Skip to main content

Characterization of MGMT and EGFR protein expression in glioblastoma and association with survival

Journal of Neuro-Oncology volume 146, pages 163–170 (2020)Cite this article

Abstract

Purpose

Understanding the molecular landscape of glioblastoma (GBM) is increasingly important in the age of targeted therapy. O-6-Methylguanine-DNA methyltransferase (MGMT) promoter methylation and EGFR amplification are markers that may play a role in prognostication, treatment, and/or clinical trial eligibility. Quantification of MGMT and EGFR protein expression may offer an alternative strategy towards understanding GBM. Here, we quantify baseline expression of MGMT and EGFR protein in newly diagnosed GBM samples using mass spectrometry. We correlate findings with MGMT methylation and EGFR amplification statuses and survival.

Methods

We retrospectively identified adult patients with newly diagnosed resected GBM. MGMT and EGFR protein expression were quantified using a selected reaction monitoring mass spectrometry assay. Protein levels were correlated with MGMT methylation and EGFR amplification and survival data.

Results

We found a statistically significant association between MGMT protein expression and promoter methylation status (p = 0.02) as well as between EGFR protein expression and EGFR amplification (p < 0.0001). EGFR protein expression and amplification were more tightly associated than MGMT protein expression and methylation. Only MGMT promoter methylation was statistically significantly associated with progression-free and overall survival.

Conclusions

Unlike EGFR protein expression and EGFR amplification which are strongly associated, only a weak association was seen between MGMT protein expression and promoter methylation. Quantification of MGMT protein expression was inferior to MGMT methylation for prognostication in GBM. Discordance was observed between EGFR amplification and EGFR protein expression; additional study is warranted to determine whether EGFR protein expression is a better biomarker than EGFR amplification for clinical decisions and trial enrollment.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2

References

  1. Stupp R, Mason WP, van den Bent MJ, Weller M, Fisher B, Taphoorn MJ, Belanger K, Brandes AA, Marosi C, Bogdahn U, Curschmann J, Janzer RC, Ludwin SK, Gorlia T, Allgeier A, Lacombe D, Cairncross JG, Eisenhauer E, Mirimanoff RO, Radiotherapy G, National Cancer Institute of Canada Clinical Trials G (2005) European Organisation for Research and Treatment of Cancer Brain Tumor. N Engl J Med 352:987–996. https://doi.org/10.1056/NEJMoa043330

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  2. Stupp R, Taillibert S, Kanner A, Read W, Steinberg D, Lhermitte B, Toms S, Idbaih A, Ahluwalia MS, Fink K, Di Meco F, Lieberman F, Zhu JJ, Stragliotto G, Tran D, Brem S, Hottinger A, Kirson ED, Lavy-Shahaf G, Weinberg U, Kim CY, Paek SH, Nicholas G, Bruna J, Hirte H, Weller M, Palti Y, Hegi ME, Ram Z (2017) Effect of tumor-treating fields plus maintenance temozolomide vs maintenance temozolomide alone on survival in patients with glioblastoma: a randomized clinical trial. JAMA 318:2306–2316. https://doi.org/10.1001/jama.2017.18718

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  3. Esteller M, Garcia-Foncillas J, Andion E, Goodman SN, Hidalgo OF, Vanaclocha V, Baylin SB, Herman JG (2000) Inactivation of the DNA-repair gene MGMT and the clinical response of gliomas to alkylating agents. N Engl J Med 343:1350–1354. doi:https://doi.org/10.1056/NEJM200011093431901

    CAS  Article  PubMed  Google Scholar 

  4. Hegi ME, Diserens AC, Gorlia T, Hamou MF, de Tribolet N, Weller M, Kros JM, Hainfellner JA, Mason W, Mariani L, Bromberg JE, Hau P, Mirimanoff RO, Cairncross JG, Janzer RC, Stupp R (2005) MGMT gene silencing and benefit from temozolomide in glioblastoma. N Engl J Med 352:997–1003. https://doi.org/10.1056/NEJMoa043331

