Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer

Conor Bradley, Philip Dunne, Stephen McQuaid, Victoria Bingham, Mark Lawler, Manuel Salto-Tellez, Patrick Johnston, Sandra Van Schaeybroeck

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

Background: The c-MET proto-oncogene is frequently overexpressed (50-60%), amplified (1-3%), and mutated (1-3%) in colorectal cancer (CRC). Hepatocyte growth factor (HGF)-dependent and independent activation of c-MET has been associated with increased survival and resistance to targeted therapies. This study aimed to investigate the role of the HGF/c-MET axis in regulating migration/invasion in CRC, using pre-clinical models and clinical samples. Methods: In order to model CRC tumour cell invasion, we have generated invasive CRC subpopulations using Boyden Invasion chambers. To model the CRC microenvironment, we have used a range of co-culture techniques with CRC cells and colon fibroblasts. Migration/invasion was determined using xCELLigence System (Roche). c-MET expression in parental and invasive cell lines was measured using Western blotting and qRT-PCR. c-MET expression in CRC FFPE tissues was measured using IHC and RNAScope®. Results: We identified marked upregulated expression of c-MET at both the protein and transcript levels in our invasive CRC cell line models. Importantly, both parental and invasive subpopulations were found to be inherently dependent on c-MET for migration, as RNAi against c-MET abrogated migration/invasion in both parental and invasive models. We also demonstrated that stimulation of CRC cells with rh-HGF resulted in increased CRC cell migration/invasion. In addition, co-culture of CRC cells with colonic myofibroblasts, resulted in marked increases in migratory and invasive capacity, and this was dependent on HGF/c-MET signaling. Interestingly, stimulation with myofibroblast conditioned medium or HGF promotes rapid degradation of c-MET at the protein level, followed by recycling, while MET transcript remains unaltered, illustrating a dynamic expression of c-MET protein in response to activation. We further showed that MET is transcriptionally upregulated in tumour budding foci at the invasive front of a cohort of stage III CRC tumors. Intriguingly, c-MET protein levels do not correlate with the transcript, most likely due to a similar protein degradation process observed in our aforementioned in vitro models. Conclusions: We show for the first time a key role for transcriptional upregulation of MET as a molecular driver of tumour invasion, both in vitro and in stage III CRC tumours.
Original languageEnglish
Title of host publicationUpregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer
PublisherProceedings: AACR Annual Meeting 2016; April 16-20 New Orleans
Editionabstract 924
Publication statusPublished - 2016

Fingerprint

Colorectal Neoplasms
Up-Regulation
Neoplasms
Hepatocyte Growth Factor
Myofibroblasts
Coculture Techniques
Proteins
Cell Line
Culture Techniques
Tumor Microenvironment
Proto-Oncogenes
Recycling
Conditioned Culture Medium
RNA Interference
Colonic Neoplasms
Proteolysis
Cell Movement
Fibroblasts
Western Blotting

