Abstract 963: Glioma treatment with temozolomide and Notch inhibition blocks tumor recovery through the induction of senescence and tumor clearance
Authors: Gilbert Candace A. , Ramirez Yulian P. , Weatherbee Jessica L. , Pierre Christine A. St. , Moser Richard P. , Kurt-Jones Evelyn A. , Ross Alonzo H. 
Published: June 2012

Abstract
Current glioma therapy relies on induction of cytotoxicity after removal of the bulk tumor through the combination of surgery, radiation and temozolomide (TMZ); however, these therapies do not result in a long-term cure. Our lab previously demonstrated that some glioma cells undergo a transient cell cycle arrest in response to chemotherapy. Treatment with TMZ decreases sphere formation; however, after a short recovery period, a small number of cells resume sphere formation and self-renewal, measured by secondary sphere formation. Blocking the Notch pathway in neurosphere cultures with gamma-secretase inhibitors (GSIs) after TMZ treatment targeted the cells capable of recovery. TMZ + GSI treated cells do not recover and are no longer capable of self-renewal. TMZ + GSI synergy is dependent on the sequence of the drug treatments. Recovery was inhibited when GSI was administered 24 hrs after TMZ treatment. TMZ + GSI treatment also decreases tumorigenicity. When glioma cell lines were treated in vitro and implanted in immunodeficient mice, TMZ + GSI treatment extended latency and greatly increased survival. In addition, in vivo TMZ + GSI treatment completely blocked tumor progression and resulted in the loss of a palpable tumor in 50% of mice, while none of the TMZ-only treated mice survived. TMZ + GSI treated cultures and xenografts display a senescent phenotype. We observed an increase in the number of cells expressing senescence-associated beta-galactosidase and a decrease in Ki67 positive cells. Gene expression was also analyzed after drug treatments to confirm the induction of senescence. p21 is upregulated in cells that have undergone either a transient cell cycle arrest or senescence. We found that upregulation of p21 occurred initially in both TMZ-only and TMZ + GSI treatments, but only remained upregulated in the TMZ + GSI samples. This demonstrates that the addition of GSIs shifts TMZ-treated cells from a transient arrested state to a permanent senescent state. New therapy combinations, such as TMZ + GSI, are arising in a promising new field of cytostatic therapy and therapy-induced senescence (TIS). A key feature of TIS is the secretory profile of senescent cells. It was previously demonstrated that senescent tumor cells secrete inflammatory cytokines and activate the innate immune system for tumor clearance. We found that TMZ + GSI treatment resulted in upregulation of secreted IL-6 and IL-8 cytokines. We are currently the effect of senescent glioma cells on the innate immune system and tumor clearance. Overall, this data demonstrates the importance of the Notch pathway in chemoprotection and maintenance of TMZ-treated gliomas. The addition of GSIs to current treatments is promising target-directed therapy to decrease the rate of brain tumor recurrence inducing senescence.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 963. doi:10.1158/1538-7445.AM2011-963