Despite the many therapeutic strategies undertaken for treatment of glioblastoma multiforme, the survival rate remains low for patients afflicted with this aggressive cerebral malignancy. Even with the combined use of several therapeutic modalities, a good prognosis is extremely rare as the remaining cancer cells inevitably infiltrate the normal brain tissue and cause tumor recurrence. Local invasion remains an important cause of morbidity and mortality from brain tumors. The long-term objective of my laboratory is to understand the cellular and molecular mechanisms that underlie tumor invasiveness in human gliomas. Radiotherapy remains a major component of treatment modalities for controlling both malignant and benign glioblastomas. In patients with residual or recurring benign tumors, there is increasing concern about radiation-related side effects that may occur even with highly accurate therapies such as radiosurgery. Despite some therapeutic effect, recent evidence has shown that irradiation may promote malignant behaviors of cancer cells both in vitro and in vivo by activating several pathways involved in tumor invasiveness, angiogenesis and metastasis. Further, the glioma initiating cells are resistant to radiotherapy. Several studies, including ours, demonstrated that significantly increased levels of cath B and uPAR in malignant gliomas and radiation further activates cath B and uPAR molecules. We hypothesize that the cath B and uPAR gene will be targeted while simultaneously combined with radiation and their anti-cancer effects will be determined.
The specific aims of the proposal are:
Specific Aim 1. Evaluate the effect of vectors expressing siRNA targeting cathepsin B and uPAR (pCU) on glioma cell adhesion, migration and invasion in in vitro models with and without radiation.
Aim 1 a. Determine the effect of pC, pU, pCU and radiation alone and in combination on the levels of cath B and uPAR in glioblastoma cell lines and glioma initiating cells (GIC).
Aim 1 b. Determine the downregulation of cath B and uPAR on glioma cell adhesion and migration in glioblastoma cell lines and glioma initiating cells with and without radiation.
Aim 1 c. Evaluate the effect of pC, pU, pCU and radiation alone and in combination on the invasive behavior of human glioblastoma cell lines and GIC.
Specific Aim 2. Determine the molecular mechanisms in cath B and uPAR inhibition-mediated changes on cell growth, proliferation and apoptosis in GIC and non-GIC with and without radiation.
Aim 2 a. Evaluate the molecular mechanisms in cath B and uPAR downregulation on cell growth and proliferation with and without radiation in non-GIC and GIC.
Aim 2 b. Determine the effect of cath B and uPAR downregulation on the molecular mechanisms of apoptosis in glioblastoma cell lines and GIC with and without radiation.
Specific Aim 3. Determine the molecular mechanisms in cath B and uPAR inhibition-mediated changes in angiogenesis and tumor growth in non-GIC and GIC with and without radiation in nude mice.
Aim 3 a. Evaluate the effect of cath B and uPAR downregulation-mediated changes in the signaling mechanisms in the regulation of cerebral angiogenesis both in vitro and in vivo with and without radiation.
Aim 3 b. Determine the optimal doses of pC, pU, pCU and radiation alone and in combination on pre-established intracranial tumor growth or the invasiveness of human glioblastoma cell lines and GIC injected intracerebrally in nude mice. The proposed studies should generate major insight into the pathogenesis of cath B and uPAR and, in turn, should suggest novel targets for therapeutic interventions of glioblastomas.
Despite the many therapeutic strategies undertaken for treatment of glioblastoma multiforme, the survival rate for patients afflicted with this aggressive cerebral malignancy remains low. Radiotherapy has remained the single most effective therapy of glioblastoma for more than 25 years. However, an increasing number of long survivors with secondary side effects and the resistance of glioma initiating cells from this treatment highlighted the need for development of novel therapeutic approaches. This proposal represents a combinational therapeutic approach using a single and bicistronic siRNA construct for cathepsin B and uPAR. This strategy may improve radiotherapy outcomes for the treatment of glioblastomas.
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