Overcoming treatment resistance in glioblastoma multiforme by tumour specific inhibition of DNA repair.
Lead Research Organisation:
University of Sussex
Department Name: Brighton and Sussex Medical School
Abstract
At the moment, most malignant brain tumours are resistant to treatment and patients live for an average of only one year. The aim of our research is to improve life expectancy for these patients by increasing the effects of radiotherapy and chemotherapy on the tumours without increasing damage to healthy tissues. Our previous work with drugs called PARP inhibitors has been promising, and we have planned a clinical trial to see if patients benefit. One aim of this project is to identify which patients benefit from PARP inhibitors, and to understand why. We will also investigate whether PARP inhibitors increase the ability of radiotherapy to eradicate the cells that are probably responsible for most cases of tumour recurrence. If PARP inhibitors are not effective we will test other drugs that affect the way cells respond to radiotherapy. Finally, we will investigate a possible new treatment for a type of brain tumour that is extremely resistant to chemotherapy. The new treatment appears to reduce levels of a protein called MGMT which otherwise protects tumour cells from chemotherapy. We need to show that the new drug increases killing of brain tumour cells without causing extra damage to healthy tissues.
Technical Summary
Aims
To identify and characterise novel, effective therapies for treating glioblastoma multiforme (GBM) by tumour-specific inhibition of DNA repair.
Objectives
(1) Demonstrate whether inhibitors of DNA repair proteins increase radiosensitivity of human glioma stem cells. (2) In vivo analysis of the effects of PARP inhibitors on DNA repair in GBM and normal tissues, alone and in combination with radiation and/or chemotherapy. Identification of biomarkers predictive of response to PARP inhibitors. (3) Establish whether tumour-specific downregulation of O6-methylguanine methyltransferase (MGMT) by modulation of PI3kinase/mTOR signalling overcomes temozolomide resistance of human glioma cells with unmethylated MGMT gene promoters.
Design and methodology
(1) Analysis of effects of inhibitors of PARP, DNA-PKcs, ATM, Chk1 and Chk2 on radiation responses of human glioma stem cells, measured by clonogenic survival, induction and repair of DNA damage (immunofluorescence and comet assays) and cell cycle parameters. Active agents will be tested in vivo by quantifying effects on tumorigenicity of irradiated glioma stem cells implanted into the forebrains of SCID mice.
(2) Immunohistochemical analysis of induction and repair of DNA double strand breaks, measured by H2AX, 53BP1 and Rad51 foci, in tumour specimens and hair follicles of patients in early phase clinical trials of the PARP inhibitor AZD2281 +/- radiotherapy and/or temozolomide. Dual staining will address cell cycle phase specificity. Repair parameters will be correlated with progression-free and overall survival.
(3) Measurement of effects of inhibitors of p38MAPkinase, mTOR or ERK signalling on MGMT gene and protein expression and temozolomide sensitivity of human glioma cells and glioma stem cells.
Scientific opportunities
Studies on glioma stem cells will verify or refute previous observations that they are radioresistant, and elucidate mechanisms underlying this phenotype. Translational studies using tumour resection specimens will interrogate in vivo effects of PARP inhibitors on DNA repair processes in GBM and normal tissues treated with radiotherapy and chemotherapy. Understanding the mechanisms underlying effects of PI3kinase/mTOR inhibition on MGMT expression will provide novel insights into interactions between oncogenic signalling pathways and protein translation, and elucidate the key factors regulating activity of this DNA repair protein.
Medical opportunities
All aspects of this proposal are aimed at improving treatment of GBM. Overcoming radioresistance of glioma stem cells is likely to be necessary to achieve tumour cure; elucidating mechanisms of sensitisation by PARP inhibitors in vivo and identifying predictive biomarkers will define their therapeutic role; and tumour-specific downregulation of MGMT may improve outcomes in GBM that are resistant to existing therapies.
To identify and characterise novel, effective therapies for treating glioblastoma multiforme (GBM) by tumour-specific inhibition of DNA repair.
Objectives
(1) Demonstrate whether inhibitors of DNA repair proteins increase radiosensitivity of human glioma stem cells. (2) In vivo analysis of the effects of PARP inhibitors on DNA repair in GBM and normal tissues, alone and in combination with radiation and/or chemotherapy. Identification of biomarkers predictive of response to PARP inhibitors. (3) Establish whether tumour-specific downregulation of O6-methylguanine methyltransferase (MGMT) by modulation of PI3kinase/mTOR signalling overcomes temozolomide resistance of human glioma cells with unmethylated MGMT gene promoters.
Design and methodology
(1) Analysis of effects of inhibitors of PARP, DNA-PKcs, ATM, Chk1 and Chk2 on radiation responses of human glioma stem cells, measured by clonogenic survival, induction and repair of DNA damage (immunofluorescence and comet assays) and cell cycle parameters. Active agents will be tested in vivo by quantifying effects on tumorigenicity of irradiated glioma stem cells implanted into the forebrains of SCID mice.
(2) Immunohistochemical analysis of induction and repair of DNA double strand breaks, measured by H2AX, 53BP1 and Rad51 foci, in tumour specimens and hair follicles of patients in early phase clinical trials of the PARP inhibitor AZD2281 +/- radiotherapy and/or temozolomide. Dual staining will address cell cycle phase specificity. Repair parameters will be correlated with progression-free and overall survival.
(3) Measurement of effects of inhibitors of p38MAPkinase, mTOR or ERK signalling on MGMT gene and protein expression and temozolomide sensitivity of human glioma cells and glioma stem cells.
Scientific opportunities
Studies on glioma stem cells will verify or refute previous observations that they are radioresistant, and elucidate mechanisms underlying this phenotype. Translational studies using tumour resection specimens will interrogate in vivo effects of PARP inhibitors on DNA repair processes in GBM and normal tissues treated with radiotherapy and chemotherapy. Understanding the mechanisms underlying effects of PI3kinase/mTOR inhibition on MGMT expression will provide novel insights into interactions between oncogenic signalling pathways and protein translation, and elucidate the key factors regulating activity of this DNA repair protein.
Medical opportunities
All aspects of this proposal are aimed at improving treatment of GBM. Overcoming radioresistance of glioma stem cells is likely to be necessary to achieve tumour cure; elucidating mechanisms of sensitisation by PARP inhibitors in vivo and identifying predictive biomarkers will define their therapeutic role; and tumour-specific downregulation of MGMT may improve outcomes in GBM that are resistant to existing therapies.