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The novel histone deacetylase inhibitor, N-hydroxy-7-(2-naphthylthio) hepatonomide, exhibits potent antitumor activity due to cytochrome-c-release-mediated apoptosis in renal cell carcinoma cells.

Park KC, Heo JH, Jeon JY, Choi HJ, Jo AR, Kim SW, Kwon HJ, Hong SJ, Han KS - BMC Cancer (2015)

Bottom Line: Renal tumors have been shown to have a higher global methylation percentage and reduced histone acetylation.An in vivo study showed that HNHA had greater anti-tumor and pro-apoptotic effects on RCC xenografts than the established HDAC inhibitors.These results suggest that HNHA may offer a new therapeutic approach to RCC.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Epigenetic modifications play a critical role in the regulation of all DNA-based processes, such as transcription, repair, and replication. Inappropriate histone modifications can result in dysregulation of cell growth, leading to neoplastic transformation and cell death. Renal tumors have been shown to have a higher global methylation percentage and reduced histone acetylation. Preclinical models have revealed that histone gene modifiers and epigenetic alterations play important roles in renal cell carcinoma (RCC) tumorigenesis. Recently, a novel HDAC inhibitor, N-hydroxy-7-(2-naphthylthio) heptanomide (HNHA), has been introduced as an example of a new class of anti-cancer agents. The anti-cancer activity of HNHA and the underlying mechanisms of action remain to be clarified.

Methods: The MTS assay using a panel of RCC cells was used to evaluate the anti-proliferative effects of HNHA. The established HDAC inhibitors, SAHA and TSA, were used for comparison. Western blotting analysis was performed to investigate the acetylation of histone H3 and the expression of apoptotic markers in vitro and in vivo. Subcellular fractionation was performed to evaluate expression of Bax and cytochrome c in the cytosol and mitochondria, and also translocation of cytochrome c from the cytoplasm to the nucleus. A confocal microscopic evaluation was performed to confirm inhibition of cell proliferation, induction of apoptosis, and the nuclear translocation of cytochrome c in RCC cells.

Results: In this study, we investigated the apoptosis-inducing activity of HNHA in cultured kidney cancer cells. Apoptosis in the HNHA-treated group was induced significantly, with marked caspase activation and Bcl-2 suppression in RCC cells in vitro and in vivo. HNHA treatment caused cytochrome c release from mitochondria, which was mediated by increased Bax expression and caspase activation. HNHA also induced nuclear translocation of cytochrome c, suggesting that HNHA can induce caspase-independent nuclear apoptosis in RCC cells. An in vivo study showed that HNHA had greater anti-tumor and pro-apoptotic effects on RCC xenografts than the established HDAC inhibitors.

Conclusions: HNHA has more potent anti-tumor activity than established HDAC inhibitors. Its activities are mediated by caspase-dependent and cytochrome-c-mediated apoptosis in RCC cells. These results suggest that HNHA may offer a new therapeutic approach to RCC.

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HNHA induced G0/G1 phase arrest and apoptosis in RCC cells. FACS flow cytometry with propidium iodide staining showed that HNHA induced G0/G1 phase arrest and increased the sub-G0 population in Caki-1 and A-498 cells (A). Western blotting showed that HNHA potently induced the expression of cell cycle arrest proteins (p53 and p21), reduced positive regulators of cell cycle progression (CDK4/6 and cyclin D1) and resulted in increased levels of anti-apoptotic (Bax and Apaf-1) proteins in RCC cells (B).
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Fig3: HNHA induced G0/G1 phase arrest and apoptosis in RCC cells. FACS flow cytometry with propidium iodide staining showed that HNHA induced G0/G1 phase arrest and increased the sub-G0 population in Caki-1 and A-498 cells (A). Western blotting showed that HNHA potently induced the expression of cell cycle arrest proteins (p53 and p21), reduced positive regulators of cell cycle progression (CDK4/6 and cyclin D1) and resulted in increased levels of anti-apoptotic (Bax and Apaf-1) proteins in RCC cells (B).

Mentions: To investigate the effects of HNHA on cell cycle progression, propidium iodide staining and flow cytometry were performed after treatment with several HDAC inhibitors, including HNHA. The HDAC inhibitors induced G0/G1 phase arrest and increased the sub G0 population (p < 0.05), suggesting induction of cell cycle arrest and apoptosis in RCC (Figure 3A). The data also suggested that HNHA was the most potent apoptosis inducer of those tested. To assess pro-apoptotic signaling pathways following treatment with these drugs, we evaluated the expression of cell-cycle arrest (p53 and p21) and anti-apoptotic (Bax and Apaf-1) proteins by Western blotting (Figure 3B). HNHA resulted in increased levels of the cell-cycle arrest (p53 and p21) and anti-apoptotic (Bax and Apaf-1) proteins in RCC cells. Furthermore, HNHA reduced CDK 4, CDK 6 and cyclin D1, a positive regulator of the cell cycle, which is comparable to the highest inhibition of viability in RCC cells.Figure 3


The novel histone deacetylase inhibitor, N-hydroxy-7-(2-naphthylthio) hepatonomide, exhibits potent antitumor activity due to cytochrome-c-release-mediated apoptosis in renal cell carcinoma cells.

