Mi Joung Kim1, Jee Suk Lee1, Sang Eun Park2, Hye-Jin Yi1, In Gab Jeong3, Jong Soon Kang4, Jieun Yun4, Joo-Yong Lee5, Seonggu Ro6, Jung Shin Lee7, Eun Kyung Choi8, Jung Jin Hwang9, Choung-Soo Kim10. 1. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. 2. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Department of Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. 3. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Department of Urology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. 4. Bio-Evaluation Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea. 5. Department of Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Asan Institute for Life Sciences, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. 6. CrystalGenomics, Inc., Gyeonggi-Do, Republic of Korea. 7. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Department of Internal Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. 8. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Department of Radiation Oncology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Center for Development and Commercialization of Anti-Cancer Therapeutics, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. 9. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Department of Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Asan Institute for Life Sciences, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. Electronic address: jjhwang@amc.seoul.kr. 10. Institute for Innovative Cancer Research, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea; Department of Urology, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Republic of Korea. Electronic address: cskim@amc.seoul.kr.
Abstract
PURPOSE: Despite several therapeutic options renal cell carcinoma is associated with a poor clinical outcome. Therefore, we investigated whether combining 5-fluorouracil with the histone deacetylase inhibitor belinostat would exert a synergistic effect on renal cell carcinoma cells in vitro and in vivo. MATERIALS AND METHODS: We used SN12C cells treated with 5-fluorouracil and/or belinostat in vitro and in xenograft experiments in vivo. Cell viability and death mechanisms were assessed by MTS assay and Western blot. To investigate the role of reactive oxygen species we used H2DCF-DA, reactive oxygen species scavengers and the roGFP2 construct. RESULTS: Belinostat potentiated the anticancer effect of 5-fluorouracil. It synergistically induced apoptosis by activating caspases and increasing the subG1 cell population. Effects on reactive oxygen species mediated DNA damage included decreased thioredoxin expression and increased levels of TBP-2, γ-H2AX and Ac-H3. Furthermore, belinostat attenuated the 5-fluorouracil mediated induction of thymidylate synthase via HSP90 hyperacetylation. Co-administration of 5-fluorouracil with belinostat similarly reduced tumor volume and weight, and increased γ-H2AX and Ac-H3 levels in the SN12C xenograft model. CONCLUSIONS: In combination with 5-fluorouracil the targeted inhibitor of histone deacetylase synergistically inhibited renal cancer cell growth by the blockade of thymidylate synthase induction and the induction of reactive oxygen species mediated DNA damage in vitro and in vivo. Our results suggest that combined treatment with belinostat and 5-fluorouracil may represent a promising new approach to renal cancer.
PURPOSE: Despite several therapeutic options renal cell carcinoma is associated with a poor clinical outcome. Therefore, we investigated whether combining 5-fluorouracil with the histone deacetylase inhibitor belinostat would exert a synergistic effect on renal cell carcinoma cells in vitro and in vivo. MATERIALS AND METHODS: We used SN12C cells treated with 5-fluorouracil and/or belinostat in vitro and in xenograft experiments in vivo. Cell viability and death mechanisms were assessed by MTS assay and Western blot. To investigate the role of reactive oxygen species we used H2DCF-DA, reactive oxygen species scavengers and the roGFP2 construct. RESULTS:Belinostat potentiated the anticancer effect of 5-fluorouracil. It synergistically induced apoptosis by activating caspases and increasing the subG1 cell population. Effects on reactive oxygen species mediated DNA damage included decreased thioredoxin expression and increased levels of TBP-2, γ-H2AX and Ac-H3. Furthermore, belinostat attenuated the 5-fluorouracil mediated induction of thymidylate synthase via HSP90 hyperacetylation. Co-administration of 5-fluorouracil with belinostat similarly reduced tumor volume and weight, and increased γ-H2AX and Ac-H3 levels in the SN12C xenograft model. CONCLUSIONS: In combination with 5-fluorouracil the targeted inhibitor of histone deacetylase synergistically inhibited renal cancer cell growth by the blockade of thymidylate synthase induction and the induction of reactive oxygen species mediated DNA damage in vitro and in vivo. Our results suggest that combined treatment with belinostat and 5-fluorouracil may represent a promising new approach to renal cancer.