risk patients. Adenocarcinoma with bronchioloalveolar features and pure bronchioloalveolar carcinoma (BAC) were analysed with fluorescence in situ hybridisation (FISH) and immunohistochemistry (IHC). RESULTS: Paxillin is overexpressed in premalignant areas of hyperplasia, squamous metaplasia and goblet cell metaplasia, as well as dysplastic lesions and carcinoma in high-risk patients. Concordance between increased paxillin gene copy number and paxillin overexpression was observed in cases of adenocarcinoma eusomic for chromosome 12. CONCLUSIONS: Paxillin overexpression occurs during the earliest stages of lung cancer development. FISH and IHC analysis of lung adenocarcinoma suggests that relatively small-scale genomic rearrangements of chromosome 12 are associated with paxillin overexpression in lung adenocarcinoma.
Asunto(s)
Adenocarcinoma Bronquioloalveolar/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas de Neoplasias/metabolismo , Paxillin/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patología , Adenocarcinoma/secundario , Adenocarcinoma del Pulmón , Adenocarcinoma Bronquioloalveolar/genética , Adenocarcinoma Bronquioloalveolar/patología , Adenocarcinoma Bronquioloalveolar/secundario , Anciano , Biopsia , Cromosomas Humanos Par 12/genética , Femenino , Dosificación de Gen , Genes erbB-1/genética , Humanos , Hibridación Fluorescente in Situ , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/secundario , Masculino , Persona de Mediana Edad , Mutación , Proteínas de Neoplasias/genética , Paxillin/genética , Lesiones Precancerosas/genética , Lesiones Precancerosas/metabolismo , Lesiones Precancerosas/patología , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas p21(ras) , Proteínas ras/genéticaRESUMEN
UNLABELLED: A 48 year-old female with chemo-refractory metastatic gastric cancer to the liver was treated on a Phase I clinical trial with MetMAb, a monoclonal antibody targeting the Met tyrosine kinase receptor. The primary tumor had high MET gene polysomy and evidence for an autocrine production of HGF, the growth factor ligand of Met. A complete response was obtained lasting two years; the cancer recurred as a peritoneal deposit invading into the transverse colon and a gastrohepatic ligament node. Compassionate use of MetMAb therapy at recurrence achieved a mixed response--a partial response of the two initial lesions, but with development of multiple new foci of carcinomatosis. Tissue and serum studies evaluating the Met signaling pathway did correlate with MetMAb treatment response initially and at the time of recurrence. SIGNIFICANCE: This research brief is the first to describe a durable complete response obtained with a molecularly targeted monoclonal antibody, MetMAb, to the receptor tyrosine kinase, Met, in a patient with chemorefractory metastatic gastric cancer. It is also the first to report biomarkers that predicted therapeutic response to Met inhibition.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Recurrencia Local de Neoplasia/terapia , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Neoplasias Gástricas/terapia , Anticuerpos Monoclonales/inmunología , Biomarcadores de Tumor/metabolismo , Ensayos Clínicos Fase I como Asunto , Ensayos Clínicos Fase II como Asunto , Resistencia a Medicamentos , Femenino , Factor de Crecimiento de Hepatocito/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/secundario , Neoplasias Hepáticas/terapia , Persona de Mediana Edad , Metástasis de la Neoplasia , Recurrencia Local de Neoplasia/metabolismo , Proteínas Proto-Oncogénicas c-met/inmunología , Proteínas Proto-Oncogénicas c-met/metabolismo , Ensayos Clínicos Controlados Aleatorios como Asunto , Transducción de Señal/efectos de los fármacos , Neoplasias Gástricas/patologíaRESUMEN
RON (MST1R) is one of two members of the MET receptor tyrosine kinase family, along with parent receptor MET. RON has a putative role in several cancers, but its expression and function is poorly characterized in gastroesophageal adenocarcinoma. A recognized functional role of MET tyrosine kinase in gastroesophageal cancer has led to early phase clinical trials using MET inhibitors, with unimpressive results. Therefore, the role of RON in gastroesophageal cancer, as well as its role in cooperative signaling with MET and as a mechanism of resistance to MET inhibition, was studied in gastroesophageal tissues and cell lines. By IHC, RON was highly over-expressed in 74% of gastroesophageal samples (n=94), and over-expression was prognostic of poor survival (p=0.008); RON and MET co-expression occurred in 43% of samples and was prognostic of worst survival (p=0.03). High MST1R gene copy number by quantitative polymerase chain reaction, and confirmed by fluorescence in situ hybridization and/or array comparative genomic hybridization, was seen in 35.5% (16/45) of cases. High MST1R gene copy number correlated with poor survival (p=0.01), and was associated with high MET and ERBB2 gene copy number. A novel somatic MST1R juxtamembrane mutation R1018G was found in 11% of samples. RON signaling was functional in cell lines, activating downstream effector STAT3, and resulted in increased viability over controls. RON and MET co-stimulation assays led to enhanced malignant phenotypes over stimulation of either receptor alone. Growth inhibition as evidenced by viability and apoptosis assays was optimal using novel blocking monoclonal antibodies to both RON and MET, versus either alone. SU11274, a classic MET small molecule tyrosine kinase inhibitor, blocked signaling of both receptors, and proved synergistic when combined with STAT3 inhibition (combination index < 1). These preclinical studies define RON as an important novel prognostic marker and therapeutic target for gastroesophageal cancer warranting further investigation.