Characterization of ERK Docking Domain Inhibitors that Induce Apoptosis by Targeting Rsk-1 and Caspase-9

<p>Abstract</p> <p>Background</p> <p>The extracellular signal-regulated kinase-1 and 2 (ERK1/2) proteins play an important role in cancer cell proliferation and survival. ERK1/2 proteins also are important for normal cell functions. Thus, anti-cancer therapies that block all ERK1/2 signaling may result in undesirable toxicity to normal cells. As an alternative, we have used computational and biological approaches to identify low-molecular weight compounds that have the potential to interact with unique ERK1/2 docking sites and selectively inhibit interactions with substrates involved in promoting cell proliferation.</p> <p>Methods</p> <p>Colony formation and water soluble tetrazolium salt (WST) assays were used to determine the effects of test compounds on cell proliferation. Changes in phosphorylation and protein expression in response to test compound treatment were examined by immunoblotting and <it>in vitro </it>kinase assays. Apoptosis was determined with immunoblotting and caspase activity assays.</p> <p>Results</p> <p><it>In silico </it>modeling was used to identify compounds that were structurally similar to a previously identified parent compound, called <b>76</b>. From this screen, several compounds, termed <b>76.2</b>, <b>76.3</b>, and <b>76.4 </b>sharing a common thiazolidinedione core with an aminoethyl side group, inhibited proliferation and induced apoptosis of HeLa cells. However, the active compounds were less effective in inhibiting proliferation or inducing apoptosis in non-transformed epithelial cells. Induction of HeLa cell apoptosis appeared to be through intrinsic mechanisms involving caspase-9 activation and decreased phosphorylation of the pro-apoptotic Bad protein. Cell-based and <it>in vitro </it>kinase assays indicated that compounds <b>76.3 </b>and <b>76.4 </b>directly inhibited ERK-mediated phosphorylation of caspase-9 and the p90Rsk-1 kinase, which phosphorylates and inhibits Bad, more effectively than the parent compound <b>76</b>. Further examination of the test compound's mechanism of action showed little effects on related MAP kinases or other cell survival proteins.</p> <p>Conclusion</p> <p>These findings support the identification of a class of ERK-targeted molecules that can induce apoptosis in transformed cells by inhibiting ERK-mediated phosphorylation and inactivation of pro-apoptotic proteins.</p

Isomers of disilabenzene (C<SUB>4</SUB>Si<SUB>2</SUB>H<SUB>6</SUB>): a computational study

Computational studies using density functional theory (B3LYP) and coupled cluster (CCSD(T)) method were performed on a large number of isomers of disilabenzene, C<SUB>4</SUB>Si<SUB>2</SUB>H<SUB>6</SUB>. In all, 78 stationary points were identified, wherein 61 of them correspond to the minima on the C<SUB>4</SUB>Si<SUB>2</SUB>H<SUB>6</SUB> potential energy surface. Although the planar forms correspond to the most stable isomers, several unconventional structures are highly competitive in energy. While the framework is an important factor, the substitution pattern seems to be equally important in deciding the relative stabilities of the isomers of disilabenzenes. The relative energies of valence isomeric forms are compared with those of benzene, silabenzene, and diphosphabenzene isomers. The relative energy orderings of all the isomers are analyzed, and conclusions about the stability are drawn. Several H-bridged isomers were computed to be very stable. Several monocyclic six-membered ring structures, where the 6π-aromaticity is disturbed, also indicated the flatness of the potential energy surface of disilabenzene and their propensity toward high reactivity and facile rearrangements

The effect of bulky group substitution on the skeleton, geometries, relative energies and the reactivities of silabenzene valence isomers

Density functional theory (B3LYP) calculations were performed on the methyl and t-butyl substituted valence isomeric forms of silabenzene. The recently identified 12 valence isomeric minima on the potential energy surface of silabenzene were considered. The calculations reveal that substitution does not affect the skeleton of all the compounds including the unusual structures, V1a and V1b, located on the silabenzene potential energy surface. Similarly, the geometric parameters and relative energies show negligible differences upon substitutions by Me and t-Bu groups except for V1b. Chemical hardness and frontier orbital energy levels are used to assess the reactivity change upon substitution. The present study concludes that the substitution by Me or t-Bu will have only the steric effect and therefore, theoretical models employing the unsubstituted analogues should adequately describe the structural and energetic characteristics of the bulky group substituted silaaromatic compounds