br The result that statin induced
The result that statin-induced cell death can be rescued by exogenous mevalonate suggests that, in part, statins act by directly inhibiting
HMGCR in tumor cells. Achieving the same effect clinically would require sufficient statin drug accumulation in the peripheral circulation and prostate tissue. For our studies, we evaluated fluvastatin, a lipo-philic statin that can achieve high maximum serum concentrations , which we hypothesized could reach the prostate. To the best of our knowledge, we report here for the first time that fluvastatin can be measured in the mouse prostate (Figure 2); however, assuming that these concentrations are similar to those achievable in the human prostate, fluvastatin monotherapy is unlikely to be effective at killing PCa Cell Counting Kit-8 (CCK-8) in every patient. Two potential strategies to increase the anti-cancer effects of statins in the prostate are: i) dose-escalation and ii) statin-drug combination therapy. In our xenograft studies, fluvastatin was administered at 2e3 the typical cholesterol-lowering dose. Evidence from a number of phase I clinical studies has demonstrated that statins are well-tolerated at these higher doses [53,54]. We, and others, have also demonstrated that the cytotoxic effects of statins are both dose- and time-dependent [19,20,49], and therefore longer treatment durations at lower doses may be equally as effective, especially for a disease with a long natural history such as PCa. Prospective dose-finding studies are necessary to evaluate how best to prescribe statins in the context of PCa. In addition to dose-escalation, statins can be combined with other agents that potentiate their anti-cancer effects. Here, we provided evidence to support that inhibiting the sterol-regulated feedback loop of the MVA pathway with dipyr-idamole can potentiate fluvastatin-induced apoptosis, both in vitro and in vivo. Furthermore, co-treatment with dipyridamole significantly re-duces the dose of fluvastatin required to induce PCa cell death.
Inhibiting the MVA pathway at the level of HMGCR with physiologically-achievable concentrations of fluvastatin is only effective at killing a minority of PCa cells. In other PCa cells, where SREBP2 is activated following fluvastatin treatment, fluvastatin-induced cell death is dampened. In these cells, inhibiting both the MVA pathway and SREBP2 is necessary to induce apoptosis, which can be achieved by combining fluvastatin with a second clinically-approved agent, dipyr-idamole. Our study provides strong pre-clinical rationale to warrant further clinical evaluation of these immediately-available and well-tolerated drugs for the treatment of PCa.
JL, PJM and LZP conceived and designed the study. JL, PJM, RY and JEV performed experiments. JL analyzed and interpreted the experi-mental data. MM, JMS and THV scored the TMAs. RY and EXC per-formed the HPLC-MS/MS analysis. YW provided the PDX model. DTSW, RJH and NEF provided clinical data and expertise. JL and LZP wrote the manuscript. All authors read and approved the manuscript. LZP su-pervised the study.
We thank all members of the Penn lab for helpful discussions. We especially thank Dr. Carolyn Goard and Melanie Peralta for their assistance with purifying and vali-dating the HMGCR antibody for IHC. We also thank Drs. Robert Bristow and Mathieu Lupien for reagents, and Dr. Miran Kenk, Wenjiang Zhang and Maria Monroy for their technical support. This work was supported by funding from the Canada Research Chairs program (L.Z. Penn), Canadian Institutes of Health Research (FRN: 142263; L.Z. Penn), CIHR Doctoral Research Awards (J. Longo and R. Yu) and Ontario Student Opportunity Trust Fund (J.E. van Leeuwen). This work was also supported by the
128 MOLECULAR METABOLISM 25 (2019) 119e1302019 University Health Network. Published by Elsevier GmbH. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Princess Margaret Cancer Foundation Hold’em for Life Prostate Cancer Research Fund.
APPENDIX A. SUPPLEMENTARY DATA
Supplementary data to this article can be found online at https://doi.org/10.1016/j.
CONFLICTS OF INTEREST
 Paller, C.J., Antonarakis, E.S., 2013. Management of biochemically recurrent prostate cancer after local therapy: evolving standards of care and new di-rections. Clinical Advances in Hematology and Oncology 11(1):14e23.
 Ettinger, S.L., Sobel, R., Whitmore, T.G., Akbari, M., Bradley, D.R., Gleave, M.E., et al., 2004. Dysregulation of sterol response element-binding proteins and downstream effectors in prostate cancer during progression to androgen independence. Cancer Research 64(6):2212e2221.