Role of c-MET inhibitors in overcoming drug resistance in spheroid models of primary human pancreatic cancer and stellate cells

Omidreza Firuzi, Pei Pei Che, Btissame el Hassouni, Mark Buijs, Stefano Coppola, Matthias Löhr, Niccola Funel, Rainer Heuchel, Ilaria Carnevale, Thomas Schmidt, Giulia Mantini, Amir Avan, Luciano Saso, Godefridus J. Peters, Elisa Giovannetti

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Pancreatic stellate cells (PSCs) are a key component of tumor microenvironment in pancreatic ductal adenocarcinoma (PDAC) and contribute to drug resistance. c-MET receptor tyrosine kinase activation plays an important role in tumorigenesis in different cancers including PDAC. In this study, effects of PSC conditioned medium (PCM) on c-MET phosphorylation (by immunocytochemistry enzyme-linked immunosorbent assay (ELISA)) and drug response (by sulforhodamine B assay) were investigated in five primary PDAC cells. In novel 3D-spheroid co-cultures of cyan fluorescence protein (CFP)-firefly luciferase (Fluc)-expressing primary human PDAC cells and green fluorescence protein (GFP)-expressing immortalized PSCs, PDAC cell growth and chemosensitivity were examined by luciferase assay, while spheroids’ architecture was evaluated by confocal microscopy. The highest phospho-c-MET expression was detected in PDAC5 and its subclone sorted for “stage specific embryonic antigen-4" (PDAC5 (SSEA4)). PCM of cells pre-incubated with PDAC conditioned medium, containing increased hepatocyte growth factor (HGF) levels, made PDAC cells significantly more resistant to gemcitabine, but not to c-MET inhibitors. Hetero-spheroids containing both PSCs and PDAC5 (SSEA4) cells were more resistant to gemcitabine compared to PDAC5 (SSEA4) homo-spheroids. However, c-MET inhibitors (tivantinib, PHA-665752 and crizotinib) were equally effective in both spheroid models. Experiments with primary human PSCs confirmed the main findings. In conclusion, we developed spheroid models to evaluate PSC-PDAC reciprocal interaction, unraveling c-MET inhibition as an important therapeutic option against drug resistant PDAC.
Original languageEnglish
Article number638
JournalCancers
Volume11
Issue number5
DOIs
Publication statusPublished - 2019

Cite this

Firuzi, Omidreza ; Che, Pei Pei ; el Hassouni, Btissame ; Buijs, Mark ; Coppola, Stefano ; Löhr, Matthias ; Funel, Niccola ; Heuchel, Rainer ; Carnevale, Ilaria ; Schmidt, Thomas ; Mantini, Giulia ; Avan, Amir ; Saso, Luciano ; Peters, Godefridus J. ; Giovannetti, Elisa. / Role of c-MET inhibitors in overcoming drug resistance in spheroid models of primary human pancreatic cancer and stellate cells. In: Cancers. 2019 ; Vol. 11, No. 5.
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title = "Role of c-MET inhibitors in overcoming drug resistance in spheroid models of primary human pancreatic cancer and stellate cells",
abstract = "Pancreatic stellate cells (PSCs) are a key component of tumor microenvironment in pancreatic ductal adenocarcinoma (PDAC) and contribute to drug resistance. c-MET receptor tyrosine kinase activation plays an important role in tumorigenesis in different cancers including PDAC. In this study, effects of PSC conditioned medium (PCM) on c-MET phosphorylation (by immunocytochemistry enzyme-linked immunosorbent assay (ELISA)) and drug response (by sulforhodamine B assay) were investigated in five primary PDAC cells. In novel 3D-spheroid co-cultures of cyan fluorescence protein (CFP)-firefly luciferase (Fluc)-expressing primary human PDAC cells and green fluorescence protein (GFP)-expressing immortalized PSCs, PDAC cell growth and chemosensitivity were examined by luciferase assay, while spheroids’ architecture was evaluated by confocal microscopy. The highest phospho-c-MET expression was detected in PDAC5 and its subclone sorted for “stage specific embryonic antigen-4{"} (PDAC5 (SSEA4)). PCM of cells pre-incubated with PDAC conditioned medium, containing increased hepatocyte growth factor (HGF) levels, made PDAC cells significantly more resistant to gemcitabine, but not to c-MET inhibitors. Hetero-spheroids containing both PSCs and PDAC5 (SSEA4) cells were more resistant to gemcitabine compared to PDAC5 (SSEA4) homo-spheroids. However, c-MET inhibitors (tivantinib, PHA-665752 and crizotinib) were equally effective in both spheroid models. Experiments with primary human PSCs confirmed the main findings. In conclusion, we developed spheroid models to evaluate PSC-PDAC reciprocal interaction, unraveling c-MET inhibition as an important therapeutic option against drug resistant PDAC.",
author = "Omidreza Firuzi and Che, {Pei Pei} and {el Hassouni}, Btissame and Mark Buijs and Stefano Coppola and Matthias L{\"o}hr and Niccola Funel and Rainer Heuchel and Ilaria Carnevale and Thomas Schmidt and Giulia Mantini and Amir Avan and Luciano Saso and Peters, {Godefridus J.} and Elisa Giovannetti",
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Role of c-MET inhibitors in overcoming drug resistance in spheroid models of primary human pancreatic cancer and stellate cells. / Firuzi, Omidreza; Che, Pei Pei; el Hassouni, Btissame; Buijs, Mark; Coppola, Stefano; Löhr, Matthias; Funel, Niccola; Heuchel, Rainer; Carnevale, Ilaria; Schmidt, Thomas; Mantini, Giulia; Avan, Amir; Saso, Luciano; Peters, Godefridus J.; Giovannetti, Elisa.

