Glioblastoma (GBM) is a malignant brain cancer that is resistant to all treatment modalities. This
resistance is due, in large part, to a population of high invasive and low proliferative cancer cells
that elude surgical resection and are refractory to chemotherapy and radiation. While a great
deal is known about oncogenes, tumor suppressors, and other pathways that promote GBM cell
proliferation, we understand relatively little about mechanisms that drive GBM cell invasion in
the brain microenvironment. Therefore, the PI's group performed genetic and biochemical
screens to identify adhesion and signaling factors that regulate invasive cell growth in GBM.
These efforts identified the non-receptor protein tyrosine phosphatase PTP-PEST/PTPN12 as a
critical signaling effector in invasive GBM cells. Here, we present a significant amount of
supporting data showing that PTP-PEST promotes GBM cell invasion by regulating the stability
of key focal adhesion signaling proteins, particularly Crk-associated substrate (p130Cas). In
particular, we have discovered that PTP-PEST in focal adhesions mediates interactions
between p130Cas and valosin containing protein (Vcp), a ubiquitin-dependent segregase and
key component of the ubiquitin proteasome system. These findings have led to our working
hypothesis that PTP-PEST is essential for GBM cell invasion by regulating the phosphorylation-
dependent ubiquitination of focal adhesion protein substrates. To test this hypothesis, we will (1)
characterize protein domains and motifs that mediate interactions between PTP-PEST,
p130Cas, and Vcp, as well as determine how these interactions modulate focal adhesion
protein stability in GBM cells; (2) identify PTP-PEST-generated phosphodegron sequences in
p130Cas and determine how they regulate focal adhesion dynamics by recruiting Vcp and
facilitating p130Cas degradation by the proteasome; (3) genetically mutate Vcp at specific sites
required for the phosphorylation-dependent ubiquitination of p130Cas and analyze cell invasion
using three-dimensional culture systems and pre-clinical mouse models of GBM; and (4)
quantify levels and spatial patterns of PTP-PEST signaling via p130Cas and Vcp in human
GBM samples and primary cancer cell culture systems. We will correlate these data with patient
survival as well as response to therapies such as temozolomide and bevacizumab. Collectively,
these experiments will not only elucidate signaling pathways that control the GBM cell invasive
state, but may lead to new strategies to target invasive cells and block tumor progression.