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cMET Activation and EGFR-Directed Therapy Resistance in Triple-Negative Breast Cancer.

Sohn J, Liu S, Parinyanitikul N, Lee J, Hortobagyi GN, Mills GB, Ueno NT, Gonzalez-Angulo AM - J Cancer (2014)

Bottom Line: However, anti-EGFR therapies have not been effective in these patients.In western blotting, combination therapy resulted in abrogation of pAKT and pMAPK while monotherapy did not.Targeting both EGFR and MET receptors may provide an effective therapeutic strategy in TNBC.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Breast Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA ; 2. Division of Medical Oncology, Department of Internal Medicine, Breast Cancer Clinic, Severance Hospital, Yonsei University College of Medicine, Seoul, Korea.

ABSTRACT

Background: EGFR expression and pathway activation are common in triple-negative breast cancer (TNBC). However, anti-EGFR therapies have not been effective in these patients. We aimed to study the efficacy of targeting MET in overcoming resistance to EGFR therapy in TNBC cell lines.

Methods: TNBC lines (MDA-MB-468, HCC-1395, and MDA-MB-231), and a hormone receptor-positive breast cancer line (T47D) were stimulated with epidermal growth factor (EGF) and hepatocyte growth factor (HGF). Lines were then treated with different concentrations of EGFR inhibitors (gefitinib or cetuximab), with or without a MET tyrosine kinase inhibitor (EMD 1214063). Proliferation was measured by MTS assay, in soft agar and with a matrigel assay. Synergy was measured with Calcusyn. Protein expression and signaling were examined with immunoblotting.

Results: There was activation of ligand-receptor-downstream signaling pathways in MDA-MB-468 and HCC-1395 upon stimulation with EGF and HGF. In these cell lines, we observed synergism when combining EGFR and MET inhibitors. These results were observed across assays. In western blotting, combination therapy resulted in abrogation of pAKT and pMAPK while monotherapy did not.

Conclusion: Our data demonstrate that dual EGFR/MET inhibition is synergistic in TNBC. Targeting both EGFR and MET receptors may provide an effective therapeutic strategy in TNBC.

No MeSH data available.


Related in: MedlinePlus

Western blot results after stimulation with EGF, HGF, and their combination in TNBC cell lines, and T47D. Cells were starved with RPMI 1640 medium overnight and next morning, they were stimulated with EGF 20ng/mL and/or recombinant human HGF 75ng/mL for 10 minutes before cell lysis. Thereafter, cell lysates were collected and loaded for Western blot with antibodies as indicated. Actin was used as a loading control.
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Figure 6: Western blot results after stimulation with EGF, HGF, and their combination in TNBC cell lines, and T47D. Cells were starved with RPMI 1640 medium overnight and next morning, they were stimulated with EGF 20ng/mL and/or recombinant human HGF 75ng/mL for 10 minutes before cell lysis. Thereafter, cell lysates were collected and loaded for Western blot with antibodies as indicated. Actin was used as a loading control.

Mentions: To determine the potential mechanisms involved with the pathway inhibition, cells were treated with EGF and/or HGF after overnight starvation with RPMI-1640 without FBS and lysed to perform western blotting. MDA-MB-468 and HCC-1395 were responsive to both EGF and HGF stimulation consistent with the presence of functional cell surface receptors able to link to downstream signals including pMAPK and pAKT. However, MDA-MB-231 and T47D showed faint or negligible expression of pEGFR and pMET even after stimulation with growth factors and there were little changes in levels of downstream targets including pMAPK and pAKT (Fig. 6). Accordingly, we designated MDA-MB-468 and HCC-1395 as cell lines that have active EGFR and MET pathways and MDA-MB-231 and T47D as cell lines with inactive EGFR and MET pathways. These results are consistent with MDA-MB-468 and HCC-1395 responding to gefitinib and EMD 121463 and with MDA-MB-231 and T47D not responding or poorly responding.


cMET Activation and EGFR-Directed Therapy Resistance in Triple-Negative Breast Cancer.

