Real-Time Methylation-Specific Polymerase Chain Reaction for MGMT Promoter Methylation Clinical Testing in Glioblastoma

An Alternative Detection Method for a Heterogeneous Process

Cristiane M. Ida, Malinda L. Butz, Robert Brian Jenkins, Jann N Sarkaria, Gaspar J. Kitange, Caterina Giannini, Benjamin R. Kipp

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Objectives: To develop and evaluate a real-time methylation-specific polymerase chain reaction (RT-MSP) MGMT assay, with a particular focus on small biopsies and indeterminate testing results.

Methods: We assessed formalin-fixed paraffin-embedded glioblastoma or gliosarcoma specimens (n = 641). A test-validation group (n = 51) with previously obtained reference laboratory (RL) results was used to determine performance characteristics of the RT-MSP assay. An indeterminate (equivocal) category was established for cases that could not be clearly classified as positive or negative.

Results: Overall agreement of RT-MSP and RL results was 91% (41/45 nonindeterminate cases). Discordant cases were tested by pyrosequencing, and results were most concordant with RT-MSP. Among cases with limited amounts of tissue (n = 7), six yielded valid results by RT-MSP (all negative); the single invalid result consisted of a stereotactic biopsy specimen obtained 14 years prior. A subset of indeterminate cases obtained during clinical testing (n = 18/575 [3%]) was also evaluated by pyrosequencing and showed a heterogeneous pattern of methylation across the eight interrogated CpG sites.

Conclusions: The RT-MSP assay that we developed in-house is a robust clinical detection method for the heterogeneous process of MGMT promoter methylation in glioblastoma.

Original languageEnglish (US)
Pages (from-to)296-307
Number of pages12
JournalAmerican Journal of Clinical Pathology
Volume148
Issue number4
DOIs
StatePublished - Oct 1 2017

Fingerprint

Glioblastoma
Methylation
Polymerase Chain Reaction
Gliosarcoma
Biopsy
Paraffin
Formaldehyde

Keywords

  • Astrocytoma
  • Brain
  • Epigenetics
  • Glioma
  • Molecular

ASJC Scopus subject areas

  • Pathology and Forensic Medicine

Cite this

@article{e271ba7e8c344dd6842d1206f7cd0c4d,
title = "Real-Time Methylation-Specific Polymerase Chain Reaction for MGMT Promoter Methylation Clinical Testing in Glioblastoma: An Alternative Detection Method for a Heterogeneous Process",
abstract = "Objectives: To develop and evaluate a real-time methylation-specific polymerase chain reaction (RT-MSP) MGMT assay, with a particular focus on small biopsies and indeterminate testing results.Methods: We assessed formalin-fixed paraffin-embedded glioblastoma or gliosarcoma specimens (n = 641). A test-validation group (n = 51) with previously obtained reference laboratory (RL) results was used to determine performance characteristics of the RT-MSP assay. An indeterminate (equivocal) category was established for cases that could not be clearly classified as positive or negative.Results: Overall agreement of RT-MSP and RL results was 91{\%} (41/45 nonindeterminate cases). Discordant cases were tested by pyrosequencing, and results were most concordant with RT-MSP. Among cases with limited amounts of tissue (n = 7), six yielded valid results by RT-MSP (all negative); the single invalid result consisted of a stereotactic biopsy specimen obtained 14 years prior. A subset of indeterminate cases obtained during clinical testing (n = 18/575 [3{\%}]) was also evaluated by pyrosequencing and showed a heterogeneous pattern of methylation across the eight interrogated CpG sites.Conclusions: The RT-MSP assay that we developed in-house is a robust clinical detection method for the heterogeneous process of MGMT promoter methylation in glioblastoma.",
keywords = "Astrocytoma, Brain, Epigenetics, Glioma, Molecular",
author = "Ida, {Cristiane M.} and Butz, {Malinda L.} and Jenkins, {Robert Brian} and Sarkaria, {Jann N} and Kitange, {Gaspar J.} and Caterina Giannini and Kipp, {Benjamin R.}",
year = "2017",
month = "10",
day = "1",
doi = "10.1093/ajcp/aqx073",
language = "English (US)",
volume = "148",
pages = "296--307",
journal = "American Journal of Clinical Pathology",
issn = "0002-9173",
publisher = "American Society of Clinical Pathologists",
number = "4",