Cite this

Bradley, C., Dunne, P., McQuaid, S., Bingham, V., Lawler, M., Salto-Tellez, M., ... Van Schaeybroeck, S. (2016). Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. In Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer (abstract 924 ed.). Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans.
Bradley, Conor ; Dunne, Philip ; McQuaid, Stephen ; Bingham, Victoria ; Lawler, Mark ; Salto-Tellez, Manuel ; Johnston, Patrick ; Van Schaeybroeck, Sandra. / Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. abstract 924. ed. Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans, 2016.
@inproceedings{063bfd8fb91e49818f65f91edf1b098b,
title = "Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer",
abstract = "Background: The c-MET proto-oncogene is frequently overexpressed (50-60{\%}), amplified (1-3{\%}), and mutated (1-3{\%}) in colorectal cancer (CRC). Hepatocyte growth factor (HGF)-dependent and independent activation of c-MET has been associated with increased survival and resistance to targeted therapies. This study aimed to investigate the role of the HGF/c-MET axis in regulating migration/invasion in CRC, using pre-clinical models and clinical samples. Methods: In order to model CRC tumour cell invasion, we have generated invasive CRC subpopulations using Boyden Invasion chambers. To model the CRC microenvironment, we have used a range of co-culture techniques with CRC cells and colon fibroblasts. Migration/invasion was determined using xCELLigence System (Roche). c-MET expression in parental and invasive cell lines was measured using Western blotting and qRT-PCR. c-MET expression in CRC FFPE tissues was measured using IHC and RNAScope{\circledR}. Results: We identified marked upregulated expression of c-MET at both the protein and transcript levels in our invasive CRC cell line models. Importantly, both parental and invasive subpopulations were found to be inherently dependent on c-MET for migration, as RNAi against c-MET abrogated migration/invasion in both parental and invasive models. We also demonstrated that stimulation of CRC cells with rh-HGF resulted in increased CRC cell migration/invasion. In addition, co-culture of CRC cells with colonic myofibroblasts, resulted in marked increases in migratory and invasive capacity, and this was dependent on HGF/c-MET signaling. Interestingly, stimulation with myofibroblast conditioned medium or HGF promotes rapid degradation of c-MET at the protein level, followed by recycling, while MET transcript remains unaltered, illustrating a dynamic expression of c-MET protein in response to activation. We further showed that MET is transcriptionally upregulated in tumour budding foci at the invasive front of a cohort of stage III CRC tumors. Intriguingly, c-MET protein levels do not correlate with the transcript, most likely due to a similar protein degradation process observed in our aforementioned in vitro models. Conclusions: We show for the first time a key role for transcriptional upregulation of MET as a molecular driver of tumour invasion, both in vitro and in stage III CRC tumours.",
author = "Conor Bradley and Philip Dunne and Stephen McQuaid and Victoria Bingham and Mark Lawler and Manuel Salto-Tellez and Patrick Johnston and {Van Schaeybroeck}, Sandra",
year = "2016",
language = "English",
booktitle = "Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer",
publisher = "Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans",
edition = "abstract 924",

}

Bradley, C, Dunne, P, McQuaid, S, Bingham, V, Lawler, M, Salto-Tellez, M, Johnston, P & Van Schaeybroeck, S 2016, Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. in Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. abstract 924 edn, Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans.

Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. / Bradley, Conor; Dunne, Philip; McQuaid, Stephen; Bingham, Victoria; Lawler, Mark; Salto-Tellez, Manuel; Johnston, Patrick; Van Schaeybroeck, Sandra.

Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. abstract 924. ed. Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans, 2016.

Research output: Chapter in Book/Report/Conference proceedingConference contribution