    CAS  Article  Google Scholar 

  5. Mansouri A, Hachem LD, Mansouri S, Nassiri F, Laperriere NJ, Xia D, Lindeman NI, Wen PY, Chakravarti A, Mehta MP, Hegi ME, Stupp R, Aldape KD, Zadeh G (2019) MGMT promoter methylation status testing to guide therapy for glioblastoma: refining the approach based on emerging evidence and current challenges. Neuro Oncol 21:167–178. https://doi.org/10.1093/neuonc/noy132

    Article  PubMed  Google Scholar 

  6. Wick W, Weller M, van den Bent M, Sanson M, Weiler M, von Deimling A, Plass C, Hegi M, Platten M, Reifenberger G (2014) MGMT testing: the challenges for biomarker-based glioma treatment. Nat Rev Neurol 10:372–385. https://doi.org/10.1038/nrneurol.2014.100

    CAS  Article  PubMed  Google Scholar 

  7. Lassman A, Dimino C, Mansukhani M, Murty V, Ansell PJ, Bain E, Holen KD, Roberts-Rapp LA, Lee J, Curran W, Mehta MP, Canoll P (2017) Concordance of Egfr and Mgmt analyses between local and central laboratories: implications for clinical trial design and precision medicine for depatuxizumab-mafodotin (Abt-414) in glioblastoma (Gbm). Neuro-Oncology 19:15–15

    Article  Google Scholar 

  8. Grasbon-Frodl EM, Kreth FW, Ruiter M, Schnell O, Bise K, Felsberg J, Reifenberger G, Tonn JC, Kretzschmar HA (2007) Intratumoral homogeneity of MGMT promoter hypermethylation as demonstrated in serial stereotactic specimens from anaplastic astrocytomas and glioblastomas. Int J Cancer 121:2458–2464. doi:https://doi.org/10.1002/ijc.23020

    CAS  Article  PubMed  Google Scholar 

  9. Preusser M, Charles Janzer R, Felsberg J, Reifenberger G, Hamou MF, Diserens AC, Stupp R, Gorlia T, Marosi C, Heinzl H, Hainfellner JA, Hegi M (2008) Anti-O6-methylguanine-methyltransferase (MGMT) immunohistochemistry in glioblastoma multiforme: observer variability and lack of association with patient survival impede its use as clinical biomarker. Brain Pathol 18:520–532. https://doi.org/10.1111/j.1750-3639.2008.00153.x

    CAS  Article  PubMed  Google Scholar 

  10. Cao VT, Jung TY, Jung S, Jin SG, Moon KS, Kim IY, Kang SS, Park CS, Lee KH, Chae HJ (2009) The correlation and prognostic significance of MGMT promoter methylation and MGMT protein in glioblastomas. Neurosurgery 65:866–875. https://doi.org/10.1227/01.NEU.0000357325.90347.A1 discussion 875

    Article  PubMed  Google Scholar 

  11. Hembrough T, Thyparambil S, Liao WL, Darfler MM, Abdo J, Bengali KM, Taylor P, Tong J, Lara-Guerra H, Waddell TK, Moran MF, Tsao MS, Krizman DB, Burrows J (2012) Selected reaction monitoring (SRM) analysis of epidermal growth factor receptor (EGFR) in formalin fixed tumor tissue. Clin Proteom 9:5. https://doi.org/10.1186/1559-0275-9-5

    CAS  Article  Google Scholar 

  12. Schwartz S, Szeto C, Tian Y, Cecchi F, Corallo S, Calegari MA, Di Bartolomeo M, Morano F, Raimondi A, Fuca G, Martinetti A, De Pascalis I, Martini M, Belfiore A, Milione M, Orlandi A, Barault L, Barone C, de Braud F, Di Nicolantonio F, Benz S, Hembrough T, Pietrantonio F (2019) Refining the selection of patients with metastatic colorectal cancer for treatment with temozolomide using proteomic analysis of O6-methylguanine-DNA-methyltransferase. Eur J Cancer 107:164–174. doi:https://doi.org/10.1016/j.ejca.2018.11.016

