Background: Radium-223 is a targeted alpha therapy that improves survival in men with mCRPC. The biologic basis for radium-223 efficacy is not completely understood. We hypothesized that PC osteomimicry, a form of epithelial plasticity leading to an osteoblastic phenotype, may contribute to the intralesional deposition of radium-223 and subsequent irradiation of the tumor microenvironment. Methods: We conducted a pharmacodynamic study of radium-223 in men with bone predominant mCRPC to investigate genomic and phenotypic alterations in circulating tumor cells (CTCs), ctDNA, and metastases. Prior to radium and 3 and 6 months after radium, liquid and metastatic biopsies were collected, including CTCs for phenotypic characterization and CTC/ctDNA genomic analysis. The primary objective was to describe the prevalence of CTC bone alkaline phosphatase (BAP) over time. We measured radium-223 decay products in tumor and surrounding normal bone during treatment. Results: We enrolled 20 men with heavily pre-treated symptomatic bone predominant mCRPC and treated with radium-223 over a median of 6 doses. 55% had elevated serum BAP. PFS was 5.5 mo; OS was 13.3 mo; 10% had unfavorable (≥5) to favorable ( < 5) Cellsearch CTC conversion; 5% had ≥30% PSA decline. We found evidence of persistent CTC BAP expression in both EpCAM+ and EpCAM- CTCs in the majority of men over time during radium treatment. We identified genomic gain of key osteomimicry regions in CTC DNA, including loci for BAP, osteopontin, and OB-cadherin. Radium-223 uptake was observed in tumor to a greater degree than surrounding normal bone. CTC DNA and matched ctDNA and CTC cultures suggested persistence of CTCs with aggressive genomic alterations such as AR, FOXA1, and MYC gain and PTEN and RB1 loss. We established multiple CTC cultures and one CTC PDX; cells exhibited evidence of epithelial plasticity with BAP expression. Conclusions: Osteomimicry may contribute to the uptake of Radium-223 within bone metastases and may thereby enhance the therapeutic benefit of radium-223. We found genomic and phenotypic evidence of osteomimicry in CTCs and CTC cultures from men with mCRPC. Clinical trial information: NCT02204943