Tetzlaff, S. K.; Reyhan, E.; Bengtson, C. P.; Schroers, J.; Wagner, J.; Schubert, M. C.; Layer, N.; Puschhof, M. C.; Faymonville, A. J.; Drewa, N.; Pramatarov, R. L.; Wissmann, N.; Alhalabi, O.; Heuer, A.; Sivapalan, N.; Campos, J.; Boztepe, B.; Scheck, J. G.; Villa, G.; Schröter, M.; Sahm, F.; Forsberg-Nilsson, K.; Breckwoldt, M. O.; Acuna, C.; Suchorska, B.; Heiland, H. D.; Saez-Rodriguez, J.; Venkataramani, V.

Glioblastomas are heterogeneous brain tumors, notorious for their invasive behavior and resistance to therapy. Neuron-to-glioma synapses have been identified to promote glioblastoma invasion and proliferation. However, a comprehensive characterization of tumor-connected neurons has been hampered by a lack of technologies. Here, we adapted retrograde tracing with a modified rabies virus system to characterize and manipulate connected neuron-tumor networks. Glioblastoma rapidly integrated into neural circuits across the brain engaging in widespread functional communication, with acetylcholinergic and glutamatergic neurons driving glioblastoma progression. We uncovered patient-specific and tumor cell state-dependent differences in synaptogenic gene expression driving neuron-tumor connectivity and subsequent invasivity. Importantly, radiotherapy enhanced neuron-tumor connectivity by increased neuronal activity. In turn, simultaneous inhibition of AMPA receptors and radiotherapy showed increased therapeutic effects, indicative of a role for neuron-to-glioma synapses in contributing to therapeutic resistance. Lastly, rabies-mediated genetic ablation of tumor-connected neurons halted glioblastoma progression, offering a novel viral strategy to target glioblastoma. Together, this study provides a comprehensive framework for basic research and clinical translation of synaptic neuron-cancer interactions to target glioblastoma.