    CAS  Article  PubMed  Google Scholar 

  13. Brell M, Tortosa A, Verger E, Gil JM, Vinolas N, Villa S, Acebes JJ, Caral L, Pujol T, Ferrer I, Ribalta T, Graus F (2005) Prognostic significance of O6-methylguanine-DNA methyltransferase determined by promoter hypermethylation and immunohistochemical expression in anaplastic gliomas. Clin Cancer Res 11:5167–5174. https://doi.org/10.1158/1078-0432.CCR-05-0230

    CAS  Article  PubMed  Google Scholar 

  14. Sasai K, Nodagashira M, Nishihara H, Aoyanagi E, Wang L, Katoh M, Murata J, Ozaki Y, Ito T, Fujimoto S, Kaneko S, Nagashima K, Tanaka S (2008) Careful exclusion of non-neoplastic brain components is required for an appropriate evaluation of O6-methylguanine-DNA methyltransferase status in glioma: relationship between immunohistochemistry and methylation analysis. Am J Surg Pathol 32:1220–1227. https://doi.org/10.1097/PAS.0b013e318164c3f0

    Article  PubMed  Google Scholar 

  15. Mason S, McDonald K (2012) MGMT testing for glioma in clinical laboratories: discordance with methylation analyses prevents the implementation of routine immunohistochemistry. J Cancer Res Clin Oncol 138:1789–1797. https://doi.org/10.1007/s00432-012-1312-1

    CAS  Article  PubMed  Google Scholar 

  16. Friedman HS, McLendon RE, Kerby T, Dugan M, Bigner SH, Henry AJ, Ashley DM, Krischer J, Lovell S, Rasheed K, Marchev F, Seman AJ, Cokgor I, Rich J, Stewart E, Colvin OM, Provenzale JM, Bigner DD, Haglund MM, Friedman AH, Modrich PL (1998) DNA mismatch repair and O6-alkylguanine-DNA alkyltransferase analysis and response to Temodal in newly diagnosed malignant glioma. J Clin Oncol 16:3851–3857. https://doi.org/10.1200/JCO.1998.16.12.3851

    CAS  Article  PubMed  Google Scholar 

  17. Belanich M, Pastor M, Randall T, Guerra D, Kibitel J, Alas L, Li B, Citron M, Wasserman P, White A, Eyre H, Jaeckle K, Schulman S, Rector D, Prados M, Coons S, Shapiro W, Yarosh D (1996) Retrospective study of the correlation between the DNA repair protein alkyltransferase and survival of brain tumor patients treated with carmustine. Cancer Res 56:783–788

    CAS  PubMed  Google Scholar 

  18. Kristensen LS, Michaelsen SR, Dyrbye H, Aslan D, Grunnet K, Christensen IJ, Poulsen HS, Gronbaek K, Broholm H (2016) Assessment of quantitative and allelic MGMT methylation patterns as a prognostic marker in glioblastoma. J Neuropathol Exp Neurol 75:246–255. https://doi.org/10.1093/jnen/nlv024

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  19. Quillien V, Lavenu A, Karayan-Tapon L, Carpentier C, Labussiere M, Lesimple T, Chinot O, Wager M, Honnorat J, Saikali S, Fina F, Sanson M, Figarella-Branger D (2012) Comparative assessment of 5 methods (methylation-specific polymerase chain reaction, MethyLight, pyrosequencing, methylation-sensitive high-resolution melting, and immunohistochemistry) to analyze O6-methylguanine-DNA-methyltranferase in a series of 100 glioblastoma patients. Cancer 118:4201–4211. https://doi.org/10.1002/cncr.27392