Sohn J, Liu S, Parinyanitikul N, Lee J, Hortobagyi GN, Mills GB, Ueno NT, Gonzalez-Angulo AM - J Cancer (2014)

Western blot results after stimulation with EGF, HGF, and their combination in TNBC cell lines, and T47D. Cells were starved with RPMI 1640 medium overnight and next morning, they were stimulated with EGF 20ng/mL and/or recombinant human HGF 75ng/mL for 10 minutes before cell lysis. Thereafter, cell lysates were collected and loaded for Western blot with antibodies as indicated. Actin was used as a loading control.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4216798&req=5

Figure 6: Western blot results after stimulation with EGF, HGF, and their combination in TNBC cell lines, and T47D. Cells were starved with RPMI 1640 medium overnight and next morning, they were stimulated with EGF 20ng/mL and/or recombinant human HGF 75ng/mL for 10 minutes before cell lysis. Thereafter, cell lysates were collected and loaded for Western blot with antibodies as indicated. Actin was used as a loading control.
Mentions: To determine the potential mechanisms involved with the pathway inhibition, cells were treated with EGF and/or HGF after overnight starvation with RPMI-1640 without FBS and lysed to perform western blotting. MDA-MB-468 and HCC-1395 were responsive to both EGF and HGF stimulation consistent with the presence of functional cell surface receptors able to link to downstream signals including pMAPK and pAKT. However, MDA-MB-231 and T47D showed faint or negligible expression of pEGFR and pMET even after stimulation with growth factors and there were little changes in levels of downstream targets including pMAPK and pAKT (Fig. 6). Accordingly, we designated MDA-MB-468 and HCC-1395 as cell lines that have active EGFR and MET pathways and MDA-MB-231 and T47D as cell lines with inactive EGFR and MET pathways. These results are consistent with MDA-MB-468 and HCC-1395 responding to gefitinib and EMD 121463 and with MDA-MB-231 and T47D not responding or poorly responding.

Bottom Line: However, anti-EGFR therapies have not been effective in these patients.In western blotting, combination therapy resulted in abrogation of pAKT and pMAPK while monotherapy did not.Targeting both EGFR and MET receptors may provide an effective therapeutic strategy in TNBC.

View Article: PubMed Central - PubMed

Affiliation: 1. Department of Breast Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA ; 2. Division of Medical Oncology, Department of Internal Medicine, Breast Cancer Clinic, Severance Hospital, Yonsei University College of Medicine, Seoul, Korea.

ABSTRACT

Background: EGFR expression and pathway activation are common in triple-negative breast cancer (TNBC). However, anti-EGFR therapies have not been effective in these patients. We aimed to study the efficacy of targeting MET in overcoming resistance to EGFR therapy in TNBC cell lines.

Methods: TNBC lines (MDA-MB-468, HCC-1395, and MDA-MB-231), and a hormone receptor-positive breast cancer line (T47D) were stimulated with epidermal growth factor (EGF) and hepatocyte growth factor (HGF). Lines were then treated with different concentrations of EGFR inhibitors (gefitinib or cetuximab), with or without a MET tyrosine kinase inhibitor (EMD 1214063). Proliferation was measured by MTS assay, in soft agar and with a matrigel assay. Synergy was measured with Calcusyn. Protein expression and signaling were examined with immunoblotting.

Results: There was activation of ligand-receptor-downstream signaling pathways in MDA-MB-468 and HCC-1395 upon stimulation with EGF and HGF. In these cell lines, we observed synergism when combining EGFR and MET inhibitors. These results were observed across assays. In western blotting, combination therapy resulted in abrogation of pAKT and pMAPK while monotherapy did not.

Conclusion: Our data demonstrate that dual EGFR/MET inhibition is synergistic in TNBC. Targeting both EGFR and MET receptors may provide an effective therapeutic strategy in TNBC.

No MeSH data available.


Related in: MedlinePlus