Structure-Based Discovery of Receptor Activator of Nuclear Factor-κB Ligand (RANKL)-Induced Osteoclastogenesis Inhibitors.

Receptor activator of nuclear factor-κB ligand (RANKL) has been actively pursued as a therapeutic target for osteoporosis, given that RANKL is the master mediator of bone resorption as it promotes osteoclast differentiation, activity and survival. We employed a structure-based virtual screening approach comprising two stages of experimental evaluation and identified 11 commercially available compounds that displayed dose-dependent inhibition of osteoclastogenesis. Their inhibitory effects were quantified through TRAP activity at the low micromolar range (IC50 < 5 µΜ), but more importantly, 3 compounds displayed very low toxicity (LC50 > 100 µΜ). We also assessed the potential of an N-(1-aryl-1H-indol-5-yl)aryl-sulfonamide scaffold that was based on the structure of a hit compound, through synthesis of 30 derivatives. Their evaluation revealed 4 additional hits that inhibited osteoclastogenesis at low micromolar concentrations; however, cellular toxicity concerns preclude their further development. Taken together with the structure-activity relationships provided by the hit compounds, our study revealed potent inhibitors of RANKL-induced osteoclastogenesis of high therapeutic index, which bear diverse scaffolds that can be employed in hit-to-lead optimization for the development of therapeutics against osteolytic diseases.

Rationally designed less toxic SPD-304 analogs and preliminary evaluation of their TNF inhibitory effects.

SPD-304 was discovered as a promising tumor necrosis factor alpha (TNF) antagonist that promotes dissociation of TNF trimers and therefore blocks the interaction of TNF and its receptor. However, SPD-304 contains a potentially toxic 3-alkylindole moiety, which can be bioactivated to a reactive electrophilic intermediate. A series of SPD-304 analogs was synthesized with the aim to diminish its toxicophore groups while maintaining the binding affinity for TNF. Incorporation of electron-withdrawing substituents at the indole moiety, in conjunction with elimination of the 6'-methyl group of the 4-chromone moiety, led to a significantly less toxic and equally potent TNF inhibitor.

Aqueous Solubility Enhancement for Bioassays of Insoluble Inhibitors and QSPR Analysis: A TNF-α Study.

The aim of this study is to improve the aqueous solubility of a group of compounds without interfering with their bioassay as well as to create a relevant prediction model. A series of 55 potential small-molecule inhibitors of tumor necrosis factor-alpha (TNF-α; SPD304 and 54 analogues), many of which cannot be bioassayed because of their poor solubility, was used for this purpose. The solubility of many of the compounds was sufficiently improved to allow measurement of their respective dissociation constants (Kd). Parameters such as dissolution time, initial state of the solute (solid/liquid), co-solvent addition (DMSO and PEG3350), and sample filtration were evaluated. Except for filtration, the remaining parameters affected aqueous solubility, and a solubilization protocol was established according to these. The aqueous solubility of the 55 compounds in 5% DMSO was measured with this protocol, and a predictive quantitative structure property relationship model was developed and fully validated based on these data. This classification model separates the insoluble from the soluble compounds and predicts the solubility of potential small-molecule inhibitors of TNF-α in aqueous solution (containing 5% DMSO as co-solvent) with an accuracy of 81.2%. The domain of applicability of the model indicates the type of compounds for which estimation of aqueous solubility can be confidently predicted.

ICPBC and C12-ICPBC: two new red emitting, fluorescent Ca2+ indicators excited with visible light.

Two new, visible-excited and red-emitting fluorescent Ca(2+) indicators were synthesized and the spectral profiles of their free and Ca(2+) bound forms were studied. The fluorescent properties of these probes are due to the extended conjugation of the chromeno[3',2':3,4]pyrido[1,2a][1,3]benzimidazole chromophore incorporated in their BAPTA-type, Ca(2+) chelating structure. The compounds, namely ICPBC and its N-dodecyl analog C12-ICPBC exhibit Ca(2+) dissociation constants of 7.7 and 18.0 microM, respectively. The fluorescence spectra of the probes showed a clear shift in excitation wavelength maxima upon Ca(2+) binding along with a large Stokes shift and changes in fluorescence intensity, indicating their potential use as Ca(2+) indicators. The ability of ICPBC to trace high calcium spikes was tested in the human HepG2 cell line with positive results.

Component-selective and stereocontrolled one-step three-component reaction among aldehydes, amines, and allenyl boronic acids or allenyl pinacolboronates.

A one-step, three-component condensation of allenyl boronic acids or allenyl pinacolboronates with amines and aldehydes affords α-allenyl or α-propargyl α-amino acids and anti-ß-amino alcohols. This process gives the allenyl or propargyl product depending on the amine and boron components. Secondary amines generate exclusively α-allenyl α-amino acids, while primary aliphatic amines lead to α-propargyl α-amino acids. Secondary aliphatic amines react with chiral α-hydroxy aldehydes and allenyl boron derivatives to form stereoselectively allenyl anti-ß-amino alcohol products.