Presentation Title

Addressing the DNA Damage Response to Irradiation and PARP1/PARG Inhibitors in Glioblastoma

Project Collaborators

Delphine Quénet (Principal Investigator), Trevor Wolf (Graduate Student)

Abstract

Grade IV glioblastoma (GBM) is one of the deadliest and fastest-progressing cancers.The mutation of the tumor suppressor PTEN is frequent (~30%) and associated with a bad prognosis. Current standard cares for GBM include tumor resection followed by chemo- and radiation therapies. The most common chemotherapeutic treatment is temozolomide (TMZ). However, GBM recurrence is almost inevitable due to TMZ resistance, highlighting the need for other therapeutic options.

Two promising drugs are Poly(ADP-ribose) Polymerase-1 (PARP1) and Poly(ADP-ribose) Glycohydrase (PARG) inhibitors. While inhibitors targeting PARP1 have been developed for decades, providing a good understand of PARP1-dependent mechanisms, potent PARG inhibitors have only recently been made available. PARP1 and PARG proteins are important for the maintenance of genome integrity. One of their main functions is in DNA repair, where they regulate chromatin structure, detect damages and recruit repair machinery. Both proteins modulate the post-translational modification called PARylation. While PARP1 catalyzes the synthesis of and covalently binds the polymer of ADP-ribose to target molecules, PARG removes those ADP-riboses when the signaling associated with PAR needs to be shut off.

To better comprehend the role of PARP1 and PARG in GBM, we are comparing the outcomes of these inhibitors. Using established GBM cell lines, we are comparing the level of DNA damage (gH2AX foci) by immunofluorescence after irradiation, and irradiation coupled with PARP and PARG inhibitors. We are also measuring genomic instability by counting micro-nuclei which result from chromosome missegregation. Finally, we are performing Annexin V/Propidium Iodide assay to determine whether these treatments induce cell death.

Preliminary data on irradiated GBM cells indicates that the number of γH2AX foci is correlated with irradiation dose (from 0Gy to 8Gy). However, the average number of micronuclei peaks around 2-4 Gy. Interestingly, irradiation activates cell death in the LN-229 GBM cell line which express a wildtype PTEN protein, but not in the U-87MG GBM cell line (PTEN-mutant). These data suggest that sensitivity to irradiation depend to PTEN status. Further study will provide insight on how PTEN status contribute to the response to irradiation in combination with PARP or PARG inhibitors.

Primary Faculty Mentor Name

Delphine Quénet

Status

Undergraduate

Student College

College of Arts and Sciences

Program/Major

Neuroscience

Primary Research Category

Biological Sciences

Second College (optional)

Honors College

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Addressing the DNA Damage Response to Irradiation and PARP1/PARG Inhibitors in Glioblastoma

Grade IV glioblastoma (GBM) is one of the deadliest and fastest-progressing cancers.The mutation of the tumor suppressor PTEN is frequent (~30%) and associated with a bad prognosis. Current standard cares for GBM include tumor resection followed by chemo- and radiation therapies. The most common chemotherapeutic treatment is temozolomide (TMZ). However, GBM recurrence is almost inevitable due to TMZ resistance, highlighting the need for other therapeutic options.

Two promising drugs are Poly(ADP-ribose) Polymerase-1 (PARP1) and Poly(ADP-ribose) Glycohydrase (PARG) inhibitors. While inhibitors targeting PARP1 have been developed for decades, providing a good understand of PARP1-dependent mechanisms, potent PARG inhibitors have only recently been made available. PARP1 and PARG proteins are important for the maintenance of genome integrity. One of their main functions is in DNA repair, where they regulate chromatin structure, detect damages and recruit repair machinery. Both proteins modulate the post-translational modification called PARylation. While PARP1 catalyzes the synthesis of and covalently binds the polymer of ADP-ribose to target molecules, PARG removes those ADP-riboses when the signaling associated with PAR needs to be shut off.

To better comprehend the role of PARP1 and PARG in GBM, we are comparing the outcomes of these inhibitors. Using established GBM cell lines, we are comparing the level of DNA damage (gH2AX foci) by immunofluorescence after irradiation, and irradiation coupled with PARP and PARG inhibitors. We are also measuring genomic instability by counting micro-nuclei which result from chromosome missegregation. Finally, we are performing Annexin V/Propidium Iodide assay to determine whether these treatments induce cell death.

Preliminary data on irradiated GBM cells indicates that the number of γH2AX foci is correlated with irradiation dose (from 0Gy to 8Gy). However, the average number of micronuclei peaks around 2-4 Gy. Interestingly, irradiation activates cell death in the LN-229 GBM cell line which express a wildtype PTEN protein, but not in the U-87MG GBM cell line (PTEN-mutant). These data suggest that sensitivity to irradiation depend to PTEN status. Further study will provide insight on how PTEN status contribute to the response to irradiation in combination with PARP or PARG inhibitors.