Molecular Characterization of A Growth-Factor Induced Glioblastoma Model Open Access

Hannikainen, Paavali (Spring 2018)

Permanent URL: https://etd.library.emory.edu/concern/etds/q524jn79p?locale=en
Published

Abstract

 

The development of therapeutics for the treatment for glioblastoma (GBM) have not been successful, with clinical trials failing by phase III. These failures in finding treatments can be attributed to the lack of an accurate preclinical animal model in which to test therapeutics. Specifically, orthotopic xenograft models do not recapitulate the heterogeneity, growth characteristics, and genetic make-up of human GBMs. Also, syngeneic models have issues with reproducibility and immunogenicity, where the model’s immune system attacks the tumor. Thus, there is a need for a better GBM model. Research on GBM pathophysiology has shown that platelet-derived growth factor (PDGF) is a key protein for GBM tumorigenesis. A viral vector expressing PDGF was previously engineered and investigated. This viral vector can infect glial cells and cause the formation of tumors in the brain parenchyma with histological features that match the human GBM. However, it is unclear if the tumors also have similar molecular characteristics to human GBM. Tyrosine kinase receptors, specifically platelet-derived growth factor receptor-alpha (PDGFR-α) and epidermal growth factor receptor (EGFR), have been extensively investigated in human GBM, in addition to p53 and isocitrate dehydrogenase-1 (IDH-1). To validate this model as an accurate preclinical testing platform, the ability to replicate previous results and re-establish this model must be shown. In addition, further characterization of PDGF-induced tumor model must be completed through investigating PDGFR-α, EGFR, p53, and IDH-1 expression in PDGF-induced tumors. The results show successful reproduction of PDGF-induced tumors, as evidenced by staining showing typical histopathologic features of GBM, most notably pseudopalidasing necrosis. In addition, characterization studies of PDGFR-α, EGFR, p53, and IDH-1 show that the model might not successfully recapitulate the molecular characteristics of GBM tumors. 

Table of Contents

 

Introduction……………………………………………………………………………………..1-6

Glioblastoma Multiforme and Failures in Treatment………………………………………1

Virally-Mediated Gene Delivery…………………………………………………………..2

Genetic Alterations in Human GBM………………………………………………………2

Platelet-Derived Growth Factor (PDGF) and Receptors………………………………….3

Epidermal Growth Factor Receptor (EGFR) in GBM…………………………………….4

p53 in GBM……………………………………………………………………………….4

Isocitrate Dehydrogenase 1 (IDH-1) in GBM…………………………………………….5

Hypotheses………………………………………………………………………………..6

Materials and Methods………………………………………………………………………...7-11

            Retrovirus Packaging……………………………………………………………………...7

            Rat Surgeries………………………………………………………………………………7

            Brain Tissue Collection……………………………………………………………………8

Hematoxylin and Eosin Staining (H&E) ………………………………………………….8

Moving from H&E Characterization to Immunohistochemical Characterization…………9

Immunohistochemistry……………………………………………………………………9

Microscopy………………………………………………………………………………10

Results and Analysis………………………………………………………………………….12-14

Hematoxylin and Eosin Stain Analysis…………………………………………………..12

Immunohistochemistry Analysis………………………………………………………...12

Tumor Size and Viral Expression………………………………………………………..14

 

Discussion…………………………………………………………………………………….15-18

Re-establishing Virally-Induced PDGF Tumor Model…………………………………..15

Tyrosine Kinase Receptors in PDGF-Induced Tumors………………………………….16

IDH-1 and p53 in PDGF-Induced Tumors………………………………………………17

PDGF-Induced Tumor Model as a Preclinical Testing Platform………………………...18

Figures and Tables…………………………………………………………………………3, 19-26

Figure 1: Tyrosine Kinase Receptor Signaling Cascade…………………………………..3

Figure 2: H&E Images of PDGF-Induced GBM in Rat Brain…………………………...19

Figure 3: Overexpression of PDGFR-α in PDGF-Induced Glioblastoma in Rat

    Brain…………………………………………………………………………...20

Figure 4: Decreased Expression of EGFR in PDGF-Induced Glioblastoma in Rat

Brain……………………………………………………………………………...21

Figure 5: Decreased Expression of p53 in PDGF-Induced Glioblastoma in Rat Brain….22

Figure 6: Inconclusive Regulation of IDH-1 in PDGF-Induced Glioblastoma in Rat

    Brain…………………………………………………………………………...23

Figure 7: Positive Immunohistochemistry Controls……………………………………..24

Figure 8: DsRED Expression in PDGF-Induced Glioblastoma in Rat Brain……………25

Table 1: Tumor Analysis Overview……………………………………………………...26

References……………………………………………………………………………………27-30

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