In malignant melanoma (MM), we have shown that elevated levels of the tumor marker, S100B, binds directly to wild-type (wt) p53, dissociates the p53 tetramer, enhances hdm2-dependent ubiquitination of p53, and down-regulates p53-dependent tumor suppression functions;therefore, it is important to develop S100B inhibitors (SBiXs;X=compound number) to restore active p53 in this deadly cancer. As a proof of principle for a drug design project, inhibiting S100B with small interfering antisense RNA (siRNAS100B) and several SBiXs was shown to restore wild-type p53 levels and its downstream gene products, as necessary to induce cell growth arrest and apoptosis in malignant melanoma. We hypothesize that low molecular weight compounds can be designed to bind the well-defined p53 binding site on Ca2+ loaded S100B with higher affinity and specifically inhibit the Ca2+dependent S100B-p53 interaction to mimic the siRNAS100B effects. In the last granting period, thirty-eight publications, ten patent applications, and twenty protein data base submissions describe our progress at addressing these goals/hypotheses. Animal model studies with several SBiXs and a human clinical trial with SBi1 are also ongoing as a result of our progress. However, it is important that we engineer and/or synthesis new and improved SBiXs with higher affinity and better specificity towards inhibiting S100B. This will be achieved with the following specific aims:
In Aim 1, computer aided drug design (CADD) combined with high-throughput screening methods (i.e. NMR, binding, and cellular assays) will be used to discover/design new compounds that bind S100B and inhibit the S100B-p53 interaction at lower concentrations and with higher specificity than those already discovered. Optimization of promising leads will be performed via chemical modifications as guided by 3D structural and computer aided drug design (CADD) lead optimization approaches.
In Aim 2, 3D structures of S100B-SBiX complexes will be determined using NMR spectroscopy and/or X-ray crystallography to further characterize the binding surface on Ca2+S100B, so improved SBiXs can be designed and synthesized. Testing new analogues will be performed using existing thermodynamic binding and biological assays (as in Aim 1) with the most promising compounds examined for their ability to suppress/eliminate tumor growth in melanoma mouse models (in Aim 3). The in vivo data will be important for focusing our design/synthesis efforts on lead compounds or classes of compounds that have efficacy in vivo, and they will be used to set priorities for additional human clinical trials to be done in the future. It is our goal to discover/synthesize and/or to improve existing SBiXs to optimally restore p53 activity in human malignant melanoma. SBiXs may also have therapeutic value for treating other cancers with elevated S100B and wt p53 such as astrocytomas, renal tumors, and some forms of leukemia, so this will be explored in future endeavors.
The ongoing project (CA107331) addresses important public health concerns including understanding how the well-established tumor marker, S100B, contributes to cancer progression by down-regulating the tumor suppressor protein p53. This information impacts how we develop drugs to treat several cancers, including malignant melanoma, which has elevated levels of the S100B growth factor. Such molecules will be tested both in vitro and in vivo as necessary to initiate human clinical trials, as was done in the past with one S100B inhibitor, SBi1.
|Cavalier, Michael C; Pierce, Adam D; Wilder, Paul T et al. (2014) Covalent small molecule inhibitors of Ca(2+)-bound S100B. Biochemistry 53:6628-40|
|Hartman, Kira G; Vitolo, Michele I; Pierce, Adam D et al. (2014) Complex formation between S100B protein and the p90 ribosomal S6 kinase (RSK) in malignant melanoma is calcium-dependent and inhibits extracellular signal-regulated kinase (ERK)-mediated phosphorylation of RSK. J Biol Chem 289:12886-95|
|Mazalouskas, Matthew D; Godoy-Ruiz, Raquel; Weber, David J et al. (2014) Small G proteins Rac1 and Ras regulate serine/threonine protein phosphatase 5 (PP5)ýýextracellular signal-regulated kinase (ERK) complexes involved in the feedback regulation of Raf1. J Biol Chem 289:4219-32|
|Yu, Wenbo; Lakkaraju, Sirish Kaushik; Raman, E Prabhu et al. (2014) Site-Identification by Ligand Competitive Saturation (SILCS) assisted pharmacophore modeling. J Comput Aided Mol Des 28:491-507|
|Dhar, Amlanjyoti; Mallick, Shampa; Ghosh, Piya et al. (2014) Simultaneous inhibition of key growth pathways in melanoma cells and tumor regression by a designed bidentate constrained helical peptide. Biopolymers 102:344-58|
|Hartman, Kira G; McKnight, Laura E; Liriano, Melissa A et al. (2013) The evolution of S100B inhibitors for the treatment of malignant melanoma. Future Med Chem 5:97-109|
|Raman, E Prabhu; MacKerell Jr, Alexander D (2013) Rapid estimation of hydration thermodynamics of macromolecular regions. J Chem Phys 139:055105|
|Zimmer, Danna B; Lapidus, Rena G; Weber, David J (2013) In vivo screening of S100B inhibitors for melanoma therapy. Methods Mol Biol 963:303-17|
|Donato, R; Cannon, B R; Sorci, G et al. (2013) Functions of S100 proteins. Curr Mol Med 13:24-57|
|Raman, E Prabhu; Yu, Wenbo; Guvench, Olgun et al. (2011) Reproducing crystal binding modes of ligand functional groups using Site-Identification by Ligand Competitive Saturation (SILCS) simulations. J Chem Inf Model 51:877-96|
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