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.
|Yu, Wenbo; MacKerell Jr, Alexander D (2017) Computer-Aided Drug Design Methods. Methods Mol Biol 1520:85-106|
|Melville, Zephan; Hernández-Ochoa, Erick O; Pratt, Stephen J P et al. (2017) The Activation of Protein Kinase A by the Calcium-Binding Protein S100A1 Is Independent of Cyclic AMP. Biochemistry 56:2328-2337|
|Cavalier, Michael C; Melville, Zephan; Aligholizadeh, Ehson et al. (2016) Novel protein-inhibitor interactions in site 3 of Ca(2+)-bound S100B as discovered by X-ray crystallography. Acta Crystallogr D Struct Biol 72:753-60|
|Roth, Braden M; Varney, Kristen M; Rustandi, Richard R et al. (2016) (1)H(N), (13)C, and (15)N resonance assignments of the CDTb-interacting domain (CDTaBID) from the Clostridium difficile binary toxin catalytic component (CDTa, residues 1-221). Biomol NMR Assign 10:335-9|
|Cavalier, Michael C; Ansari, Mohd Imran; Pierce, Adam D et al. (2016) Small Molecule Inhibitors of Ca(2+)-S100B Reveal Two Protein Conformations. J Med Chem 59:592-608|
|Faller, Christina E; Raman, E Prabhu; MacKerell Jr, Alexander D et al. (2015) Site Identification by Ligand Competitive Saturation (SILCS) simulations for fragment-based drug design. Methods Mol Biol 1289:75-87|
|Yu, Wenbo; Lakkaraju, Sirish Kaushik; Raman, E Prabhu et al. (2015) Pharmacophore modeling using site-identification by ligand competitive saturation (SILCS) with multiple probe molecules. J Chem Inf Model 55:407-20|
|Bresnick, Anne R; Weber, David J; Zimmer, Danna B (2015) S100 proteins in cancer. Nat Rev Cancer 15:96-109|
|Lakkaraju, Sirish Kaushik; Yu, Wenbo; Raman, E Prabhu et al. (2015) Mapping functional group free energy patterns at protein occluded sites: nuclear receptors and G-protein coupled receptors. J Chem Inf Model 55:700-8|
|He, Xinhua; Lakkaraju, Sirish K; Hanscom, Marie et al. (2015) Acyl-2-aminobenzimidazoles: a novel class of neuroprotective agents targeting mGluR5. Bioorg Med Chem 23:2211-20|
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