| Abstract: | 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.Mutational and copy number analyses from epithelial tumors have identified several activating tyrosine kinase mutations and amplifications, such as epidermal growth factor receptor (EGFR) mutations in lung adenocarcinoma and erythroblastic leukemia viral oncogene homolog 2 (ERBB2 or HER2/neu) gene amplification in breast cancer (1). The dependence on these tyrosine kinases for tumor growth and survival has led to successful clinical treatment with tyrosine kinase inhibitors (TKIs) (2, 3). However, recent genomic analyses of prostate adenocarcinoma revealed that activating tyrosine kinase mutations or amplifications are very rare (1, 4–6).Despite the scarcity of tyrosine kinase amplifications or activating mutations in prostate cancer, tyrosine kinase expression and activity has been shown to play an important role in disease progression. For example, coexpression of wild-type SRC tyrosine kinase and androgen receptor (AR) can synergistically drive the formation of mouse prostate adenocarcinoma (7). Evaluation of nontyrosine-kinase–initiated mouse models of prostate cancer further identified activation of the nonreceptor tyrosine kinases SRC, ABL1, and Janus kinase 2 (JAK2) (8). We also observed increased tyrosine phosphorylation in nearly 50% of castration-resistant prostate cancer (CRPC) tissues examined compared with hormone-naïve prostate cancer (8). These studies suggest that comprehensive evaluation of metastatic CRPC samples for tyrosine kinase activity may lead to the identification of new drug targets.Studies in melanoma and breast cancer have revealed that despite heterogeneity in primary, localized disease, metastases seem to arise from a single precursor cell (9, 10). The multifocal nature of organ-confined prostate cancer poses a question as to the clonality of metastatic disease (11). Investigation into clonality in metastatic CRPC has found that tumors isolated from anatomically different lesions in the same patient bear similar copy number, mutational status, erythroblast transformation specific (ETS) rearrangements, and methylation patterns from multiple metastatic lesions supporting their clonal origins (6, 12–14). In addition, these studies found a remarkable amount of interpatient heterogeneity, suggesting that personalized medicine approaches may be necessary to efficiently target metastatic lesions. Previous observations of intrapatient similarity hold promise with regard to treatment strategies for metastatic CRPC patients by means of systematically attacking the cancer cell clone contributing to disease.This led us to investigate whether actionable targets such as tyrosine kinases also maintain similar activation patterns across anatomically distinct metastases from the same patient. With access to rare metastatic CRPC tissue from the University of Michigan’s Rapid Autopsy Program (15), we evaluated global tyrosine phosphorylation patterns in lethal metastatic CRPC patients. Phosphotyrosine peptide enrichment and quantitative mass spectrometry (MS) identified diverse phosphorylation events in the metastatic tissues compared with naive primary prostate tissue and prostate cancer cell line-derived xenografts. Validation of activated kinases that were identified via either MS or kinase–substrate relationships revealed intrapatient similarity and interpatient heterogeneity across a large panel of targets. Interestingly, these kinase activities are a result not of mutation (6) but rather of pathway activation within the tumors themselves. In summary, the observation that similar tyrosine kinase activities are present in most if not all anatomically disparate metastatic lesions from the same patient reveals that (i) CRPC lesions may be clonal in origin and (ii) kinase activation patterns observed in these lesions should be prioritized for further evaluation as new targeted therapeutic strategies. |
