Institute of Biomedical Sciences

Title

Liquid biopsy for detection and monitoring of driver mutations in Diffuse Intrinsic Pontine Gliomas (DIPGs)

Poster Number

17

Document Type

Poster

Keywords

Liquid biopsy, pediatric brain tumor, histone 3, diffuse intrinsic pontine glioma, digital droplet PCR

Publication Date

4-2017

Abstract

Background: Diffuse Intrinsic Pontine Glioma (DIPG) is a deadly pediatric brain cancer that makes up 10-15% of all central nervous system (CNS) tumors in children. Surgery is not an option due to its anatomical location and infiltrative nature. It is amongst the most challenging tumors to treat. Combination of chemotherapy and focal radiation therapy is the primary therapy for DIPG, but the benefits of the radiation therapy is temporary. Immunotherapy is a technique that is gaining more interest in CNS tumors. Identification of tumor associated antigens is one of many requirements in developing an effective immunotherapy.

Objective: To validate Wilms' tumor protein (WT1) as a potential tumor associated antigen in DIPGs using patient derived cell lines and patient formalin fixed paraffin embedded (FFPE) specimens. Methods/Design: DIPG patient FFPE specimens were immunohistochemically stained for WT1 using mouse monoclonal anti-WT1 antibody. FFPE specimens and patient derived cell lines were also co-stained for K27M histone H3 protein mutation (p.Lys27Met) and WT1 using immunofluorescent (IF) staining methods. The fluorescence intensity was measured to compare WT1 level in the two histone mutation subgroups of DIPG (H3.3K27M and H3.1K27M). In addition, frozen tumor tissues and patient derived cell lines were used to validate WT1 protein levels in western blot.

Results/Discussion: Immunohistochemistry (IHC) staining of patient FFPE specimens showed strong WT1 immunoreactivity in tumor compared to adjacent normal tissue. In addition, our IHC staining showed weak to absent WT1 immunoreactivity in H3.1K27M subtype tumor specimens compared to strong to moderate in H3.3K27M subtype tumor specimens. IF staining of cell lines confirm this differential WT1 levels in the subtypes. Western blot of tumor tissues and cell lines were performed to further validate WT1 levels. These results suggest that WT1 is a potential DIPG tumor associated antigen which can be utilized for immunotherapy.

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Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 4.0 License.

Open Access

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Comments

To be presented at GW Annual Research Days 2017.

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Liquid biopsy for detection and monitoring of driver mutations in Diffuse Intrinsic Pontine Gliomas (DIPGs)

Background: Diffuse Intrinsic Pontine Glioma (DIPG) is a deadly pediatric brain cancer that makes up 10-15% of all central nervous system (CNS) tumors in children. Surgery is not an option due to its anatomical location and infiltrative nature. It is amongst the most challenging tumors to treat. Combination of chemotherapy and focal radiation therapy is the primary therapy for DIPG, but the benefits of the radiation therapy is temporary. Immunotherapy is a technique that is gaining more interest in CNS tumors. Identification of tumor associated antigens is one of many requirements in developing an effective immunotherapy.

Objective: To validate Wilms' tumor protein (WT1) as a potential tumor associated antigen in DIPGs using patient derived cell lines and patient formalin fixed paraffin embedded (FFPE) specimens. Methods/Design: DIPG patient FFPE specimens were immunohistochemically stained for WT1 using mouse monoclonal anti-WT1 antibody. FFPE specimens and patient derived cell lines were also co-stained for K27M histone H3 protein mutation (p.Lys27Met) and WT1 using immunofluorescent (IF) staining methods. The fluorescence intensity was measured to compare WT1 level in the two histone mutation subgroups of DIPG (H3.3K27M and H3.1K27M). In addition, frozen tumor tissues and patient derived cell lines were used to validate WT1 protein levels in western blot.

Results/Discussion: Immunohistochemistry (IHC) staining of patient FFPE specimens showed strong WT1 immunoreactivity in tumor compared to adjacent normal tissue. In addition, our IHC staining showed weak to absent WT1 immunoreactivity in H3.1K27M subtype tumor specimens compared to strong to moderate in H3.3K27M subtype tumor specimens. IF staining of cell lines confirm this differential WT1 levels in the subtypes. Western blot of tumor tissues and cell lines were performed to further validate WT1 levels. These results suggest that WT1 is a potential DIPG tumor associated antigen which can be utilized for immunotherapy.