}

TY - JOUR

T1 - Real-Time Methylation-Specific Polymerase Chain Reaction for MGMT Promoter Methylation Clinical Testing in Glioblastoma

T2 - An Alternative Detection Method for a Heterogeneous Process

AU - Ida, Cristiane M.

AU - Butz, Malinda L.

AU - Jenkins, Robert Brian

AU - Sarkaria, Jann N

AU - Kitange, Gaspar J.

AU - Giannini, Caterina

AU - Kipp, Benjamin R.

PY - 2017/10/1

Y1 - 2017/10/1

N2 - Objectives: To develop and evaluate a real-time methylation-specific polymerase chain reaction (RT-MSP) MGMT assay, with a particular focus on small biopsies and indeterminate testing results.Methods: We assessed formalin-fixed paraffin-embedded glioblastoma or gliosarcoma specimens (n = 641). A test-validation group (n = 51) with previously obtained reference laboratory (RL) results was used to determine performance characteristics of the RT-MSP assay. An indeterminate (equivocal) category was established for cases that could not be clearly classified as positive or negative.Results: Overall agreement of RT-MSP and RL results was 91% (41/45 nonindeterminate cases). Discordant cases were tested by pyrosequencing, and results were most concordant with RT-MSP. Among cases with limited amounts of tissue (n = 7), six yielded valid results by RT-MSP (all negative); the single invalid result consisted of a stereotactic biopsy specimen obtained 14 years prior. A subset of indeterminate cases obtained during clinical testing (n = 18/575 [3%]) was also evaluated by pyrosequencing and showed a heterogeneous pattern of methylation across the eight interrogated CpG sites.Conclusions: The RT-MSP assay that we developed in-house is a robust clinical detection method for the heterogeneous process of MGMT promoter methylation in glioblastoma.

AB - Objectives: To develop and evaluate a real-time methylation-specific polymerase chain reaction (RT-MSP) MGMT assay, with a particular focus on small biopsies and indeterminate testing results.Methods: We assessed formalin-fixed paraffin-embedded glioblastoma or gliosarcoma specimens (n = 641). A test-validation group (n = 51) with previously obtained reference laboratory (RL) results was used to determine performance characteristics of the RT-MSP assay. An indeterminate (equivocal) category was established for cases that could not be clearly classified as positive or negative.Results: Overall agreement of RT-MSP and RL results was 91% (41/45 nonindeterminate cases). Discordant cases were tested by pyrosequencing, and results were most concordant with RT-MSP. Among cases with limited amounts of tissue (n = 7), six yielded valid results by RT-MSP (all negative); the single invalid result consisted of a stereotactic biopsy specimen obtained 14 years prior. A subset of indeterminate cases obtained during clinical testing (n = 18/575 [3%]) was also evaluated by pyrosequencing and showed a heterogeneous pattern of methylation across the eight interrogated CpG sites.Conclusions: The RT-MSP assay that we developed in-house is a robust clinical detection method for the heterogeneous process of MGMT promoter methylation in glioblastoma.

KW - Astrocytoma

KW - Brain

KW - Epigenetics

KW - Glioma

KW - Molecular

UR - http://www.scopus.com/inward/record.url?scp=85031786454&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85031786454&partnerID=8YFLogxK

U2 - 10.1093/ajcp/aqx073

DO - 10.1093/ajcp/aqx073

M3 - Article

VL - 148

SP - 296

EP - 307

JO - American Journal of Clinical Pathology

JF - American Journal of Clinical Pathology

SN - 0002-9173

IS - 4

ER -