Abstract Neurons form functional synapses with glioblastoma and metastatic brain cancer cells. While pharmacological disruption of these connections can reduce tumor growth in preclinical models, we lack methods to identify which specific neuronal and tumor cell types form these pathological connections. This knowledge gap limits our ability to develop targeted therapies that selectively disrupt tumor-supporting neural circuits while preserving normal brain function. We developed Interactome-seq, a technology that combines engineered synaptic proteins with RNA barcoding to map cell-cell connections at single-cell resolution. Our constructs use split-fluorescent proteins that reconstitute only at cell-cell contact sites, enabling isolation of intact synapses for molecular analysis. Unique RNA barcodes identify the specific cell types forming each connection. We validated this system in cell culture, demonstrating selective labeling of cell-cell interfaces and efficient RNA barcode capture. We are now deploying Interactome-seq in mouse glioblastoma models to create comprehensive maps of neuron-tumor connectivity. This technology will reveal which neuronal subtypes preferentially synapse with tumor cells and identify cancer cell subpopulations with enhanced capacity for neural integration. These insights may guide development of precision therapies targeting specific neuron-tumor interactions while minimizing effects on healthy neural circuits. Beyond brain tumors, Interactome-seq could be adopted to map any cell-cell interaction in complex tissues, including immune-tumor contacts, neurodevelopmental connections, and disease-associated cellular networks, opening new avenues for understanding and treating diverse disease conditions. Citation Format: Boxuan Zhao. Interactome-seq: A novel technology for mapping neuron-cancer synapses at single-cell resolution abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Brain Cancer; 2026 Mar 23-25; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86 (6Suppl): Abstract nr PR004.
Boxuan Zhao (Mon,) studied this question.