Abstract LB020: Role of acetylcholine in glioblastoma pathophysiology: Emerging insights

Abstract BACKGROUND: Glioblastoma (GBM) is the most common primary brain tumor in adults and associated with abysmal patient outcomes. While it is well-known that glutamatergic neurons can form functional synapses with GBM cells to enhance tumor growth, less is known about the effects of the activity of other neuronal types on GBM biology. Here, we examined the effect of acetylcholine on the growth of GBM cells belonging to different molecular subtypes. We also explored ways to modulate acetylcholine produced by cholinergic neurons in mouse basal forebrain to further study the effect on the pathophysiology of GBM. METHODS: Patient-derived GBM cell lines were exposed to varying concentrations of neurotransmitters (acetylcholine & glutamate) and assessed for cell viability using CCK-8 and EdU assays. Atropine, an antagonist of muscarinic receptors, was used to analyze the effect of cholinergic receptor inhibition on acetylcholine-mediated GBM cell growth in vitro and on the survival of mice bearing human GBM xenografts in vivo. Cholinergic neurons in the mouse basal forebrain were targeted for modulation of acetylcholine production by means of chemogenetics, as well as an attention-related behavior paradigm such as a novel object recognition test (NORT). We also examined possible dysregulation of cholinergic signaling in a preneoplastic glioma mouse model using RNA-seq analysis. RESULTS: RNA sequencing analyses revealed altered expression of cholinergic receptors in preneoplastic glioma mouse models, suggesting a potential role of cholinergic signaling in glioma formation. GBM cell growth was increased upon incubation with acetylcholine. Consistently, pharmacologic inhibition of muscarinic receptors prolonged the survival of mice bearing human GBM xenografts, suggesting a potential role of cholinergic signaling in glioma growth. Last, we tested the possibility of targeting cholinergic neurons in mouse basal forebrain using AAV-mediated chemogenetics and attention-associated behavior tasks. CONCLUSIONS: Our findings suggest that GBM cells, irrespective of molecular subtype, may leverage cholinergic signaling mechanisms to support their pathophysiology. Future studies will investigate the role of cholinergic neuron activity in GBM initiation and growth. Citation Format: Khushboo Irshad, Aishani S. Gargapati, Sanjay Singh, Khalil Ali Ahmad Kasm, Kechen Ban, Benjamin E. Aghoghovwia, Rajasekaran Mahalingam, Momo K. Harris, Sama Alnassiry, Renae Bertrand, Alexia T. Mendoza, Valeria S. Vega Sánchez, Nathan Bui, Truc Kuo, Frederick Lang, Yuan Pan. Role of acetylcholine in glioblastoma pathophysiology: Emerging insights [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr LB020.