Document Type

Dissertation

Date of Degree

Summer 2013

Degree Name

PhD (Doctor of Philosophy)

Degree In

Pharmacy

First Advisor

Mahfoud Assem

Abstract

Poor prognosis and resistance to therapy in malignant gliomas is mainly due to highly dispersive nature of glioma cells. Aggressive infiltration in the brain parenchyma poses a serious challenge to complete tumor resection and effective treatment. Moreover, absence of valid biomarkers confounds definitive diagnosis and therapy response prediction. Identification of novel markers will require improved understanding of glioma genetics, particularly as it relates to key regulatory signals that control glioma cell migration. Since, these tumors are genetically heterogeneous, genomic explorations could improve understanding of molecular mechanisms underlying glioma migration and aid in discovery of molecular markers of clinical impact. Thus the central aim of our study is to identify novel genetic markers in patients with glioma for better prognosis, diagnosis and prediction of response to treatment.

In this study we screened for genome wide copy number alterations and loss of heterozygosity in a representative glioma population of sixty patients. Alterations were detected at multiple chromosomal loci with putative tumor suppressor genes and oncogenes. In relation to molecular determinants of glioma cell migration, alterations at the Protein Tyrosine Phosphatase Receptor type Kappa (PTPRK) locus were the most frequent in our glioma population. PTPRK alterations are relevant to glioma biology as PTPRK is a cell adhesion molecule that is highly expressed in brain. However, function of PTPRK has not been described previously in gliomas. This led to our central hypothesis that PTPRK is a novel biomarker that suppresses diffusive capacity of glioma cells and thereby may improve glioma therapeutic outcome.

Overexpression and knockdown experiments were performed to study PTPRK function using malignant glioma cell lines. We explored molecular mechanisms underlying PTPRK function and its effect on response to therapy. Moreover, we discovered novel PTPRK mutations by sequencing full length PTPRK transcripts in numerous glioma biopsies. Effect of these mutations on PTPRK function and response to anti-glioma therapeutics were subsequently analyzed using in vitro cell based assays.

Our results suggest that PTPRK is an independent prognostic marker and a glioma tumor suppressor that is altered at both transcriptional and post-translational levels. Proteolytic processing of transmembrane PTPRK protein generates a series of fragments which are the only detectable PTPRK forms in glioblastoma. Short hairpin RNA (shRNA) mediated downregulation of PTPRK reduces glioblastoma cell migration suggesting that proteolysis of PTPRK contribute to migration of glioblastoma cells. Additionally, overexpression of wild-type PTPRK suppresses growth and migration of malignant glioma cells which correlates with inhibition of EGFR and beta-catenin signaling, and improves effect of conventional therapies for glioma. However, PTPRK mutations abrogate tumor suppressive effects of wild-type PTPRK and alter sensitivity of glioma cells to chemotherapy.

These results indicate that the cell surface PTPRK protein is a tumor suppressor of prognostic and predictive value which is frequently inactivated in malignant gliomas.

Pages

132 pages

Comments

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Copyright

Copyright 2013 Supreet Agarwal

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