TY - GEN

T1 - Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer

AU - Bradley, Conor

AU - Dunne, Philip

AU - McQuaid, Stephen

AU - Bingham, Victoria

AU - Lawler, Mark

AU - Salto-Tellez, Manuel

AU - Johnston, Patrick

AU - Van Schaeybroeck, Sandra

PY - 2016

Y1 - 2016

N2 - Background: The c-MET proto-oncogene is frequently overexpressed (50-60%), amplified (1-3%), and mutated (1-3%) in colorectal cancer (CRC). Hepatocyte growth factor (HGF)-dependent and independent activation of c-MET has been associated with increased survival and resistance to targeted therapies. This study aimed to investigate the role of the HGF/c-MET axis in regulating migration/invasion in CRC, using pre-clinical models and clinical samples. Methods: In order to model CRC tumour cell invasion, we have generated invasive CRC subpopulations using Boyden Invasion chambers. To model the CRC microenvironment, we have used a range of co-culture techniques with CRC cells and colon fibroblasts. Migration/invasion was determined using xCELLigence System (Roche). c-MET expression in parental and invasive cell lines was measured using Western blotting and qRT-PCR. c-MET expression in CRC FFPE tissues was measured using IHC and RNAScope®. Results: We identified marked upregulated expression of c-MET at both the protein and transcript levels in our invasive CRC cell line models. Importantly, both parental and invasive subpopulations were found to be inherently dependent on c-MET for migration, as RNAi against c-MET abrogated migration/invasion in both parental and invasive models. We also demonstrated that stimulation of CRC cells with rh-HGF resulted in increased CRC cell migration/invasion. In addition, co-culture of CRC cells with colonic myofibroblasts, resulted in marked increases in migratory and invasive capacity, and this was dependent on HGF/c-MET signaling. Interestingly, stimulation with myofibroblast conditioned medium or HGF promotes rapid degradation of c-MET at the protein level, followed by recycling, while MET transcript remains unaltered, illustrating a dynamic expression of c-MET protein in response to activation. We further showed that MET is transcriptionally upregulated in tumour budding foci at the invasive front of a cohort of stage III CRC tumors. Intriguingly, c-MET protein levels do not correlate with the transcript, most likely due to a similar protein degradation process observed in our aforementioned in vitro models. Conclusions: We show for the first time a key role for transcriptional upregulation of MET as a molecular driver of tumour invasion, both in vitro and in stage III CRC tumours.

AB - Background: The c-MET proto-oncogene is frequently overexpressed (50-60%), amplified (1-3%), and mutated (1-3%) in colorectal cancer (CRC). Hepatocyte growth factor (HGF)-dependent and independent activation of c-MET has been associated with increased survival and resistance to targeted therapies. This study aimed to investigate the role of the HGF/c-MET axis in regulating migration/invasion in CRC, using pre-clinical models and clinical samples. Methods: In order to model CRC tumour cell invasion, we have generated invasive CRC subpopulations using Boyden Invasion chambers. To model the CRC microenvironment, we have used a range of co-culture techniques with CRC cells and colon fibroblasts. Migration/invasion was determined using xCELLigence System (Roche). c-MET expression in parental and invasive cell lines was measured using Western blotting and qRT-PCR. c-MET expression in CRC FFPE tissues was measured using IHC and RNAScope®. Results: We identified marked upregulated expression of c-MET at both the protein and transcript levels in our invasive CRC cell line models. Importantly, both parental and invasive subpopulations were found to be inherently dependent on c-MET for migration, as RNAi against c-MET abrogated migration/invasion in both parental and invasive models. We also demonstrated that stimulation of CRC cells with rh-HGF resulted in increased CRC cell migration/invasion. In addition, co-culture of CRC cells with colonic myofibroblasts, resulted in marked increases in migratory and invasive capacity, and this was dependent on HGF/c-MET signaling. Interestingly, stimulation with myofibroblast conditioned medium or HGF promotes rapid degradation of c-MET at the protein level, followed by recycling, while MET transcript remains unaltered, illustrating a dynamic expression of c-MET protein in response to activation. We further showed that MET is transcriptionally upregulated in tumour budding foci at the invasive front of a cohort of stage III CRC tumors. Intriguingly, c-MET protein levels do not correlate with the transcript, most likely due to a similar protein degradation process observed in our aforementioned in vitro models. Conclusions: We show for the first time a key role for transcriptional upregulation of MET as a molecular driver of tumour invasion, both in vitro and in stage III CRC tumours.

UR - http://www.abstractsonline.com/Plan/ViewAbstract.aspx?mID=4017&sKey=60767c34-b158-4c83-93ca-003da4a7211e&cKey=720ea5f8-a4aa-4874-911d-907715f1e6b5&mKey=1d10d749-4b6a-4ab3-bcd4-f80fb1922267

M3 - Conference contribution

BT - Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer

PB - Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans

ER -

Bradley C, Dunne P, McQuaid S, Bingham V, Lawler M, Salto-Tellez M et al. Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. In Upregulation of the MET transcript is consistently associated with invasion and tumor budding in colorectal cancer. abstract 924 ed. Proceedings: AACR Annual Meeting 2016; April 16-20 New Orleans. 2016