Armstrong, Brock

Abstract BACKGROUND/PURPOSE The Mayo Clinic has developed a large panel of patient-derived xenografts (PDX) from patients with glioblastoma multiforme (GBM). Here we report initial molecular profiling of the Mayo PDX panel and a comparison of patient and PDX molecular profile and response to therapy. METHODS & MATERIALS Clinical data was retrieved by retrospective chart review. Animal data from all PDX investigations conducted between 2004-2014 were retrieved from experimental logs and subsequently consolidated into a database for analysis. RESULTS From 1999 to 2014, 182 patient tumor samples of varying histological grades were attempted for xenograft with 73 resulting viable PDX lines. Viable xenografts were only produced from WHO grade IV tumor specimens, yielding an overall success rate of 49% for these tumors. GBM patients that produced viable xenografts compared to those that did not exhibited a trend for decreased overall survival (P = 0.18). There was no significant association between successful xenografting and whether tissue was from newly diagnosed (45/93%) or recurrent (20/40; 50%) tumors. Patient age > 45 at diagnosis was correlated with increased PDX viability in GBMs (p = 0.05). Of the viable PDX models analyzed, EGFR mutation was identified in 17 lines, TERT mutation was found in 13 lines, IDH mutation in 1 line, and MGMT hypermethylation in 25 lines. RNAseq was performed on orthotopic tumor samples from 53 PDX models. After excluding contaminating murine sequence reads, expression analysis demonstrated 32 models with a mesenchymal phenotype. Array comparative genomic hybridization was performed on 9 patient samples and derivative early, mid and late passage PDX tumors. Using unbiased hierarchical clustering, there was a high concordance between patient and xenograft models. Within the PDX panel, fractionated radiation (RT) alone and RT combined with temozolomide (TMZ) was tested in orthotopic tumors in 38 lines. The overall median survival benefit (ratio of median survival for treated vs. placebo) in PDX lines treated with RT only was 1.6 (range: 0.9-2.5) and with RT/TMZ was 2.5 (range: 1.1 - 8.9). There was a positive association between observed patient survival and the corresponding survival benefit in the PDX for subjects treated with RT/TMZ (r = 0.2; n = 17). Response to adjuvant TMZ was evaluated in 42 tumor lines, and response to bevacizumab was tested in 33 tumor lines, and correlations with clinical treatment response are being evaluated. CONCLUSIONS: The Mayo GBM PDX panel is widely used in the neuro-oncology community. The initial molecular analysis suggests a good correlation between patient and PDX at the level of genotypic characteristics and therapeutic sensitivity. Citation Format: Dioval A. Remonde, Brett L. Carlson, Mark A. Schroeder, Brock Armstrong, Sen Peng, Lisa Evers, Paul A. Decker, Jeanette Eckel Passow, Michael E. Berens, Nhan L. Tran, Robert B. Jenkins, Jann N. Sarkaria. Characterization of patient-derived xenograft (PDX) models to evaluate clinical and therapeutic responses of glioblastoma multiforme. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1468. doi:10.1158/1538-7445.AM2015-1468

Abstract Glioblastoma multiforme (GBM) is the most common and most lethal primary malignant brain tumor, affecting 25,000 patients per year. Approximately 87% of GBMs have abnormalities in the p53 pathway that affects G2 checkpoint. In the absence of p53, Wee1 kinase has been shown to regulate G2 checkpoint upon chemotherapeutic insult. Wee1 inhibits CDC2 function, thereby promoting G2 phase cell cycle arrest, thus preventing damaged cells from entering into premature mitosis without repairing the DNA. We hypothesize that in p53-deficient tumor cells, inhibitors of Wee1 would be effective to heighten DNA damage due to genetic and pharmacologic loss of both the G1 and G2 checkpoints, respectively. MK-1775 (Merck) is a potent and selective small molecule inhibitor of the Wee1 kinase that has demonstrated enhanced cytotoxic effect in combination with several chemotherapeutic agents in preclinical studies in vitro and in vivo. Here, we selected an array of short term cultures of glioma patient derived xenograft (PDX) models (GBM12, GBM43, GBM79, GBM84) and long term established glioma cell lines (U87, SF767, T98G) with aberrant and wild type G1 checkpoint pathways and investigated cytotoxic effects of MK-1775 as a single agent and in combination with Temozolomide (TMZ). Our results demonstrated differential sensitivity to MK-1775 and TMZ as single agents for different glioma cell lines in vitro. However, no correlation between MK-1775, TMZ sensitivity and p53 status of the cell lines was observed. Combination therapy with MK-1775 and TMZ showed a synergistic effect for one PDX model, GBM43, with combination index (CI) of 0.4. Combination therapy of all other glioma cell lines either showed an additive effect or antagonistic effect with CI of >0.75 irrespective of their p53 status. Our results demonstrated that synergistic effects of MK-1775 in combination with chemotherapy (TMZ) are not solely dependent on p53 status of glioma cells and require a more comprehensivce understanding of the molecular mechanism that dictates GBM's dependence on Wee1 for escaping DNA damage therapy. Supported by the Ben & Catherine Ivy Foundation Citation Format: Harshil D. Dhruv, Andrew D. Nelson, Brock Armstrong, Julie L. Boerner, Jann N. Sarkaria, Nhan L. Tran, Michael Berens. Vulnerability and resistance in glioblastoma to G2-specific drugs. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1319. doi:10.1158/1538-7445.AM2014-1319

Abstract A critical problem in the treatment of malignant gliomas is the extensive infiltration of individual tumor cells into adjacent brain tissues. This invasive phenotype severely limits all current therapies, and to date, no treatment is available to control the spread of this disease. Members of the tumor necrosis factor (TNF) ligand superfamily and their cognate receptors regulate various cellular responses including proliferation, migration, differentiation, and apoptosis. Specifically, the TNFRSF19/TROY gene encodes a type I cell surface receptor that is expressed on migrating or proliferating progenitor cells of the hippocampus, thalamus, and cerebral cortex. Here, we show that levels of TROY mRNA expression directly correlate with increasing glial tumor grade. Among malignant gliomas, TROY expression correlates inversely with overall patient survival. In addition, we show that TROY overexpression in glioma cells activates Rac1 signaling in a Pyk2-dependent manner to drive glioma cell invasion and migration. Pyk2 coimmunoprecipitates with the TROY receptor, and depletion of Pyk2 expression by short hairpin RNA interference oligonucleotides inhibits TROY-induced Rac1 activation and subsequent cellular migration. These findings position aberrant expression and/or signaling by TROY as a contributor, and possibly as a driver, of the malignant dispersion of glioma cells. Mol Cancer Res; 8(11); 1558–67. ©2010 AACR.