Park KC, Heo JH, Jeon JY, Choi HJ, Jo AR, Kim SW, Kwon HJ, Hong SJ, Han KS - BMC Cancer (2015)

HNHA induced G0/G1 phase arrest and apoptosis in RCC cells. FACS flow cytometry with propidium iodide staining showed that HNHA induced G0/G1 phase arrest and increased the sub-G0 population in Caki-1 and A-498 cells (A). Western blotting showed that HNHA potently induced the expression of cell cycle arrest proteins (p53 and p21), reduced positive regulators of cell cycle progression (CDK4/6 and cyclin D1) and resulted in increased levels of anti-apoptotic (Bax and Apaf-1) proteins in RCC cells (B).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4318161&req=5

Fig3: HNHA induced G0/G1 phase arrest and apoptosis in RCC cells. FACS flow cytometry with propidium iodide staining showed that HNHA induced G0/G1 phase arrest and increased the sub-G0 population in Caki-1 and A-498 cells (A). Western blotting showed that HNHA potently induced the expression of cell cycle arrest proteins (p53 and p21), reduced positive regulators of cell cycle progression (CDK4/6 and cyclin D1) and resulted in increased levels of anti-apoptotic (Bax and Apaf-1) proteins in RCC cells (B).
Mentions: To investigate the effects of HNHA on cell cycle progression, propidium iodide staining and flow cytometry were performed after treatment with several HDAC inhibitors, including HNHA. The HDAC inhibitors induced G0/G1 phase arrest and increased the sub G0 population (p < 0.05), suggesting induction of cell cycle arrest and apoptosis in RCC (Figure 3A). The data also suggested that HNHA was the most potent apoptosis inducer of those tested. To assess pro-apoptotic signaling pathways following treatment with these drugs, we evaluated the expression of cell-cycle arrest (p53 and p21) and anti-apoptotic (Bax and Apaf-1) proteins by Western blotting (Figure 3B). HNHA resulted in increased levels of the cell-cycle arrest (p53 and p21) and anti-apoptotic (Bax and Apaf-1) proteins in RCC cells. Furthermore, HNHA reduced CDK 4, CDK 6 and cyclin D1, a positive regulator of the cell cycle, which is comparable to the highest inhibition of viability in RCC cells.Figure 3

Bottom Line: Renal tumors have been shown to have a higher global methylation percentage and reduced histone acetylation.An in vivo study showed that HNHA had greater anti-tumor and pro-apoptotic effects on RCC xenografts than the established HDAC inhibitors.These results suggest that HNHA may offer a new therapeutic approach to RCC.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Epigenetic modifications play a critical role in the regulation of all DNA-based processes, such as transcription, repair, and replication. Inappropriate histone modifications can result in dysregulation of cell growth, leading to neoplastic transformation and cell death. Renal tumors have been shown to have a higher global methylation percentage and reduced histone acetylation. Preclinical models have revealed that histone gene modifiers and epigenetic alterations play important roles in renal cell carcinoma (RCC) tumorigenesis. Recently, a novel HDAC inhibitor, N-hydroxy-7-(2-naphthylthio) heptanomide (HNHA), has been introduced as an example of a new class of anti-cancer agents. The anti-cancer activity of HNHA and the underlying mechanisms of action remain to be clarified.

Methods: The MTS assay using a panel of RCC cells was used to evaluate the anti-proliferative effects of HNHA. The established HDAC inhibitors, SAHA and TSA, were used for comparison. Western blotting analysis was performed to investigate the acetylation of histone H3 and the expression of apoptotic markers in vitro and in vivo. Subcellular fractionation was performed to evaluate expression of Bax and cytochrome c in the cytosol and mitochondria, and also translocation of cytochrome c from the cytoplasm to the nucleus. A confocal microscopic evaluation was performed to confirm inhibition of cell proliferation, induction of apoptosis, and the nuclear translocation of cytochrome c in RCC cells.

Results: In this study, we investigated the apoptosis-inducing activity of HNHA in cultured kidney cancer cells. Apoptosis in the HNHA-treated group was induced significantly, with marked caspase activation and Bcl-2 suppression in RCC cells in vitro and in vivo. HNHA treatment caused cytochrome c release from mitochondria, which was mediated by increased Bax expression and caspase activation. HNHA also induced nuclear translocation of cytochrome c, suggesting that HNHA can induce caspase-independent nuclear apoptosis in RCC cells. An in vivo study showed that HNHA had greater anti-tumor and pro-apoptotic effects on RCC xenografts than the established HDAC inhibitors.

Conclusions: HNHA has more potent anti-tumor activity than established HDAC inhibitors. Its activities are mediated by caspase-dependent and cytochrome-c-mediated apoptosis in RCC cells. These results suggest that HNHA may offer a new therapeutic approach to RCC.

Show MeSH
Related in: MedlinePlus