Blocking the Notch Pathway with Gamma-Secretase Inhibitors Enhances Temozolomide Treatment of Gliomas through Therapy-Induced Senescence: A Dissertation

Abstract

Glioma therapy relies on induction of cytotoxicity; however, the current combination of surgery, irradiation (IR) and temozolomide (TMZ) treatment does not result in a long-term cure. Our lab previously demonstrated that a small population of glioma cells enters a transient cell cycle arrest in response to chemotherapy. Treatment with TMZ significantly decreases initial neurosphere formation; however, after a short recovery period, a small number of cells resume neurosphere formation and repopulate the culture. This recovery of neurosphere growth recapitulates the inevitable glioma recurrence in the clinic. The focus of our laboratory is to study direct-target therapies that can be combined with TMZ to inhibit neurosphere recovery. The Notch pathway is a promising target because it is involved in cell growth and survival. Here, we demonstrate that blocking the Notch pathway using gamma-secretase inhibitors (GSIs) enhances TMZ treatment. The combination of TMZ and GSI treatments targets the cells capable of recovery. TMZ + GSI treated cells do not recover and are no longer capable of self-renewal. Interestingly, recovery is inhibited when the GSI is administered 24 hrs after TMZ treatment, demonstrating a sequence-dependent mechanism. TMZ + GSI treatment also decreases tumorigenicity. When glioma cell lines were treated in vitro and implanted in NU/NU nude 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. Cultures treated with TMZ + GSI have decreased proliferation, but no increase in cell death. We observed an increase in the number of cells expressing senescence-associated β-galactosidase in vitro and in vivo. This demonstrates that inhibition of the Notch pathway shifts TMZ-treated cells from a transient cell cycle arrest into a permanent senescent state. Senescent cells can stimulate the innate immune system. Here we demonstrate that TMZ + GSI treatment increases phagocytosis in vitro. New therapy combinations, such as TMZ + GSI, are arising in the field of therapy-induced senescence (TIS). 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 a promising target-directed therapy to decrease the rate of brain tumor recurrence by inducing senescence and tumor clearance.