Molecular diagnostics inpediatric glial tumors

Joon-Hyung Kim, MSIVWeill Cornell Medical College

Tumor type Molecular phenotype

Association

Oligodendroglioma

1p/19q codeletion Adults > Peds

Ganglioglioma BRAF V600E Peds > Adults

PXA BRAF V600E Peds = AdultsMesenchymal histology

Pilocytic astrocytoma

BRAF-KIAA1549 Tumor location/resectability (Cerebellar > Non-cerebellar)

DIPG 1q gain, 17p lossH3F3A, HIST1H3B mutations (K27M)

Peds > AdultsDIPG > Non-Brainstem pGBM

pGBM H3F3A mutation

ATRX, DAXX mutationATRX mutation

Peds > AdultsGBM > WHO I,II,III gliomasPeds > AdultsType of H3F3A mutation (G34R/V > K27M)

Key molecular alterations in pediatric glial tumors

1p/19q codeletion1p/19q codeletion

with polysomy “relative deletion”

No 1p/19q codeletion

Snuderl et al. Clin Cancer Res 2009.

Progression Free Survival Overall Survival

Anaplastic oligodendroglioma in adults (n=64)

1p and 19q deletions are infrequent in pediatric oligodendroglioma

Series N Age 1p del

19q del

1p/19q del

MGMT met

IDH1 mut

TP53 mut

Pollack et al., 2003

8 NA 1 2 NA NA NA NA

Raghavan et al. 2003

15 2-9 0 0 0 NA NA NA

11 10-18 2 0 3 NA NA NA

Kreiger et al., 2005

13 5-18 1 0 0 NA NA NA

Suri et al., 2011 7 3-18 0 0 0 5 0 0

Creach et al., 2012

15 1-18 NA NA 2 NA NA NA

Total 69 1-18 4 (7.4%)

2 (3.7%)

5 (8.2%)

5 (71%) 0 0

Tumor type Molecular phenotype

Association

Oligodendroglioma

1p/19q codeletion Adults > Peds

Ganglioglioma BRAF V600E Peds > Adults

PXA BRAF V600E Peds = AdultsMesenchymal histology

Pilocytic astrocytoma

BRAF-KIAA1549 Tumor location/resectability (Cerebellar > Non-cerebellar)

DIPG 1q gain, 17p lossH3F3A, HIST1H3B mutations (K27M)

Peds > AdultsDIPG > Non-Brainstem pGBM

pGBM H3F3A mutation

ATRX, DAXX mutationATRX mutation

Peds > AdultsGBM > WHO I,II,III gliomasPeds > AdultsType of H3F3A mutation (G34R/V > K27M)

MacConaill et al. 2009

BRAF V600E in Ganglioglioma in Children

Schindler et al. 2011

Pediatric Ganglioglioma % BRAF V600E

MacConaill et al., 2009 57% (8/14)

Dougherty et al., 2010 50% (9/18)

Schindler et al., 2011 13% (3/24)

Total 36% (20/56)

Pediatric PXA

Schindler et al., 2011 78% (28/36) Dias-Santagata et al, 2011 57% (4/7) Total 74% (32/43)

BRAF V600E in Ganglioglioma and Pleomorphic Xanthoastrocytoma in Children < 18 years

• Initially described in melanoma, colon and papillary thyroid carcinoma• Vemurafenib (“V600E mutated BRAF inhibitor”)

– FDA approved for late-stage or unresectable melanoma (Aug 2011)

Tumor type Molecular phenotype

Association

Ganglioglioma BRAF V600E Peds > Adults

PXA BRAF V600E Peds = AdultsMesenchymal histology

Pilocytic astrocytoma

BRAF-KIAA1549 Tumor location/resectability (Cerebellar > Non-cerebellar)

Oligodendroglioma

1p/19q codeletion Adults > Peds

DIPG 1q gain, 17p loss

H3F3A, HIST1H3B mutations (K27M)

Peds > Adults

DIPG > Non-Brainstem pGBM

pGBM H3F3A mutation

ATRX, DAXX mutation

ATRX mutation

Peds > AdultsGBM > WHO I,II,III gliomas

Peds > Adults

Type of H3F3A mutation (G34R/V > K27M)Unlike in adults, EGFR amplification, PTEN deletion, IDH1 mutations

are rarely observed in pGBM.

`

Schwartzentruber et al., 2012

H3F3A mutations are exclusive to high grade tumors and occur in the pediatric setting.

H3F3A, ATRX, and DAXX mutations distinguish pediatric from adult GBM.

Tumor type Molecular phenotype

Association

Ganglioglioma BRAF V600E Peds > Adults

PXA BRAF V600E Peds = AdultsMesenchymal histology

Pilocytic astrocytoma

BRAF-KIAA1549 Tumor location/resectability (Cerebellar > Non-cerebellar)

Oligodendroglioma

1p/19q codeletion Adults > Peds

DIPG 1q gain, 17p loss

H3F3A, HIST1H3B mutations (K27M)

Peds > Adults

DIPG > Non-Brainstem pGBM

pGBM H3F3A mutation

ATRX, DAXX mutation

ATRX mutation

Peds > AdultsGBM > WHO I,II,III gliomas

Peds > Adults

Type of H3F3A mutation (G34R/V > K27M)H3F3A is located on chromosome 1q, a region of large-scale chromosomal gain in DIPG.

Adults Peds

Epigenome MGMT promoter methylation in adult GBM (associated with pseudoprogression s/p TMZ/RT and improved survival)

H3.3-ATRX-DAXX chromatin remodeling pathway in pGBM

Chromosome

1p/19q codeletion in adult OGD

1q gain, 17p loss in DIPG

Gene Adult AA with EGFR amplification behaves like GBM

EGFR amp, PTEN del are rare in pGBM/DIPG

PDGFRA amp in DIPG

Nucleotide IDH1 mutation (R132H) in secondary GBM and LGG in adults

BRAF mutation (V600E) in ganglioglioma and PXA

Protein BRAF-KIAA1549 in cerebellar PAPARP-1 overexpression in DIPG

Tumor-derived Exosomes

• Exosomes are small membrane vesicles (30-100 nm) derived from luminal membranes of multivesicular bodies and released constitutively by fusion with cell membrane

• Released from tumor cells, exosomes mediate local and systemic cell communication through horizontal transfer of information, such as mRNA, miRNA, proteins, DNA

Electron microscopy of exosomes derived from U87 cells

Research questions:1. Can exosomes in peripheral blood of glioma patients serve as biomarkers of tumor progression? 2. Can exosomes in patient plasma reliably predict parent tumor mutational status in brain?

-IDH1 R132H mutation-BRAF V600E mutation

gDNA exoDNA- - ++DNase

3kb10kb

1kb

DNA is present in tumor cell derived exosomes. ExoDNA exists predominantly as methylated, single stranded DNA

Courtesy of Haiying Zhang and David Lyden

exoDNA-1+S1 nuclease

3kb10kb

1kb

-exoDNA-2

+ - gDNA

+ -

exoDNA

gDNA

Anti-5’metCytosine Anti-DNA

Acknowledgements

Neurosurgery• Jeffrey Greenfield, MD,

PhD• Michael Kaplitt, MD, PhD• Philip Stieg, MD, PhD

Pediatrics• David Lyden, MD, PhD

Neurosurgery• Philip Gutin, MD

Pathology• Jason Huse, MD, PhD

Neuroradiology• Andrei Holodny, MD