In prostate cancer, multiple metastases from the same patient share similar copy number, mutational status, erythroblast transformation specific (ETS) rearrangements, and methylation patterns supporting their clonal origins. Whether actionable targets such as tyrosine kinases are also similarly expressed and activated in anatomically distinct metastatic lesions of the same patient is not known. We evaluated active kinases using phosphotyrosine peptide enrichment and quantitative mass spectrometry to identify druggable targets in metastatic castration-resistant prostate cancer obtained at rapid autopsy. We identified distinct phosphopeptide patterns in metastatic tissues compared with treatment-naive primary prostate tissue and prostate cancer cell line-derived xenografts. Evaluation of metastatic castration-resistant prostate cancer samples for tyrosine phosphorylation and upstream kinase targets revealed SRC, epidermal growth factor receptor (EGFR), rearranged during transfection (RET), anaplastic lymphoma kinase (ALK), and MAPK1/3 and other activities while exhibiting intrapatient similarity and interpatient heterogeneity. Phosphoproteomic analyses and identification of kinase activation states in metastatic castration-resistant prostate cancer patients have allowed for the prioritization of kinases for further clinical evaluation.