|Abstract or Summary
- Histone deacetylase (HDAC) inhibitors have the potential to de-repress epigenetically silenced genes in cancer cells, leading to cell cycle arrest and apoptosis. Dietary HDAC inhibitors derived from natural phytochemicals are promising anticancer agents. In this thesis, metabolites from natural organosulfur and organoselenium compounds, i.e. allyl mercaptan (AM), β-methylselenopyruvate (MSP) and α-keto-γ-methylselenobutyrate (KMSB), were discovered to serve as HDAC inhibitors and exhibit anticancer activities in human colon cancer cells.
AM is a metabolite of garlic-derived organosulfur compounds, whereas MSP and KMSB are the newly discovered α-keto acid metabolites of Se-methylselenocysteine (MSC) and selenomethionine (SM) respectively. In this thesis research, all three compounds were shown to inhibit HDAC activity in a competitive manner at micromolar levels. Molecular modeling suggested they can fit into the active site of HDAC enzymes and chelate catalytic Zn²⁺ via sulfhydryl group (AM) or keto acid group (MSP and KMSB). Studies on the structural analogs indicated that the selenium atom was also important for MSP/KMSB's HDAC inhibitory effects.
In human colon cancer cells, AM, MSP and KMSB decreased HDAC activities, and induced rapid histone hyperacetylation in a dose-dependent manner. All three compounds induced rapid and sustained expression of the cell cycle inhibitor p21 at both mRNA and protein levels. There was enhanced P21 promoter activity, and hyperacetylated histone H3 was associated with the gene promoter. The induction of p21 required a Sp1/Sp3 binding sites but was independent of p53 status. P21 induction may mediate cell cycle arrest in AM/MSP/KMSB-treated colon cancer cells. MSP and KMSB also induced apoptosis in colon cancer cells, as evidenced by morphological changes, Annexin V staining and increased cleaved caspase-3, -6, -7, -9 and poly(ADP-ribose)polymerase. MSP dramatically induced the expression of pro-apoptotic Bcl-2 family gene Bmf, and knocking down Bmf expression by siRNA significantly decreased caspase activation in MSP-treated colon cancer cells. As a result of cell cycle arrest and/or apoptosis induction, these compounds significantly inhibited colon cancer cell growth.
Formation of MSP was directly detected in MSC-treated colon cancer cells. MSC, the parent compound also induced histone hyperacetylation, p21 and Bmf expression in the cells. Knocking down Bmf expression reduced MSC's apoptotic effects. In colon cancer cells, SM cannot be converted to KMSB, and histone acetylation remained unchanged in SM-treated colon cancer cells. Histone hyperacetylation was also observed in the tissues of the mice gavaged with AM and its parent organosulfur compounds. These results indicate that AM/MSP/KMSB could be active metaobolites of organosulfur or organoselenium compounds contributing to their chemopreventive effects.