Identifying novel therapeutic targets for seizures and brain cancers

Abstract

Epilepsy describes a range of conditions characterised by unprovoked recurring seizures that are due to multiple factors including the excitatory neurotransmitter glutamate. A subgroup of patients with epilepsy may not respond to systemic treatment and therefore would become candidates for surgical therapy. A less invasive approach is vagal nerve stimulation, which I demonstrated is safe and effective in 100 patients, in the first UK based large case series.

Another group of patients that suffer from intractable epilepsy are those with tumour associated seizures (TAS). In particular, patients diagnosed with glioblastoma multiforme can initially present with unexplained seizures or develop seizures later in the course of the disease. TAS is often pharmaco-resistant and treatment options are limited. Animal models of TAS demonstrated that the systemic Xc- cysteine-glutamate exchange transporter (XCT) antagonist sulfasalazine reduced the frequency of seizures. In human studies of TAS, increased levels of glutamate, reduced excitatory amino acid transporter 2 (EAAT2) expression and reduced XCT were associated with greater frequency of TAS.

We found that EAAT2 single nucleotide mutations correlated with an increased susceptibility to cerebral palsy in pre-term infants, implicating glutamate excess as the underlying cause. Together with evidence of downregulation of EAAT at the peri-tumoural region in TAS, it became evident that glutamate mediated excitotoxicity caused by dysfunctional EAAT2 may be a therapeutic target in such diseases. Therefore, I investigated strategies to modulate the expression of EAAT2, identifying peroxisome proliferator gamma (PPARγ) agonists as a promising candidate.

I utilised Immortalised glioblastoma cell lines to investigate the role of the PPARγ agonist pioglitazone on glutamate transport in vitro. Pioglitazone was chosen for its established safety profile in type 2 diabetes and widely reported anti-neoplastic effects. I demonstrated that pioglitazone increases EAAT2 expression and reduces extracellular concentrations of glutamate in doses that do not affect cell viability. At the time of publication, this work identified a second drug candidate for the treatment of TAS in addition to sulfasalazine, which proved to be potentially unsafe in a clinical trial.

Glioblastoma is associated with a very poor prognosis due to its invasiveness and presence of cancer stem cells. We showed that the use of multimodal MRI can help delineate the invasive margin of glioblastoma but further therapeutic strategies would be needed. As such, my focus shifted to cancer cell migration since glioblastoma is known to be a highly infiltrative and diffuse disease, where glutamate mediate excitotoxicity was thought to facilitate this process. The concept of cancer stem cells offered a new explanation for treatment resistance, where this subpopulation of cells was shown to be resistant to conventional treatments. Further, I demonstrated that neural stem cells are directed by electric fields (EFs). As such, I investigated the role of EFs on both differentiated and cancer stem cells, finding that each cell type had opposing responses to EFs. I subsequently investigated the role of pioglitazone, finding that directed cell migration was inhibited in both cells types, indicating that this drug class may have a further role in preventing glioblastoma recurrence.