Abstract A039: Arginyltransferase Ate1 reduction drives prostate cancer metastasis

Abstract

Abstract This study demonstrates that diminished Arginyltransferase (Ate1) expression in prostate cancer is sufficient to drive progression and distal metastasis, as shown via in vitro and in vivo models, and in correlation analysis with human patient samples. While prostate cancer affects a large population of men, it is usually a dormant disease and is only lethal in the small subpopulation in which distal metastasis occurs. Unfortunately, distal metastasis is currently difficult to predict and treat. Understanding what provokes metastasis in prostate cancer is therefore clinically important. Ate1 is the enzyme solely responsible for mediating post-translational protein arginylation in most eukaryotic cells, including mammalian cells. Previous studies show that a complete loss of Ate1 in yeast and mammalian cells increases resistance to stress-induced cell death and promotes genomic instability. Ate1 loss further promotes cancer-relevant phenotypes in mouse fibroblasts, such as persisting cellular replication despite stress or cell-cell contacts. Furthermore, a lower level of Ate1 appears to correlate with shorter survival in patients with several types of cancer, including prostate cancer. However, the role of Ate1 in progression and metastasis of prostate cancer (or any cancer) remained unknown. Here we demonstrate in prostate-relevant models that a reduction of Ate1 drives prostate cancer progression and metastasis. We first examined Ate1’s impact on stress-induced cell death on prostate cancer cells in vitro in models relevant to the tumor microenvironment and cancer treatments. PC-3 and LNCaP prostate cancer cells with shRNA targeting Ate1 (or nontargeting control) were stressed with cellular oxidant hydrogen peroxide, gamma radiation, and apoptotic inducer staurosporine. Consistent with our hypothesis, the knockdown of Ate1 resulted in less cell death in these cells. Interestingly, while a reduction of Ate1 in these cell lines does not affect proliferation, it does cause an increase in anchorage-independent growth in LNCaP cells via the soft agar assay, and an increase in PC-3 cell invasion through Matrigel. Additional analysis of prostate cancer cell lines PC-3, LNCaP, metastatic PC-3-derivative PC3-ML, metastatic LNCaP-derivative C4-2B, and metastatic line DU145 showed a distinct and significant relationship between increased malignancy and decreased Ate1 expression. Next, to study the effect of Ate1 on metastasis in vivo, PC-3 cells expressing luciferase and with shRNA targeting Ate1 (or control) were injected into prostates of immune-compromised (nu/nu) mouse (orthotopic xenografts). PC3-ML, a derivative of PC-3 known to be metastasis-competent, was used as a positive metastatic control. We used luciferin luminescence to measure tumor growth and the presence of local invasions to adjacent tissues such as seminal vesicles and testicles, as well as metastasis to distal sites including lymph nodes, spleen, and lung. Sixty days after tumor inoculation, all three cell types consistently formed similar primary tumors but varied greatly in their metastatic behaviors. The PC-3 cells with nontargeting control shRNA largely failed to metastasize to distal sites and only occasionally exhibited local invasion, as expected from previous reports. However, PC-3 cells with Ate1 downregulated by shRNA consistently formed distal metastases in lungs, spleen, and lymph nodes with similar frequency to the positive control, PC3-ML. As further support for Ate1’s role in prostate cancer progression, results from data mining and immune-histologic examination of human tissue samples revealed a consistent downregulation of Ate1 protein and mRNA during prostate cancer progression and metastasis, offering promising value of predicting metastatic events before they occur. In conclusion, Ate1 reduction may be a potent indicator of prostate cancer progression, and provide insight into the molecular mechanism of prostate cancer metastasis. Citation Format: Michael D. Birnbaum, Ning Zhou, Kerry L. Burnstein, Fangliang Zhang. Arginyltransferase Ate1 reduction drives prostate cancer metastasis [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A039.