Limits...
Identification of human IKK-2 inhibitors of natural origin (part I): modeling of the IKK-2 kinase domain, virtual screening and activity assays.

Sala E, Guasch L, Iwaszkiewicz J, Mulero M, Salvadó MJ, Pinent M, Zoete V, Grosdidier A, Garcia-Vallvé S, Michielin O, Pujadas G - PLoS ONE (2011)

Bottom Line: Notably, when these 1,061 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.We demonstrated 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 and the pharmacophore) will be made available to interested readers upon request.

View Article: PubMed Central - PubMed

Affiliation: Grup de Recerca en Nutrigenòmica, Departament de Bioquímica i Biotecnologia, Universitat Rovira i Virgili, Campus de Sescelades, Tarragona, Catalonia, Spain.

ABSTRACT

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. Consequently, there has been great interest in determining whether such molecules show biological activity toward protein targets of pharmacological relevance. One target of particular interest is hIKK-2, a serine-threonine protein kinase belonging to the IKK complex that is the primary component responsible for activating NF-κB in response to various inflammatory stimuli. Indeed, this has led to the development of synthetic ATP-competitive inhibitors for hIKK-2. Therefore, the main goals of this study were (a) to use virtual screening 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 the in vitro activity of selected natural-product hits.

Methodology/principal findings: We thus predicted that 1,061 out of the 89,425 natural products present in the studied database would inhibit hIKK-2 with good ADMET properties. Notably, when these 1,061 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 molecules from these three clusters (for which no anti-inflammatory activity has been previously described) were then selected for in vitro activity testing, in which three out of the five molecules were shown to inhibit hIKK-2.

Conclusions/significance: We demonstrated 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 and the pharmacophore) will be made available to interested readers upon request.

Show MeSH
2D structures of the VS hits for which hIKK-2-inhibition activity was tested in an in vitro assay.The IC50 and pIC50 values are also shown for those molecules that inhibit hIKK-2 and were calculated with GraphPad Prism v4.0.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3044726&req=5

pone-0016903-g004: 2D structures of the VS hits for which hIKK-2-inhibition activity was tested in an in vitro assay.The IC50 and pIC50 values are also shown for those molecules that inhibit hIKK-2 and were calculated with GraphPad Prism v4.0.

Mentions: One of the most important challenges of any VS workflow is its ability to find molecules with the required activity but without trivial similarity (in terms of chemical structure) to known active compounds. Thus, to determine which of the 1,061 potential hIKK-2 inhibitors predicted by our VS workflow could be considered as new lead molecules that may be of use in drug design and development, we merged these 1,061 NPs with the 98 known hIKK-2 inhibitors used either for validation or for pharmacophore-generation purposes, and the resulting set was classified into clusters (data not shown). Notably, three out of the ten clusters obtained consisted exclusively of NPs (i.e., Clusters 1, 2 and 3 contain 120, 88 and 38 NPs, respectively) and, therefore, these 246 molecules are scaffold-hopping candidates for hIKK-2 inhibition (see Table S3). To prove the reliability of our predictions, we selected 5 of these 246 molecules (ZINC00058225, ZINC01669260 and ZINC16946275 from Cluster 1; ZINC03683886 from Cluster 2; and ZINC03871389 from Cluster 3; see Figure 4) and tested their effects on the hIKK-2 activity using an in vitro assay (see Figure 5). The results of this experiment showed that three out of the five molecules (i.e., ZINC01669260 from Cluster 1, ZINC03683886 from Cluster 2 and ZINC03871389 from Cluster 3) inhibited hIKK-2, with IC50 values ranging from 183.8 to 3,325 µM. Thus, the highest inhibition rate achieved for ZINC03871389 (ethacridine, ethodin or rivanol) was 93% at a 15-mM inhibitor concentration. Similarly, the highest inhibition rates achieved for ZINC01669260 (lupinine) and ZINC03683886 were 91 and 92% at 10- and 15-mM inhibitor concentrations, respectively. Remarkably, a SciFinder search (Chemical Abstracts Service, Columbus, Ohio, USA; http://www.cas.org/products/sfacad) of the literature in which the three active molecules are cited revealed that none of them have been described as anti-inflammatory drugs. Therefore, this result supports the ability of the electrostatic-potential and shape comparison performed by EON to smooth differences in chemical structures and translate them into criteria important for their intermolecular interactions with the ligand-binding site (see Figure 6).


