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Background: Licochalconce (LC) H is an artificial compound in the course of synthesizing LCC in 2013. So far, few studies on the effects of LCH have been found in the literature. Despite progress in treatment modalities for oral cancer, the cure from cancer has still limitations.Purpose: The effects of LCH were investigated on human oral squamous cell carcinoma (OSCC) cells to elucidate its mechanisms.Study Design: We explored the mechanism of action of LCH by which it could have effects on JAK2/STAT3 signaling pathway.Methods: To confirm LCH anti-cancer effect, analyzed were MTT assay, DAPI staining, soft agar, kinase assay, molecular docking simulation, flow cytometry and Western blotting analysis.Results: According to docking and molecular dynamics simulations, the predicted pose of the complex LCH and JAK2 seems reasonable and LCH is strongly bound to active JAK2 with opened activation loop. The LCH inhibitor is surrounded by specific ATP-binding pocket in which it is stabilized by forming hydrogen bonds and hydrophobic interactions. It is shown that LCH plays as a competitive inhibitor in an active state of JAK2. LCH caused a dose-dependent decrease in phosphorylation of JAK2 and STAT3. More interestingly, LCH suppressed JAK2 kinase activity in vitro by its direct binding to the JAK2. LCH significantly inhibited the JAK2/STAT3 signaling pathway, causing the down-regulation of target genes such as Bcl-2, survivin, cyclin D1, p21 and p27. In addition, LCH inhibited cell proliferation and colony formation of OSCC cells in a dose- and time-dependent manner, as well as induction of cell apoptosis through extrinsic and intrinsic pathway. The induction of apoptosis in OSCC cells by LCH was evident in the increased production of ROS, loss of mitochondrial membrane potential, release of cyto c, variation of apoptotic proteins and activation of caspase cascade.Conclusion: LCH not only induces apoptosis in OSCC cells through the JAK/STAT3 signaling pathway but also inhibits cell growth. It is proposed that LCH has a promising use for the chemotherapeutic agent of oral cancer. [ABSTRACT FROM AUTHOR]

Mitochondria play an important role in the integration and transmission of cell death signals, activating caspases and other cell death execution events by releasing apoptogenic proteins from the intermembrane space. The BCL-2 family of proteins localize (or can be targeted) to mitochondria and regulate the permeability of the mitochondrial outer membrane to these apoptotic factors. Recent evidence suggests that multiple mechanisms may regulate the release of mitochondrial factors, some of which depend on the action of caspases. [Copyright &y& Elsevier]

Members of the Bcl-2 protein family regulate apoptosis by controlling the release of apoptogenic proteins such as cytochrome c from the mitochondrial intermembrane space. Proapoptotic members induce release by increasing outer membrane permeability, while antiapoptotic members prevent this. The activity of Bcl-2 proteins depends mostly on their insertion into the mitochondrial membrane, which is reported to occur via putative channels formed by the two central hydrophobic helices. The pro- and antiapoptotic activity of Bcl-2 proteins can also be modulated by heterodimerization between antagonists through the BH3 domain of proapoptotic members, though the position of the heterodimer with respect to the membrane has never been elucidated. In this work, the membrane insertion capacity of the antiapoptotic Bcl-2 related protein Nr-13 was explored, using monolayer expansion measurements. Nr-13 penetrates into the monolayer with a molecular cross-section of 1100 A˚2, thereby implicating almost all α-helical domains of the molecule in this process. A mutant protein, bearing neutral instead of acidic residues in the loop between the two putative channel-forming fifth and sixth α-helices, retained the ability to interact with the lipid monolayer, suggesting that the membrane insertion of Nr-13 is not exclusively α5-α6-dependent. In contrast, the specific interaction of Nr-13 with the monolayer was prevented by heterodimer formation with the BH3 domain of proapoptotic Bax. These findings are discussed in terms of a model for monolayer insertion in which the antiapoptotic Nr-13 and proapoptotic proteins exert their antagonistic effects by preventing each other from reaching the membrane. [Copyright &y& Elsevier]

To detect the posttranslational N-myristoylation of caspase substrates, the susceptibility of the newly exposed N-terminus of known caspase substrates to protein N-myristoylation was evaluated by in vivo metabolic labeling with [3H]myristic acid in transfected cells using a fusion protein in which the query sequence was fused to a model protein. As a result, it was found that the N-terminal nine residues of the newly exposed N-terminus of the caspase-cleavage product of cytoskeletal actin efficiently direct the protein N-myristoylation. Metabolic labeling of COS-1 cells transiently transfected with cDNA coding for full-length truncated actin (tActin) revealed the efficient incorporation of [3H]myristic acid into this molecule. When COS-1 cells transiently transfected with cDNA coding for full-length actin were treated with staurosporine, an apoptosis-inducing agent, an N-myristoylated tActin was generated. Immunofluorescence staining coupled with MitoTracker or fluorescence tagged-phalloidin staining revealed that exogenously expressed tActin colocalized with mitochondria without affecting cellular and actin morphology. Taken together, these results demonstrate that the C-terminal 15 kDa fragment of cytoskeletal actin is posttranslationally N-myristoylated upon caspase-mediated cleavage during apoptosis and targeted to mitochondria. [Copyright &y& Elsevier]