| dc.description.abstract | Pediatric high-grade glioma (pHGG) accounts for 3-15% of all primary pediatric central nervous system tumors and is the most aggressive pediatric brain tumor. The standard therapy for pHGG includes maximal safe tumor resection followed by radiation therapy and intensive chemotherapy using oral temozolomide (TMZ). Despite aggressive therapy, the prognosis is still poor and the five-year survival rate is dismal (< 5%). Thus, there is an urgent need to identify effective and efficient combination therapies for treatment of pHGG. The objective of this study is to develop an unbiased, target-based approach to the discovery of novel combination therapies for pHGG. We are using pHGG cell lines from patient-derived orthotopic xenografts (PDOX) growing both as 3D neurospheres and 2D monolayers. The neurospheres are thought to represent predominantly cancer-based stem cells. For drug screening, we use 2-step strategy, first carrying out a single agent screen with a library of cancer-related therapeutics. We use the single agent screen to identify classes of pharmacologic agents with growth inhibitory/cytotoxic activity against the PDOX pHGG cells. Combinations of prototypic agents from the major classes of active drugs are then tested for synergistic activity in the second stage of combinatorial screening studies. The first stage single agent screening studies, carried out with 1863 well-characterized cancer therapeutics, identified several classes of active compounds including HDAC inhibitors, proteasome inhibitors, HSP90 inhibitors, PI3K/mTOR/Akt inhibitors and cell cycle inhibitors as the most active classes. We selected a minimum of 2 representatives of each of these classes for combinatorial screens and discovered that combinations of proteasome inhibitors (CEP-18770, MLN 2238) and HDAC inhibitors (Panobinostat, JNJ-26481585) and also a Wee1 inhibitor (MK-1775) and combined Chk1 inhibitors (AZD7762, MK-8776) exhibited the greatest synergistic lethality activity.
Both of these sets of combinations exhibited selective activity, inhibiting the viability of pHGG cells much more than normal astrocytes. Because of their profound synergistic activity and their targeted molecular activity, we focused subsequent mechanistic studies on the combination of Wee1 and Chk1 inhibitors. In pHGG cells, the combination of these two classes of drugs not only selectively suppressed cell growth in the pHGG tumor cells, but it also promoted genomic instability, induced the G2/M cell cycle arrest, slowed down the speed of DNA replication during DNA replication and caused mitotic catastrophe. These effects were selective for pHGG tumor cells and were not detected in normal astrocyte cells. In addition, Wee1 and Chk1 inhibitors had a selective effect on the down-regulation ATP production from mitochondrial respiration in tumor cells but not in astrocytes. In conclusion, these studies have identified the combination of Wee1 and Chk1 inhibitors as selective inducers of lethality in pediatric high-grade glioma cells by a mechanism that includes their combined effects on metabolic inhibition and the induction of mitotic catastrophe. | en |