Identification of human IKK-2 inhibitors of natural origin (part I): modeling of the IKK-2 kinase domain, virtual screening and activity assays.

Sala E, Guasch L, Iwaszkiewicz J, Mulero M, Salvadó MJ, Pinent M, Zoete V, Grosdidier A, Garcia-Vallvé S, Michielin O, Pujadas G - PLoS ONE (2011)

2D structures of the VS hits for which hIKK-2-inhibition activity was tested in an in vitro assay.The IC50 and pIC50 values are also shown for those molecules that inhibit hIKK-2 and were calculated with GraphPad Prism v4.0.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3044726&req=5

pone-0016903-g004: 2D structures of the VS hits for which hIKK-2-inhibition activity was tested in an in vitro assay.The IC50 and pIC50 values are also shown for those molecules that inhibit hIKK-2 and were calculated with GraphPad Prism v4.0.
Mentions: One of the most important challenges of any VS workflow is its ability to find molecules with the required activity but without trivial similarity (in terms of chemical structure) to known active compounds. Thus, to determine which of the 1,061 potential hIKK-2 inhibitors predicted by our VS workflow could be considered as new lead molecules that may be of use in drug design and development, we merged these 1,061 NPs with the 98 known hIKK-2 inhibitors used either for validation or for pharmacophore-generation purposes, and the resulting set was classified into clusters (data not shown). Notably, three out of the ten clusters obtained consisted exclusively of NPs (i.e., Clusters 1, 2 and 3 contain 120, 88 and 38 NPs, respectively) and, therefore, these 246 molecules are scaffold-hopping candidates for hIKK-2 inhibition (see Table S3). To prove the reliability of our predictions, we selected 5 of these 246 molecules (ZINC00058225, ZINC01669260 and ZINC16946275 from Cluster 1; ZINC03683886 from Cluster 2; and ZINC03871389 from Cluster 3; see Figure 4) and tested their effects on the hIKK-2 activity using an in vitro assay (see Figure 5). The results of this experiment showed that three out of the five molecules (i.e., ZINC01669260 from Cluster 1, ZINC03683886 from Cluster 2 and ZINC03871389 from Cluster 3) inhibited hIKK-2, with IC50 values ranging from 183.8 to 3,325 µM. Thus, the highest inhibition rate achieved for ZINC03871389 (ethacridine, ethodin or rivanol) was 93% at a 15-mM inhibitor concentration. Similarly, the highest inhibition rates achieved for ZINC01669260 (lupinine) and ZINC03683886 were 91 and 92% at 10- and 15-mM inhibitor concentrations, respectively. Remarkably, a SciFinder search (Chemical Abstracts Service, Columbus, Ohio, USA; http://www.cas.org/products/sfacad) of the literature in which the three active molecules are cited revealed that none of them have been described as anti-inflammatory drugs. Therefore, this result supports the ability of the electrostatic-potential and shape comparison performed by EON to smooth differences in chemical structures and translate them into criteria important for their intermolecular interactions with the ligand-binding site (see Figure 6).

Bottom Line: Notably, when these 1,061 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.We demonstrated 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 and the pharmacophore) will be made available to interested readers upon request.

View Article: PubMed Central - PubMed

Affiliation: Grup de Recerca en Nutrigenòmica, Departament de Bioquímica i Biotecnologia, Universitat Rovira i Virgili, Campus de Sescelades, Tarragona, Catalonia, Spain.

ABSTRACT

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. Consequently, there has been great interest in determining whether such molecules show biological activity toward protein targets of pharmacological relevance. One target of particular interest is hIKK-2, a serine-threonine protein kinase belonging to the IKK complex that is the primary component responsible for activating NF-κB in response to various inflammatory stimuli. Indeed, this has led to the development of synthetic ATP-competitive inhibitors for hIKK-2. Therefore, the main goals of this study were (a) to use virtual screening 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 the in vitro activity of selected natural-product hits.

Methodology/principal findings: We thus predicted that 1,061 out of the 89,425 natural products present in the studied database would inhibit hIKK-2 with good ADMET properties. Notably, when these 1,061 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 molecules from these three clusters (for which no anti-inflammatory activity has been previously described) were then selected for in vitro activity testing, in which three out of the five molecules were shown to inhibit hIKK-2.

Conclusions/significance: We demonstrated 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 and the pharmacophore) will be made available to interested readers upon request.

Show MeSH