Cell-Autonomous EPHB2 signaling sustains neuroendocrine prostate cancer

Neuroendocrine prostate cancer (NEPC) is a lethal subtype of prostate cancer that commonly develops from prostatic adenocarcinoma after long-term androgen deprivation therapy (ADT). Its incidence has risen markedly over the past decade with the widespread use of potent AR pathway inhibitors (ARPIs), yet no effective, approved therapies exist. Most studies focus on terminal NEPC, while the early drivers and transitional cellular states remain poorly defined. To address this gap, we used the LTL331/LTL331R patient-derived xenograft (PDX) model, the only available PDX system that faithfully recapitulate ADT-induced adenocarcinoma-to-NEPC transdifferentiation. Through longitudinal single cell transcriptomic sequencing across the entire process of adenocarcinoma-to-NEPC transdifferentiation of human prostate cancer, we identified a distinct intermediate, transitional cell state in the lineage shift. Within this state, EPHB2 emerged as a potential lineage-determining receptor activated at the onset of NE transdifferentiation. Mechanistically, we found that endothelial cells (ECs) in the tumor microenvironment (TME) initiate NE transdifferentiation by activating an EFNB2-EFNA5-EPHB2 axis in cancer cells. EC-derived EFNB2 upregulates EFNA5, which subsequently activates EPHB2 in a cell-autonomous manner. This signaling pair suppresses AR signaling and luminal lineage programs while promoting NE lineage initiation. In terminal NEPC, sustained EFNA5-driven EPHB2 activation maintains NE identity and enhances aggressiveness through a self-reinforcing loop. Functionally, EPHB2 inhibitor significantly reduced NEPC cell proliferation and NE marker expression. Importantly, combining an EPHB2 inhibitor with an EZH2 inhibitor partially reversed the NE phenotype, restored AR signaling, and resensitized NEPC cells to ARPIs.In conclusion, EPHB2 functions as a dual-phase regulator in NEPC, initially activated by TME-derived ephrin ligands to trigger NE transdifferentiation and later sustained through cell-autonomous signaling to maintain terminal NEPC. These findings position EPHB2 as a promising therapeutic target in intercepting NEPC development and progression.