Background Their large scaffold diversity and properties such as structural complexity and drug similarity form the basis of claims that natural products are ideal starting points for drug design and development. to the development of synthetic ATP-competitive inhibitors for hIKK-2. Therefore the main goals of this study were (a) to use virtual screening Paeoniflorin to identify potential hIKK-2 inhibitors of natural origin that compete with ATP and (b) to evaluate the reliability of our virtual-screening protocol by experimentally testing Paeoniflorin the activity of selected natural-product hits. Methodology/Principal Findings We thus predicted that 1 61 out of the 89 425 natural products present in the studied database would inhibit hIKK-2 with good ADMET properties. Notably when these 1 61 molecules were merged with the 98 synthetic hIKK-2 inhibitors used in this study and the resulting set was classified into ten clusters according to chemical similarity there were three clusters that contained only natural products. Five Paeoniflorin molecules from these three clusters (for which no anti-inflammatory activity has been previously described) were then selected for activity testing in which three out of the five molecules were shown to inhibit hIKK-2. Conclusions/Significance We exhibited that our virtual-screening protocol was successful in identifying lead compounds for developing new inhibitors for hIKK-2 a target of great interest in medicinal chemistry. Additionally all the tools developed during the current study (i.e. the homology model for the hIKK-2 kinase domain name and the pharmacophore) will be made available to interested readers upon request. Introduction Natural products (NPs) are a valuable source of inspiration as lead compounds for the design and development of new drug candidates [1]. In fact Mouse monoclonal to WNT5A over 60% of the current anticancer drugs are natural-product-related molecules (activity of selected NP hits. To achieve these goals we (1) developed a homology model for the hIKK-2 kinase domain name which could stand the test of our validation criteria (2) docked ATP-competitive molecules known to be potent and specific inhibitors of hIKK-2 with this model [10] [11] [13] [15] [16] [18] [20]-[31] (3) identified which of the resulting poses were by analyzing whether they satisfied the experimentally known generic binding Paeoniflorin features of ATP-competitive inhibitors of kinases [45] (4) used the knowledge-based coherent poses to derive a structure-based common pharmacophore made up of the key intermolecular interactions between hIKK-2 and its inhibitors (5) obtained exclusion volumes from our homology model and added them to the pharmacophore (6) validated the selectivity of the resulting pharmacophore and of the VS process using a large database of kinase decoys [46] and ATP-competitive inhibitors for hIKK-2 that were not used during the pharmacophore building [47] (7) used the previously validated structure-based pharmacophore and VS protocol to find ATP-competitive inhibitors for hIKK-2 in a database of NPs [48] and finally (8) proved the reliability of the prediction by testing the inhibitory effect of some selected hits on hIKK-2 [18]) (b) one hydrogen bond between its amide group and the side-chain hydroxyl group of Tyr98 (c) one hydrogen bond between the nitrogen from the amide group and the backbone oxygen atom of Gln100 and (d) one hydrogen bond on the other side of the binding pocket with the Asn150 and Asp166 side chains. Here we Paeoniflorin note that the relevance of Cys99 and Gln100 in this intermolecular conversation has been reported [18]. Furthermore inhibitor 12 has hydrophobic interactions with the Leu21 Val29 Ala42 Asp145 Val152 and Ile165 side chains. Inhibitor 4a [23] (Physique 2E) is usually a pyridine derivate that has a very different chemical scaffold than the other inhibitors studied and moreover it belongs to a family of very active hIKK-2 inhibitors. The Cys99 backbone atoms are involved in two hydrogen-bonding interactions Paeoniflorin one of which is between the nitrogen and the hydroxyl group in the 2′ position of the benzyl moiety of 4a (which is an important group for this family of hIKK-2 inhibitors [22]-[24] [37]) and the other is between the carbonyl oxygen and one of two amine.