Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2017 Jan 10;8(2):2104-2113.
doi: 10.18632/oncotarget.12881.

Droplet digital PCR is a powerful technique to demonstrate frequent FGFR1 duplication in dysembryoplastic neuroepithelial tumors

Frédéric Fina  1 , Doriane Barets  2 , Carole Colin  3 , Corinne Bouvier  2   3 , Laëtitia Padovani  4 , Isabelle Nanni-Metellus  1 , L'Houcine Ouafik  1 , Didier Scavarda  5 , Andrey Korshunov  6 , David T W Jones  6 , Dominique Figarella-Branger  2   3
Affiliations

Droplet digital PCR is a powerful technique to demonstrate frequent FGFR1 duplication in dysembryoplastic neuroepithelial tumors

Frédéric Fina et al. Oncotarget. .

Abstract

Dysembryoplastic neuroepithelial tumors (DNT) share V600E mutation in the BRAF gene with other low grade neuroepithelial tumors (LGNTs). FGFR1 internal tandem duplication of the tyrosine-kinase domain (FGFR1-ITD), another genetic alteration that also leads to MAP kinase pathway alteration, has been previously reported in LGNTs by whole-genome sequencing. In the present study we searched for FGFR1-ITD by droplet digital PCR (DDPCR™) and for FGFR1 point mutations by HRM-sequencing in a series of formalin-fixed paraffin-embedded (FFPE) LGNTs including 12 DNT, 2 oligodendrogliomas lacking IDH mutation and 1p/19q co- deletion (pediatric-type oligodendrogliomas; PTOs), 3 pediatric diffuse astrocytomas (PDAs), 14 gangliogliomas (GGs) and 5 pilocytic astrocytomas (PAs). We showed by DDPCR™ that 5/12 DNT, but none of the other LGNTs, demonstrated FGFR1-ITD. In addition, these cases also accumulated phosphorylated-FGFR1 protein as shown by immunohistochemistry. FGFR1G539R point mutation was only recorded in one DNT that also showed FGFR1-ITD. Interestingly, these FGFR1 alterations were mutually exclusive from BRAFV600E mutation that was recorded in 13 LGNTs (3 DNTs, 1 PTO, 2 PDAs, 5 GGs and 2 PAs). Therefore, FGFR1 alteration mainly represented by FGFR1-ITD is a frequent event in DNT. DDPCR™ is an easy and alternative method than whole-genome sequencing to detect FGFR1-ITD in FFPE brain tumors, in routine practice.

Keywords: FGFR1; MAP kinase pathway; droplet digital PCR (DDPCR™); dysembryoplastic neuroepithelial tumor (DNT); low grade neuroepithelial tumor (LGNT).

PubMed Disclaimer

Conflict of interest statement

CONFLICTS OF INTEREST

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
(A) Determination of copy number variation states of FGFR1 exon 16 compared to FGFR1 exon 8 for the 36 FFPE cases and 6 frozen additional samples (*), the placenta sample (normal control) and the positive control (PA89). When replicates were done, CNV value was the mean of replicates (additional table). Otherwise, CNV measurement was performed from a single DDPCR™ well of around 15 000 droplets. Error bars indicate the Poisson 95% confidence intervals for each DNA copy number determination. For the cases (#PTO01, #GG30; #GG04) without duplication but with low CNV value (that could demonstrate a deletion for FGFR1) it is worth noting that the DNA level was particularly low, making conclusion difficult. (B) Determination of the absolute quantification of FGFR1 exon 16 (grey triangle) and FGFR1 exon 8 (black square) in copies/μl for the 36 FFPE cases and 6 frozen additional samples (*), the placenta sample (normal control) and the positive control (PA89). Error bars indicate the Poisson 95% confidence intervals for each DNA copy number determination. Abbreviations: DNTs: dysembryoplastic neuroepithelial tumors; GGs: gangliogliomas, PAs: pilocytic astrocytomas; PDAs: pediatric diffuse astrocytomas; PTOs: pediatric-type oligodendrogliomas;
Figure 2
Figure 2. Phospho-FGFR1 protein expression detected by immunohistochemistry on two DNT cases: the immunoreactivity was restricted to the glial compartment (A and B) especially the oligodendroglial-like cells in the glial nodule (A) and the GNE (B).
Floating neurons were negative (B, arrows), as well as cortex (C). Scale bars: 50 μm.
Figure 3
Figure 3. Schematic representation of the normal and the duplicated FGFR1 gene
Figure 4
Figure 4
(A) Distribution of the CNV values for the 36 FFPE samples studied the 19 cancer cell lines, the placenta sample (normal control) and the PA89 positive control. The CNV distribution shows very few intermediate values, reflecting the powerful capacity of the droplet digital PCR to discriminate duplicated samples (2.75-3) from non-duplicated samples (1.75-2.25). The cut-off value of the CNV reflecting FGFR1 duplication that we chose was CNV > 2.25. (B) Distribution of the CNV min values for the 36 FFPE samples studied, the 19 cancer cell lines, the placenta sample (normal control) and the PA89 positive control. The cut-off value of the CNV min reflecting FGFR1 duplication that we chose was CNV min > 2.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Daumas-Duport C, Scheithauer BW, Chodkiewicz JP, Laws ER, Jr, Vedrenne C. Dysembryoplastic neuroepithelial tumor: a surgically curable tumor of young patients with intractable partial seizures. Report of thirty-nine cases Neurosurgery. 1988;23:545–556. - PubMed
    1. Fernandez C, Girard N, Paz Paredes A, Bouvier-Labit C, Lena G, Figarella-Branger D. The usefulness of MR imaging in the diagnosis of dysembryoplastic neuroepithelial tumor in children: a study of 14 cases. AJNR Am J Neuroradiol. 2003;24:829–834. - PMC - PubMed
    1. Louis DN, Perry A, Reifenberger G, von Deimling A, Figarella-Branger D, Cavenee WK, Ohgaki H, Wiestler OD, Kleihues P, Ellison DW. The 2016 World Health Organization Classification of Tumors of the Central Nervous System: a summary. Acta Neuropathol. 2016;131:803–820. - PubMed
    1. Louis DN, Ohgaki H, Wiestler OD, Cavenee WK, Ellison DW, Figarella-Branger D, Perry A, Reifenberger G, von Deimling A. (2016) World Health Organization Classification of Tumours of the Central Nervous System. (Lyon: International Agency for Research on Cancer (IARC)) - PubMed
    1. Padovani L, Colin C, Fernandez C, Maues de Paula A, Mercurio S, Scavarda D, Frassineti F, Adelaide J, Loundou A, Intagliata D, Bouvier C, Lena G, Birnbaum D, et al. Search for distinctive markers in DNT and cortical grade II glioma in children: same clinicopathological and molecular entities? Curr Top Med Chem. 2012;12:1683–1692. - PubMed

MeSH terms

Substances

-