Afatinib and Temozolomide combination inhibits tumorigenesis by targeting EGFRvIII-cMet signaling in glioblastoma cells

Raghupathy Vengoji, Muzafar A Macha, Rama Krishna Nimmakayala, Satyanarayana Rachagani, Jawed A. Siddiqui, Kavita Mallya, Santhi Gorantla, Maneesh Jain, Moorthy Palanimuthu Ponnusamy, Surinder Kumar Batra, Nicole A Shonka

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Abstract

Background: Glioblastoma (GBM) is an aggressive brain tumor with universal recurrence and poor prognosis. The recurrence is largely driven by chemoradiation resistant cancer stem cells (CSCs). Epidermal growth factor receptor (EGFR) and its mutant EGFRvIII are amplified in ~ 60% and ~ 30% of GBM patients, respectively; however, therapies targeting EGFR have failed to improve disease outcome. EGFRvIII-mediated cross-activation of tyrosine kinase receptor, cMET, regulates GBM CSC maintenance and promote tumor recurrence. Here, we evaluated the efficacy of pan-EGFR inhibitor afatinib and Temozolomide (TMZ) combination on GBM in vitro and in vivo. Methods: We analyzed the effect of afatinib and temozolomide (TMZ) combination on GBM cells U87MG and U251 engineered to express wild type (WT) EGFR, EGFRvIII or EGFRvIII dead kinase, CSCs isolated from U87 and U87EGFRvIII in vitro. The therapeutic utility of the drug combination was investigated on tumor growth and progression using intracranially injected U87EGFRvIII GBM xenografts. Results: Afatinib and TMZ combination synergistically inhibited the proliferation, clonogenic survival, motility, invasion and induced senescence of GBM cells compared to monotherapy. Mechanistically, afatinib decreased U87EGFRvIII GBM cell proliferation and motility/invasion by inhibiting EGFRvIII/AKT, EGFRvIII/JAK2/STAT3, and focal adhesion kinase (FAK) signaling pathways respectively. Interestingly, afatinib specifically inhibited EGFRvIII-cMET crosstalk in CSCs, resulting in decreased expression of Nanog and Oct3/4, and in combination with TMZ significantly decreased their self-renewal property in vitro. More interestingly, afatinib and TMZ combination significantly decreased the xenograft growth and progression compared to single drug alone. Conclusion: Our study demonstrated significant inhibition of GBM tumorigenicity, CSC maintenance in vitro, and delayed tumor growth and progression in vivo by combination of afatinib and TMZ. Our results warrant evaluation of this drug combination in EGFR and EGFRvIII amplified GBM patients.

Original languageEnglish (US)
Article number266
JournalJournal of Experimental and Clinical Cancer Research
Volume38
Issue number1
DOIs
StatePublished - Jun 18 2019

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temozolomide
Glioblastoma
Carcinogenesis
Neoplastic Stem Cells
Epidermal Growth Factor Receptor
Drug Combinations
Heterografts
Recurrence
Growth
Maintenance
epidermal growth factor receptor VIII
BIBW 2992
Focal Adhesion Protein-Tyrosine Kinases
Neoplasms
Cell Aging

Keywords

  • Afatinib
  • Cancer stem cells
  • Epidermal growth factor receptor
  • Glioblastoma
  • Temozolomide