    CAS  Article  PubMed  Google Scholar 

  20. Maxwell JA, Johnson SP, Quinn JA, McLendon RE, Ali-Osman F, Friedman AH, Herndon JE 2, Bierau K, Bigley J, Bigner DD, Friedman HS (2006) Quantitative analysis of O6-alkylguanine-DNA alkyltransferase in malignant glioma. Mol Cancer Ther 5:2531–2539. https://doi.org/10.1158/1535-7163.MCT-06-0106

    CAS  Article  PubMed  Google Scholar 

  21. Karayan-Tapon L, Quillien V, Guilhot J, Wager M, Fromont G, Saikali S, Etcheverry A, Hamlat A, Loussouarn D, Campion L, Campone M, Vallette FM, Gratas-Rabbia-Re C (2010) Prognostic value of O6-methylguanine-DNA methyltransferase status in glioblastoma patients, assessed by five different methods. J Neurooncol 97:311–322. https://doi.org/10.1007/s11060-009-0031-1

    CAS  Article  PubMed  Google Scholar 

  22. Lavon I, Fuchs D, Zrihan D, Efroni G, Zelikovitch B, Fellig Y, Siegal T (2007) Novel mechanism whereby nuclear factor kappaB mediates DNA damage repair through regulation of O(6)-methylguanine-DNA-methyltransferase. Cancer Res 67:8952–8959. https://doi.org/10.1158/0008-5472.CAN-06-3820

    CAS  Article  PubMed  Google Scholar 

  23. Blough MD, Zlatescu MC, Cairncross JG (2007) O6-Methylguanine-DNA methyltransferase regulation by p53 in astrocytic cells. Cancer Res 67:580–584. https://doi.org/10.1158/0008-5472.CAN-06-2782

    CAS  Article  PubMed  Google Scholar 

  24. Everhard S, Tost J, El Abdalaoui H, Criniere E, Busato F, Marie Y, Gut IG, Sanson M, Mokhtari K, Laigle-Donadey F, Hoang-Xuan K, Delattre JY, Thillet J (2009) Identification of regions correlating MGMT promoter methylation and gene expression in glioblastomas. Neuro Oncol 11:348–356. https://doi.org/10.1215/15228517-2009-001

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  25. Bady P, Sciuscio D, Diserens AC, Bloch J, van den Bent MJ, Marosi C, Dietrich PY, Weller M, Mariani L, Heppner FL, McDonald DR, Lacombe D, Stupp R, Delorenzi M, Hegi ME (2012) MGMT methylation analysis of glioblastoma on the Infinium methylation BeadChip identifies two distinct CpG regions associated with gene silencing and outcome, yielding a prediction model for comparisons across datasets, tumor grades, and CIMP-status. Acta Neuropathol 124:547–560. https://doi.org/10.1007/s00401-012-1016-2

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  26. van den Bent MJ, Gravendeel LA, Gorlia T, Kros JM, Lapre L, Wesseling P, Teepen JL, Idbaih A, Sanson M, Smitt PA, French PJ (2011) A hypermethylated phenotype is a better predictor of survival than MGMT methylation in anaplastic oligodendroglial brain tumors: a report from EORTC study 26951. Clin Cancer Res 17:7148–7155. https://doi.org/10.1158/1078-0432.CCR-11-1274

    Article  PubMed  Google Scholar 

  27. McCormack AI, Wass JA, Grossman AB (2011) Aggressive pituitary tumours: the role of temozolomide and the assessment of MGMT status. Eur J Clin Invest 41:1133–1148. https://doi.org/10.1111/j.1365-2362.2011.02520.x

