Metastatic prostate cancer remains incurable and inevitably progresses to lethal disease despite expanding treatment options. Standard of care includes therapies targeting the androgen receptor (AR) pathway, a key regulator of prostate lineage and survival. Despite a decade of approvals of more potent AR Signaling Inhibitors (ARSI), nearly all patients with metastatic prostate cancer develop recurrence due to the emergence of castration-resistant prostate cancer (mCRPC). Even with recent advances, the mechanisms driving metastatic progression and the diverse molecular underpinnings of ARSI resistance remain to be fully elucidated, thereby complicating the development of therapies to overcome resistance. Within the tumor microenvironment (TME), cancer associated fibroblasts (CAF) are phenotypically and functionally heterogenous and have varying roles in tumor progression and therapeutic resistance, however, many of these mechanisms remain elusive. We and others have revealed that Asporin (ASPN)+ CAF are associated with worse oncological outcomes including metastasis, and we have further demonstrated that they are enriched in therapy resistant metastases. As a secreted protein, ASPN has been shown to communicate throughout the TME to orchestrate cancer cell migration and metastatic development. Our new data indicate that ASPN also protects prostate cancer cells from enzalutamide, a second-generation ARSI. Preliminary data support that ASPN in the TME regulates metastasis and therapy resistance through the activation of HER2/3, a targetable pathway with reported involvement in migration, metastasis, and ARSI resistance. Consistent with this, preliminary analyses show that ASPN+ CAF co-localize with HER2 and HER3 expression in mCRPC. Preliminary mechanistic data indicate that ASPN binds to HER3 and induces the activation of HER2/3 and downstream pathways including MAPK, PI3K, and calcium signaling. Our preliminary findings highlight the importance of ASPN-HER2/3 mediated release of intracellular calcium stores in both HER2/3 signal amplification and calcium microdomain oscillations which are critical for migration. Collectively, our data strongly support our central hypothesis that ASPN+ CAF potentiate prostate cancer migration, metastasis, and ARSI resistance by activating HER2/3 signaling in adjacent prostate cancer cells. Our goals are to 1) Define the molecular mechanisms of ASPN-induced HER2/3 activation, 2) Determine mechanisms of ASPN-HER2/3 regulated calcium oscillations in cellular migration, and 3) Elucidate the functional role of ASPN-HER2/3 in prostate cancer metastasis and therapy resistance. We expect that the outcomes from the proposed studies will provide 1) critical information on a novel mechanism of HER2/3 activation by ASPN in the TME and 2) opportunities for repurposing HER2/3 directed therapies for innovative therapeutic approaches in prostate cancer. By using cutting edge technologies, we will expand current paradigms pertainin...