Targeting Glutamate to Treat Brain Cancer – Repurposing Clinically Approved Glutamate Antagonists to Treat Glioblastoma

Glioblastoma represents the most common primary malignant brain tumor in adults and is characterized by an aggressive clinical course, with a median overall survival of approximately twelve months. The current standard-of-care treatment consists of maximal safe surgical resection followed by chemoradiotherapy with temozolomide and temozolomide alone. Despite these interventions, most patients experience tumor recurrence within one year after diagnosis, and therapeutic options for recurrent disease remain extremely limited. Consequently, the overall prognosis for individuals diagnosed with glioblastoma is unfavorable. Glioblastoma is thought to owe part of its devastating clinical course to its ability to intricately interact with the neural environment, mimicking neuronal processes and integrating into brain circuits to drive its own growth and invasion. Glioma cells form functional synapses with neurons, establish interconnected tumor microtube networks, and even generate intrinsic rhythmic electrical activity that promotes malignant features. These interactions are further enhanced by paracrine factors such as NLGN-3, IGF1, BDNF, and TSP-1, which facilitate synaptogenesis and tumor progression. At the center of these mechanisms lies glutamate, the brain’s primary excitatory neurotransmitter, which glioblastoma cells secrete into the tumor microenvironment to induce neuronal hyperexcitability, excitotoxic neuronal death, and calcium-dependent signaling that fuels proliferation and invasion. The pleiotropic role of glutamate highlights this metabolite as a central factor driving glioblastoma’s aggressiveness and poor patient outcomes and has been the main rationale for this thesis and the ongoing phase Ib/II randomized clinical trial GLUGLIO (NCT05664464), which explores the efficacy of combined memantine, sulfasalazine, and gabapentin in conjunction with standard chemoradiotherapy in patients with newly diagnosed glioblastoma. In this project, we used an RCAS/tv-a glioblastoma mouse model, which recapitulates key glioblastoma oncogenic drivers and relevant drug targets, to evaluate the therapeutic potential of the triple combination of anti-glutamatergic drugs. Our results demonstrated that daily treatment with this triple combination prolonged median survival of tumor-bearing mice. This survival benefit was not observed with single or double drug combinations, nor with the standard antiepileptic drug levetiracetam, suggesting a synergistic effect of this drug combination. Importantly, we found that none of the drugs, individually or in combination, affected tumor cell viability under acute or chronic exposure conditions in vitro. Electrophysiological analyses further revealed that the drug combination modulates neuronal network activity, shifting it towards a less synchronized and less hyperexcitable state. Crucially, this treatment reversed glioblastoma-induced alterations observed in neuron–tumor co-cultures. In relation to glutamate, we further confirmed that sulfasalazine is capable of modulating both intra- and extracellular glutamate concentrations in vitro. In contrast, memantine and gabapentin did not regulate glutamate concentrations. Consistent with these findings, in vivo analyses revealed that treatment with the triple drug combination did not result in measurable alterations in glutamate concentrations, suggesting that the observed therapeutic benefits are unlikely to be mediated through direct modulation of overall glutamate levels. Collectively, these results suggest that the triple drug combination represents a promising therapeutic strategy for glioblastoma, with demonstrated an albeit modest survival benefit in mice. Furthermore, data imply that the mechanism of action extends beyond glutamate metabolism regulation and may involve the regulation of neuronal input to the tumor and its microenvironment. $\textit{Full-text embargoed until: 2026-05-12}$