    CAS  Article  PubMed  Google Scholar 

  28. Bengtsson D, Schroder HD, Andersen M, Maiter D, Berinder K, Feldt Rasmussen U, Rasmussen AK, Johannsson G, Hoybye C, van der Lely AJ, Petersson M, Ragnarsson O, Burman P (2015) Long-term outcome and MGMT as a predictive marker in 24 patients with atypical pituitary adenomas and pituitary carcinomas given treatment with temozolomide. J Clin Endocrinol Metab 100:1689–1698. https://doi.org/10.1210/jc.2014-4350

    CAS  Article  PubMed  Google Scholar 

  29. Jackson C, Noorbakhsh S, Sundaram R, Kalathil A, Bindra R (2018) Synthetic sensitization of Mgmt-deficient tumor cells to temozolomide using Atr inhibitors. Neuro-Oncology 20:222–222

    Article  Google Scholar 

  30. Felsberg J, Hentschel B, Kaulich K, Gramatzki D, Zacher A, Malzkorn B, Kamp M, Sabel M, Simon M, Westphal M, Schackert G, Tonn JC, Pietsch T, von Deimling A, Loeffler M, Reifenberger G, Weller M, German Glioma N (2017) Epidermal growth factor receptor variant III (EGFRvIII) positivity in EGFR-amplified glioblastomas: prognostic role and comparison between primary and recurrent tumors. Clin Cancer Res 23:6846–6855. https://doi.org/10.1158/1078-0432.CCR-17-0890

    CAS  Article  Google Scholar 

  31. Sawaya R (1999) Extent of resection in malignant gliomas: a critical summary. J Neurooncol 42:303–305

    CAS  Article  Google Scholar 

  32. Sanai N, Berger MS (2008) Glioma extent of resection and its impact on patient outcome. Neurosurgery 62:753–764. https://doi.org/10.1227/01.neu.0000318159.21731.cf discussion 264–756

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

The authors acknowledge the support of the NIH Cancer Center Support Grant P30 CA008748. The authors wish to thank Susan Atlas and Moira McGrath for their editorial assistance.

Funding

This study was funded in part by NantOmics which holds the patent for the SRM assay discussed in the manuscript. The authors acknowledge the support of the NIH Cancer Center Support Grant P30 CA008748.

Author information

Affiliations

  1. Department of Neurology, Memorial Sloan Kettering Cancer Center, New York, NY, 10065, USA

    Lauren R. Schaff & Andrew L. Lin

  2. NantOmics, Culver City, CA, 90230, USA

    Dongyao Yan, Sheeno Thyparambil, Yuan Tian, Fabiola Cecchi & Todd Hembrough

  3. Department of Pathology, Memorial Sloan Kettering Cancer Center, New York, NY, 10065, USA

    Marc Rosenblum

  4. Department of Biostatistics, Memorial Sloan Kettering Cancer Center, New York, NY, 10065, USA

    Anne S. Reiner & Katherine S. Panageas

Authors
  1. Lauren R. Schaff
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Dongyao Yan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sheeno Thyparambil
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yuan Tian
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Fabiola Cecchi
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Marc Rosenblum
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Anne S. Reiner
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Katherine S. Panageas
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Todd Hembrough
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Andrew L. Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Lauren R. Schaff.

Ethics declarations

Conflict of interest

Authors Schaff, Rosenblum, Reiner, Panageas, and Lin have no conflict of interest. Authors Thyparambil and Hembrough have a patent for the SRM assay discussed in this manuscript and are employees of NantOmics. Authors Yan, Tian, and Cecchi are employees of NantOmics.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors. This study was approved by the Internal Review Board at MSKCC.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 38.4 kb)

Supplementary material 2 (XLSX 18.2 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Schaff, L.R., Yan, D., Thyparambil, S. et al. Characterization of MGMT and EGFR protein expression in glioblastoma and association with survival. J Neurooncol 146, 163–170 (2020). https://doi.org/10.1007/s11060-019-03358-x

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11060-019-03358-x

Keywords

  • Glioblastoma
  • MGMT
  • EGFR
  • Mass spectrometry
  • Proteomics