# Dynamic Reprogramming of Immune-Related Signaling During Progression to Enzalutamide Resistance in Prostate Cancer

## TL;DR

This study shows that early enzalutamide treatment boosts immune signaling in prostate cancer, but resistance later suppresses it, suggesting a window for combining therapies to prevent treatment failure.

## Contribution

The study identifies a transient early window of immune responsiveness during enzalutamide treatment that could be therapeutically exploited to prevent resistance.

## Key findings

- Early enzalutamide treatment enhances IFNγ and IL6 signaling in prostate cancer cells.
- Enzalutamide-resistant cells show suppressed immune signaling and increased lineage plasticity.
- Early-stage cells are highly sensitive to STAT1 inhibition by fludarabine, but this sensitivity is lost in resistant cells.

## Abstract

Enzalutamide treatment can drive lineage plasticity that gives rise to treatment-induced neuroendocrine prostate cancer (t-NEPC), a lethal subtype of prostate cancer. Despite its clinical importance, the immune-related signaling dynamics underlying this enzalutamide-driven plasticity remain poorly understood. This study uncovers a temporal window during early enzalutamide therapy where inflammatory signaling pathways are transiently activated and therapeutically targetable, offering a strategy to delay or prevent neural lineage plasticity transformation and treatment resistance.

Background: Treatment with androgen receptor (AR) signaling inhibitors, such as enzalutamide, can induce neural lineage plasticity in prostate cancer, potentially progressing to t-NEPC. However, the molecular mechanisms underlying this enzalutamide-driven plasticity, particularly the contribution of immune signaling pathways, remain poorly understood. Methods: We analyzed transcriptomic profiles of patient samples and prostate cancer cell lines to investigate changes in immune signaling pathways. Interferon gamma (IFNγ), interferon alpha (IFNα), and interleukin 6 (IL6)-Janus kinase (JAK)-signal transducer and activator of transcription 3 (STAT3) signaling were assessed in enzalutamide-sensitive and -resistant prostate cancer cells. Functional assays were conducted to examine cell responsiveness to cytokine stimulation and susceptibility to STAT1 inhibition using fludarabine. Results: Immune-related pathways, including IFNγ, IFNα, IL6-JAK-STAT3, and inflammatory responses, were significantly suppressed in NEPC patient samples compared to those with castration-resistant prostate cancer (CRPC). Enzalutamide-resistant and NEPC cells exhibited markedly impaired IFNγ and IL6 signaling. In contrast, early-stage enzalutamide treatment paradoxically enhanced IFNγ and IL6 responsiveness. Transcriptomic profiling revealed coordinated upregulation of E2F target genes and activation of IFNα/IFNγ and JAK/STAT signaling pathways during early treatment. Importantly, these early-stage cells remained highly sensitive to IFNγ and IL6 stimulation and showed increased susceptibility to STAT1 inhibition by fludarabine, a sensitivity that was lost in resistant cells. Conclusions: Early enzalutamide treatment enhances immune responsiveness, while the development of resistance is associated with suppressed immune signaling and increased lineage plasticity. These results suggest a therapeutic window where combining enzalutamide with STAT inhibitors may delay or prevent lineage plasticity and resistance.