title: Epigenetic biomarkers of T-cells in human glioma.
abstract: Immune factors are thought to influence glioma risk and outcomes, but immune profiling studies to further our understanding of the immune response are limited by current immunodiagnostic methods. We developed a new assay to capture glioma immune biology based on quantitative methylation specific PCR (qMSP) of two T-cell genes (CD3Z: T-cells, and FOXP3: Tregs). Flow cytometry of T-cells correlated well with the CD3Z demethylation assay (r = 0.93; p < 2.2 × 10 (-16) ), demonstrating the validity of the assay. Furthermore, there was a high correlation between qMSP and immunohistochemistry (IHC) in quantifying tumor infiltrating T-cells (r = 0.85; p = 3.4 × 10 (-11) ). Applying our qMSP methods to archival whole blood from 65 glioblastoma multiforme (GBM) cases and 94 non-diseased controls, GBM cases had highly statistically significantly lower T-cells (p = 1.7 × 10 (-9) ) as well as Tregs (p = 5.2 × 10 (-11) ) and a modestly lower ratio of Tregs/T-cells (p = 0.024). Applying the methods to 120 excised glioma tumors, we observed that tumor infiltrating CD3+ T-cells were positively correlated with glioma tumor grade (p = 5.7 × 10 (-7) ), and that Tregs were enriched in tumors compared with peripheral blood indicating active chemoattraction of suppressive Tregs into the tumor compartment. Poorer patient survival was correlated with higher levels of tumor infiltrating T-cells (p = 0.01) and Tregs (p = 0.04). DNA methylation based immunodiagnostics represent a new generation of powerful laboratory tools offering many advantages over conventional methods that will facilitate large clinical epidemiologic studies and capitalize on stored archival blood and tissue banks.
authors.affiliation: Department of Neurological Surgery, University of California San Francisco, San Francisco, CA USA. John.Wiencke@ucsf.edu
authors.forename: John K
authors.initials: JK
authors.lastname: Wiencke
authors.forename: William P
authors.initials: WP
authors.lastname: Accomando
authors.forename: Shichun
authors.initials: S
authors.lastname: Zheng
authors.forename: Joe
authors.initials: J
authors.lastname: Patoka
authors.forename: Xiaoqin
authors.initials: X
authors.lastname: Dou
authors.forename: Joanna J
authors.initials: JJ
authors.lastname: Phillips
authors.forename: George
authors.initials: G
authors.lastname: Hsuang
authors.forename: Brock C
authors.initials: BC
authors.lastname: Christensen
authors.forename: E Andres
authors.initials: EA
authors.lastname: Houseman
authors.forename: Devin C
authors.initials: DC
authors.lastname: Koestler
authors.forename: Paige
authors.initials: P
authors.lastname: Bracci
authors.forename: Joseph L
authors.initials: JL
authors.lastname: Wiemels
authors.forename: Margaret
authors.initials: M
authors.lastname: Wrensch
authors.forename: Heather H
authors.initials: HH
authors.lastname: Nelson
authors.forename: Karl T
authors.initials: KT
authors.lastname: Kelsey
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000032:analysis / Q000235:genetics*; D001932:Brain Neoplasms / Q000235:genetics* / Q000401:mortality / Q000473:pathology / Q000601:surgery; D017252:CD3 Complex / Q000235:genetics*; D016022:Case-Control Studies; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D005434:Flow Cytometry; D051858:Forkhead Transcription Factors / Q000235:genetics*; D005909:Glioblastoma / Q000235:genetics* / Q000276:immunology / Q000401:mortality / Q000473:pathology* / Q000601:surgery; D005910:Glioma / Q000235:genetics* / Q000473:pathology; D006801:Humans; D016246:Lymphocytes, Tumor-Infiltrating / Q000473:pathology; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods; D011401:Promoter Regions, Genetic; D050378:T-Lymphocytes, Regulatory; D055815:Young Adult
chemical_list: D014408:Biomarkers, Tumor; D017252:CD3 Complex; C121704:CD3 antigen, zeta chain; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors