We provide evidence that irradiating tumors at a time point when formation of androgen-induced double-strand breaks is ongoing provides superior control when compared with radiation that is delivered when tumors are fully deprived of androgens. These results may have significant implications for altering current clinical management of intermediate-and high-risk prostate cancer treated with definitive radiotherapy.
Raju R. Raval, MD, DPhil, Mohammad Hedayati, PhD, Michael C. Haffner, MD, Yonggang Zhang, MD, Haoming Zhou, BS, Emma J. Knight, BS, Michele E. Wabler, MS, Susan Dalrymple, BS, John T. Isaacs, PhD, Aileen Santos, BS, Russell Hales, MD, Srinivasan Yegnasubramanian, MD, PhD, Theodore L. DeWeese, MD; Department of Radiation Oncology and Molecular Radiation Sciences, Department of Medical Oncology, The Johns Hopkins University School of Medicine
Purpose: Androgen suppression (AS) combined with radiation therapy is currently a standard of care in the treatment of intermediate- and high-risk prostate cancer (PCa), and this combination compared with radiation alone has been shown to improve therapeutic outcomes. Historically, the optimal sequencing of the two modalities has not been thoroughly tested. It was recently discovered by our group that androgen stimulation of PCa cells can result in the formation of topoisomerase II beta (TOP2B)-mediated DNA double-strand breaks (DSBs). Here, aiming to exploit this finding, we investigated the extent and timing of TOP2B-mediated DSBs in vitro, as stimulated by androgens. In addition, we explored the combination of androgen stimulation with radiation and its effects on tumor growth in an animal model.
Methods: Androgen-sensitive LAPC4 PCa cells were grown in androgen-free media for 3 days. Androgen (R1881) was added at a final concentration of 100 nM, and the formation of DSBs at various time points was monitored by visualization of Î³H2AX foci and by comet assay analysis. For in vivo xenografts, nude mice were castrated, and silastic implants containing androgen were implanted. Cells were injected in the mouse flank. When tumor volumes reached approximately 0.1 cm3, the implants were removed for 16 days prior to reimplantation. A total of 8 Gy of ionizing radiation was delivered 4 days prior to reimplantation (traditional group) or at 12 hours after reimplantation (experimental group). The control group did not receive radiation; tumors were measured every other day, and time to 4× initial tumor volume was determined.
Results: Addition of androgen to LAPC4 cells resulted in a significant increase in the number of Î³H2AX foci per cell: untreated (11.6 ± 3.3), 2-hour treatment (19.3 ± 5.4), and 6-hour treatment (45.2 ± 8.1) (P < .01). Comet assay (tail moments) results were consistent with Î³H2AX findings: untreated (14.3 ± 6.4), 2-hour treatment (29.8 ± 7.4), and 6-hour treatment (39.4 ± 11.5) (P < .03). The time to 4× tumor growth was significantly delayed in the experimental group (63.0 d) compared with the traditional (31.5 d) and control groups (23.0 d) (P < .01).
Conclusions: We provide evidence that irradiating tumors at a time point when formation of androgen-induced DSBs is ongoing provides superior control when compared with radiation that is delivered when tumors are fully deprived of androgens. These results may have significant implications for altering current clinical management of intermediate-and high-risk prostate cancer treated with definitive radiotherapy. There are numerous approaches that could be utilized in leading to transient increases in testosterone in patients, along with AS cycling, that are currently being tested in animal models and being proposed for human trials. In light of the new details uncovered about the mechanisms of androgen receptor (AR) signaling and downstream affects at the level of DNA processing, taking advantage of the very mechanism by which prostate cancers may grow (AR pathway) and using this knowledge to further improve therapeutic ratios in the setting of definitive radiotherapy is an attractive approach.