In: Cancers, Vol. 11, No. 5, 638, 2019.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Role of c-MET inhibitors in overcoming drug resistance in spheroid models of primary human pancreatic cancer and stellate cells

AU - Firuzi, Omidreza

AU - Che, Pei Pei

AU - el Hassouni, Btissame

AU - Buijs, Mark

AU - Coppola, Stefano

AU - Löhr, Matthias

AU - Funel, Niccola

AU - Heuchel, Rainer

AU - Carnevale, Ilaria

AU - Schmidt, Thomas

AU - Mantini, Giulia

AU - Avan, Amir

AU - Saso, Luciano

AU - Peters, Godefridus J.

AU - Giovannetti, Elisa

PY - 2019

Y1 - 2019

N2 - Pancreatic stellate cells (PSCs) are a key component of tumor microenvironment in pancreatic ductal adenocarcinoma (PDAC) and contribute to drug resistance. c-MET receptor tyrosine kinase activation plays an important role in tumorigenesis in different cancers including PDAC. In this study, effects of PSC conditioned medium (PCM) on c-MET phosphorylation (by immunocytochemistry enzyme-linked immunosorbent assay (ELISA)) and drug response (by sulforhodamine B assay) were investigated in five primary PDAC cells. In novel 3D-spheroid co-cultures of cyan fluorescence protein (CFP)-firefly luciferase (Fluc)-expressing primary human PDAC cells and green fluorescence protein (GFP)-expressing immortalized PSCs, PDAC cell growth and chemosensitivity were examined by luciferase assay, while spheroids’ architecture was evaluated by confocal microscopy. The highest phospho-c-MET expression was detected in PDAC5 and its subclone sorted for “stage specific embryonic antigen-4" (PDAC5 (SSEA4)). PCM of cells pre-incubated with PDAC conditioned medium, containing increased hepatocyte growth factor (HGF) levels, made PDAC cells significantly more resistant to gemcitabine, but not to c-MET inhibitors. Hetero-spheroids containing both PSCs and PDAC5 (SSEA4) cells were more resistant to gemcitabine compared to PDAC5 (SSEA4) homo-spheroids. However, c-MET inhibitors (tivantinib, PHA-665752 and crizotinib) were equally effective in both spheroid models. Experiments with primary human PSCs confirmed the main findings. In conclusion, we developed spheroid models to evaluate PSC-PDAC reciprocal interaction, unraveling c-MET inhibition as an important therapeutic option against drug resistant PDAC.

AB - Pancreatic stellate cells (PSCs) are a key component of tumor microenvironment in pancreatic ductal adenocarcinoma (PDAC) and contribute to drug resistance. c-MET receptor tyrosine kinase activation plays an important role in tumorigenesis in different cancers including PDAC. In this study, effects of PSC conditioned medium (PCM) on c-MET phosphorylation (by immunocytochemistry enzyme-linked immunosorbent assay (ELISA)) and drug response (by sulforhodamine B assay) were investigated in five primary PDAC cells. In novel 3D-spheroid co-cultures of cyan fluorescence protein (CFP)-firefly luciferase (Fluc)-expressing primary human PDAC cells and green fluorescence protein (GFP)-expressing immortalized PSCs, PDAC cell growth and chemosensitivity were examined by luciferase assay, while spheroids’ architecture was evaluated by confocal microscopy. The highest phospho-c-MET expression was detected in PDAC5 and its subclone sorted for “stage specific embryonic antigen-4" (PDAC5 (SSEA4)). PCM of cells pre-incubated with PDAC conditioned medium, containing increased hepatocyte growth factor (HGF) levels, made PDAC cells significantly more resistant to gemcitabine, but not to c-MET inhibitors. Hetero-spheroids containing both PSCs and PDAC5 (SSEA4) cells were more resistant to gemcitabine compared to PDAC5 (SSEA4) homo-spheroids. However, c-MET inhibitors (tivantinib, PHA-665752 and crizotinib) were equally effective in both spheroid models. Experiments with primary human PSCs confirmed the main findings. In conclusion, we developed spheroid models to evaluate PSC-PDAC reciprocal interaction, unraveling c-MET inhibition as an important therapeutic option against drug resistant PDAC.

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U2 - 10.3390/cancers11050638

DO - 10.3390/cancers11050638

M3 - Article

VL - 11

JO - Cancers (Basel)

JF - Cancers (Basel)

SN - 2072-6694

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