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

@article{77857dd8e99140dcb749f7a4bdcdbe38,
title = "Afatinib and Temozolomide combination inhibits tumorigenesis by targeting EGFRvIII-cMet signaling in glioblastoma cells",
abstract = "Background: Glioblastoma (GBM) is an aggressive brain tumor with universal recurrence and poor prognosis. The recurrence is largely driven by chemoradiation resistant cancer stem cells (CSCs). Epidermal growth factor receptor (EGFR) and its mutant EGFRvIII are amplified in ~ 60{\%} and ~ 30{\%} of GBM patients, respectively; however, therapies targeting EGFR have failed to improve disease outcome. EGFRvIII-mediated cross-activation of tyrosine kinase receptor, cMET, regulates GBM CSC maintenance and promote tumor recurrence. Here, we evaluated the efficacy of pan-EGFR inhibitor afatinib and Temozolomide (TMZ) combination on GBM in vitro and in vivo. Methods: We analyzed the effect of afatinib and temozolomide (TMZ) combination on GBM cells U87MG and U251 engineered to express wild type (WT) EGFR, EGFRvIII or EGFRvIII dead kinase, CSCs isolated from U87 and U87EGFRvIII in vitro. The therapeutic utility of the drug combination was investigated on tumor growth and progression using intracranially injected U87EGFRvIII GBM xenografts. Results: Afatinib and TMZ combination synergistically inhibited the proliferation, clonogenic survival, motility, invasion and induced senescence of GBM cells compared to monotherapy. Mechanistically, afatinib decreased U87EGFRvIII GBM cell proliferation and motility/invasion by inhibiting EGFRvIII/AKT, EGFRvIII/JAK2/STAT3, and focal adhesion kinase (FAK) signaling pathways respectively. Interestingly, afatinib specifically inhibited EGFRvIII-cMET crosstalk in CSCs, resulting in decreased expression of Nanog and Oct3/4, and in combination with TMZ significantly decreased their self-renewal property in vitro. More interestingly, afatinib and TMZ combination significantly decreased the xenograft growth and progression compared to single drug alone. Conclusion: Our study demonstrated significant inhibition of GBM tumorigenicity, CSC maintenance in vitro, and delayed tumor growth and progression in vivo by combination of afatinib and TMZ. Our results warrant evaluation of this drug combination in EGFR and EGFRvIII amplified GBM patients.",
keywords = "Afatinib, Cancer stem cells, Epidermal growth factor receptor, Glioblastoma, Temozolomide",
author = "Raghupathy Vengoji and Macha, {Muzafar A} and Nimmakayala, {Rama Krishna} and Satyanarayana Rachagani and Siddiqui, {Jawed A.} and Kavita Mallya and Santhi Gorantla and Maneesh Jain and {Palanimuthu Ponnusamy}, Moorthy and Batra, {Surinder Kumar} and Shonka, {Nicole A}",
year = "2019",
month = "6",
day = "18",
doi = "10.1186/s13046-019-1264-2",
language = "English (US)",
volume = "38",
journal = "Journal of Experimental and Clinical Cancer Research",
issn = "0392-9078",
publisher = "BioMed Central",
number = "1",

}

TY - JOUR

T1 - Afatinib and Temozolomide combination inhibits tumorigenesis by targeting EGFRvIII-cMet signaling in glioblastoma cells

AU - Vengoji, Raghupathy

AU - Macha, Muzafar A

AU - Nimmakayala, Rama Krishna

AU - Rachagani, Satyanarayana

AU - Siddiqui, Jawed A.

