Background: Glioblastoma is a fatal malignant primary brain tumour. Standard therapy includes maximal surgical resection, radiotherapy and adjuvant chemotherapy with temozolomide. However, glioblastoma cells can be resistant to adjuvant treatments, therefore tumour recurrence is inevitable resulting to an incurable disease. This clearly reflects the pressing need for more effective treatment strategies. Here, we investigated two drugs: the first-line glioblastoma chemotherapeutic temozolomide, and gemcitabine, a small, brain-penetrant, cytosine analogue; both induce DNA damage and trigger the DNA damage response pathway. Inhibition of the DNA damage response pathway has been shown to sensitise other adult cancers to conventional DNA-damaging chemotherapeutics.
Aim: We explored the hypothesis that an inhibitor of cell cycle checkpoint kinase 1 and 2 (CHK1/2), called LY2606368, will inhibit the DNA damage response following temozolomide- or gemcitabine-induced DNA damage, leading to an accumulation of DNA damage and enhanced glioblastoma cell death.
Methods/Results: Immunoblotting confirmed LY2606368 inhibition of CHK1 in GMB6 patient-derived glioblastoma cells. In vitro drug interaction assays showed that LY2606368 synergised with temozolomide or gemcitabine in murine and human glioblastoma cell lines. Also, when combined with temozlomide or gemcitabine, phosphorylation of CDC2 was reduced while gammaH2AX levels were enhanced. These data suggest that LY2606368 blocks DNA-damage induced cell cycle arrest, leading to an accumulation of DNA damage. This was also observed in vivo, where immunohistochemistry showed that at 24 hours post-treatment, LY2606368 increased the extent of temozolomide- and gemcitabine-induced DNA damage, leading to reduced tumour proliferation. Most-importantly, combination therapy significantly extended survival of multiple orthotopic mouse models of glioblastoma. We also examined LY2606368 combined with radiation which consistently showed that LY2606368 improved the therapeutic efficacy.
Conclusions: This study has established the therapeutic benefit of combining temozolomide or gemcitabine with LY2606368 and provides robust data that is informing the rational design of a potential clinical trial for relapsed glioblastoma patients.