ABSTRACT The role of tumor-macrophage hybrid cells in prostate cancer metastasis AR signaling inhibitors (ARSIs) are commonly used to treat recurrent prostate cancer whose growth depends on AR activities. However, ARSIs can result in selective pressure, transitioning AR-dependent tumors to AR- independent tumors. Extensive studies have revealed signal-switching mechanisms by which ARSIs exert the phenotypic transition leading to metastatic castration-resistant prostate cancer (mCRPC). Apart from these signaling circumventions, here we propose that tumor-macrophage hybrid (TMH) cells are a novel cellular contributor to mCRPC. Our preliminary data suggest that TMH cells adopt macrophages' characteristics to evade immune surveillance and withstand hemodynamic stress in the vasculature for distant colonization. In addition, TMH cells intrinsically express very low AR/AR-targets, resembling an adenocarcinoma-immune subtype of mCRPC that resists ARSIs. Three specific aims will use systems-level approaches to mechanistically define the role of TMH cells in mCRPC. In Aim 1, we will demonstrate that TMH cells leverage abundant c-Myc from parental cancer cells to upregulate M1/M2 polarization loci derived from parental macrophages. This unique transcription machinery, not observed in either parental cells, is being exploited to confound the host immune system, thereby enhancing the immunoevasive features of TMH cells. In this regard, c-Myc acts as an immunoevasive driver, not primarily as an oncogenic driver per se, in TMH cells. In Aim 2, we will show that as a result of cellular fusions, TMH cells undergo transcriptomic rewiring of the AR cistrome, leading to treatment resistance of ARSIs. Consequently, these cells overtake AR-dependent cancer cell populations for malignant outgrowth. In Aim 3, we will evaluate whether integrin-mediated mechanotransduction in TMH cells strengthens their resistance to fluid shear stress in blood circulation. The signaling cascade enhances a mesenchymal feature of TMH cells, endowing them with higher viability and motility than circulating tumor cells (CTCs). Therefore, our proposed study highlights an overlooked cell-intrinsic factor contributing to mCRPC. Furthermore, enumeration and molecular profiling of TMH cells represent a new type of liquid biopsy for prostate cancer patients.

PROJECT NARRATIVE The U01 study will mechanistically characterize tumor-macrophage hybrid cells as a new and unexpected player in metastatic castration-resistant prostate cancer (mCRPC). Molecular profiling of these malignant cells may reveal their Achilles' heels, presenting opportunities to develop new agents for treating this mCRPC subtype.