AU - Mallya, Kavita

AU - Gorantla, Santhi

AU - Jain, Maneesh

AU - Palanimuthu Ponnusamy, Moorthy

AU - Batra, Surinder Kumar

AU - Shonka, Nicole A

PY - 2019/6/18

Y1 - 2019/6/18

N2 - Background: Glioblastoma (GBM) is an aggressive brain tumor with universal recurrence and poor prognosis. The recurrence is largely driven by chemoradiation resistant cancer stem cells (CSCs). Epidermal growth factor receptor (EGFR) and its mutant EGFRvIII are amplified in ~ 60% and ~ 30% of GBM patients, respectively; however, therapies targeting EGFR have failed to improve disease outcome. EGFRvIII-mediated cross-activation of tyrosine kinase receptor, cMET, regulates GBM CSC maintenance and promote tumor recurrence. Here, we evaluated the efficacy of pan-EGFR inhibitor afatinib and Temozolomide (TMZ) combination on GBM in vitro and in vivo. Methods: We analyzed the effect of afatinib and temozolomide (TMZ) combination on GBM cells U87MG and U251 engineered to express wild type (WT) EGFR, EGFRvIII or EGFRvIII dead kinase, CSCs isolated from U87 and U87EGFRvIII in vitro. The therapeutic utility of the drug combination was investigated on tumor growth and progression using intracranially injected U87EGFRvIII GBM xenografts. Results: Afatinib and TMZ combination synergistically inhibited the proliferation, clonogenic survival, motility, invasion and induced senescence of GBM cells compared to monotherapy. Mechanistically, afatinib decreased U87EGFRvIII GBM cell proliferation and motility/invasion by inhibiting EGFRvIII/AKT, EGFRvIII/JAK2/STAT3, and focal adhesion kinase (FAK) signaling pathways respectively. Interestingly, afatinib specifically inhibited EGFRvIII-cMET crosstalk in CSCs, resulting in decreased expression of Nanog and Oct3/4, and in combination with TMZ significantly decreased their self-renewal property in vitro. More interestingly, afatinib and TMZ combination significantly decreased the xenograft growth and progression compared to single drug alone. Conclusion: Our study demonstrated significant inhibition of GBM tumorigenicity, CSC maintenance in vitro, and delayed tumor growth and progression in vivo by combination of afatinib and TMZ. Our results warrant evaluation of this drug combination in EGFR and EGFRvIII amplified GBM patients.

AB - Background: Glioblastoma (GBM) is an aggressive brain tumor with universal recurrence and poor prognosis. The recurrence is largely driven by chemoradiation resistant cancer stem cells (CSCs). Epidermal growth factor receptor (EGFR) and its mutant EGFRvIII are amplified in ~ 60% and ~ 30% of GBM patients, respectively; however, therapies targeting EGFR have failed to improve disease outcome. EGFRvIII-mediated cross-activation of tyrosine kinase receptor, cMET, regulates GBM CSC maintenance and promote tumor recurrence. Here, we evaluated the efficacy of pan-EGFR inhibitor afatinib and Temozolomide (TMZ) combination on GBM in vitro and in vivo. Methods: We analyzed the effect of afatinib and temozolomide (TMZ) combination on GBM cells U87MG and U251 engineered to express wild type (WT) EGFR, EGFRvIII or EGFRvIII dead kinase, CSCs isolated from U87 and U87EGFRvIII in vitro. The therapeutic utility of the drug combination was investigated on tumor growth and progression using intracranially injected U87EGFRvIII GBM xenografts. Results: Afatinib and TMZ combination synergistically inhibited the proliferation, clonogenic survival, motility, invasion and induced senescence of GBM cells compared to monotherapy. Mechanistically, afatinib decreased U87EGFRvIII GBM cell proliferation and motility/invasion by inhibiting EGFRvIII/AKT, EGFRvIII/JAK2/STAT3, and focal adhesion kinase (FAK) signaling pathways respectively. Interestingly, afatinib specifically inhibited EGFRvIII-cMET crosstalk in CSCs, resulting in decreased expression of Nanog and Oct3/4, and in combination with TMZ significantly decreased their self-renewal property in vitro. More interestingly, afatinib and TMZ combination significantly decreased the xenograft growth and progression compared to single drug alone. Conclusion: Our study demonstrated significant inhibition of GBM tumorigenicity, CSC maintenance in vitro, and delayed tumor growth and progression in vivo by combination of afatinib and TMZ. Our results warrant evaluation of this drug combination in EGFR and EGFRvIII amplified GBM patients.

KW - Afatinib

KW - Cancer stem cells

KW - Epidermal growth factor receptor

KW - Glioblastoma

KW - Temozolomide

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U2 - 10.1186/s13046-019-1264-2

DO - 10.1186/s13046-019-1264-2

M3 - Article

VL - 38

JO - Journal of Experimental and Clinical Cancer Research

JF - Journal of Experimental and Clinical Cancer Research

SN - 0392-9078

IS - 1

M1 - 266

ER -