Enzalutamide‐Resistant STEAP4+ MyoCAF Secrete Phosphatidylcholine to Foster Progression by Activating Stemness in Hormone‐Sensitive Prostate Cancer

Expanding enzalutamide (ENZ) use in hormone‐sensitive prostate cancer (HSPC) faces resistance. Single‐cell profiling identifies therapy‐resistant STEAP4+ myofibroblastic cancer‐associated fibroblast (SETAP4+ myoCAF). These myoCAFs drive resistance via TFE3‐mediated autophagy and PCYT1A‐led phosphatidylcholine overproduction, creating a phospholipid‐rich niche that activates tumor HSP90/HIF1A, promoting stemness and revealing actionable therapeutic targets for HSPC. Abstract Despite the expanding clinical application of second‐generation anti‐androgens like enzalutamide (ENZ) in hormone‐sensitive prostate cancer (HSPC), therapeutic resistance culminating in castration‐resistant prostate cancer (CRPC) persists as an unresolved clinical crisis. Through comprehensive single‐cell transcriptomic profiling of ENZ‐naïve and ENZ‐treated tumors, an expansion of ENZ‐resistant myofibroblastic cancer‐associated fibroblast (designated STEAP4+ myoCAF) is identified that correlates with adverse clinical outcomes. Strikingly, STEAP4+ myoCAF demonstrated intrinsic ENZ resistance through a mechanistically novel pathway involving transcription factor binding to IGHM enhancer 3 (TFE3)‐mediated autophagy activation. Integrated lipidomic and functional analyses revealed that TFE3 activation drives phosphatidylcholine overproduction via direct upregulation of phosphate cytidylyltransferase 1A (PCYT1A), establishing a tumor‐promoting feedforward loop. The resultant phospholipid‐rich microenvironment activates an HSP90/HIF1A signaling axis in malignant epithelial cells, fueling cancer stemness and therapeutic escape. These findings position the STEAP4+ myoCAF‐TFE3/tumor‐HIF1A axis as a master regulator of anti‐androgen resistance, offering clinically actionable targets to extend treatment efficacy in advanced prostate cancer. Expanding enzalutamide (ENZ) use in hormone-sensitive prostate cancer (HSPC) faces resistance. Single-cell profiling identifies therapy-resistant STEAP4 + myofibroblastic cancer-associated fibroblast (SETAP4 + myoCAF). These myoCAFs drive resistance via TFE3-mediated autophagy and PCYT1A-led phosphatidylcholine overproduction, creating a phospholipid-rich niche that activates tumor HSP90/HIF1A, promoting stemness and revealing actionable therapeutic targets for HSPC. Abstract Despite the expanding clinical application of second-generation anti-androgens like enzalutamide (ENZ) in hormone-sensitive prostate cancer (HSPC), therapeutic resistance culminating in castration-resistant prostate cancer (CRPC) persists as an unresolved clinical crisis. Through comprehensive single-cell transcriptomic profiling of ENZ-naïve and ENZ-treated tumors, an expansion of ENZ-resistant myofibroblastic cancer-associated fibroblast (designated STEAP4 + myoCAF) is identified that correlates with adverse clinical outcomes. Strikingly, STEAP4 + myoCAF demonstrated intrinsic ENZ resistance through a mechanistically novel pathway involving transcription factor binding to IGHM enhancer 3 (TFE3)-mediated autophagy activation. Integrated lipidomic and functional analyses revealed that TFE3 activation drives phosphatidylcholine overproduction via direct upregulation of phosphate cytidylyltransferase 1A (PCYT1A), establishing a tumor-promoting feedforward loop. The resultant phospholipid-rich microenvironment activates an HSP90/HIF1A signaling axis in malignant epithelial cells, fueling cancer stemness and therapeutic escape. These findings position the STEAP4 + myoCAF-TFE3/tumor-HIF1A axis as a master regulator of anti-androgen resistance, offering clinically actionable targets to extend treatment efficacy in advanced prostate cancer. Advanced Science, Volume 12, Issue 44, November 27, 2025.