Despite advances in the development of highly effective androgen receptor (AR)-directed therapies for the treatment of men with advanced prostate cancer, acquired resistance to such therapies frequently ensues. A significant subset of patients with resistant disease develop AR-negative, androgen-indifferent tumors that lose their luminal identity and display neuroendocrine features of (neuroendocrine (NE) prostate cancer (NEPC)). The cellular heterogeneity and the molecular evolution during the progression from AR-positive, adenocarcinoma to AR-negative NEPC has yet to be characterized. Moreover, understanding the role of NE signaling in carcinogenesis and cell reprogramming during this progression following anti-androgen therapy remains an unmet need. Utilizing a new genetically engineered mouse model, we have characterized the synergy between Rb1 loss and MYCN (encodes N-Myc) overexpression which results in the formation of AR-negative, poorly differentiated tumors with high metastatic potential. Preliminary single-cell-based approaches revealed striking observations including subsets of NEPC tumor cells.  We have also designed a new mouse model allowing for controlled prostate-specific ablation of NE cells.  However, more work is needed to characterize these models and to understand the heterogeneity of temporal changes to the transcriptome and chromatin accessibility during the emergence of distinct cell populations that give rise to NEPC.

Aim 1. Uncover the temporal progression to AR-indifferent prostate cancer at the single cell level after AR targeted therapies. We will explore the cell hierarchy of the proximal and distal prostate during development of NEPC driven by MYCN-induced/Pten/Rb1 co-loss loss using a new genetically engineered mouse model (GEM) in the Rickman lab (Fig. 1A) or another GEM with Trp53/Rb1 co-loss (Nikitin). This will involve longitudinal single-cell analyses (RNAseq and ATACseq, Rickman and Nikitin) and detailed immunofluorescent assessments (Nikitin) using lineage markers to track primary tumor and metastatic cell evolution. This analysis will reveal biomarkers and potential targets that will be validated using a large clinical cohort of locally advanced, CRPC and NEPC samples (Rickman).

Aim 2. Characterize the role of NE signaling in prostatic regeneration and carcinogenesis. This will include NE cell ablation in models based on MYCN-induced/Pten/Rb1 co-loss (Rickman) and by Trp53/Rb1 co-loss (Nikitin), and longitudinal evaluation of changes in NE networks associated with regeneration and malignant transformation. These studies will facilitate identification of carcinogenesis stages and/or treatment regiments critical for overt lineage plasticity and appearance of NE phenotype.