Insight into cancer targets and ligand binding landscape using bioinformatics and integrated molecular modeling tools.
The alarming rate of varying types of cancer diseases in human remains a global burden requiring drastic treatment in which, a prominent method of combating it is through enzymebased drug design. Metastatic castration-resistant prostate cancer (mCRPC) and triple-negative breast cancer (TNBC) are deadly forms of prostate and breast cancers, respectively. The later cancerous growth has been linked to non-receptor tyrosine (Src/p38) kinase as a potential targeted enzyme for possible chemotherapeutic control while, mCRPC have recently been linked to retinoic acid-related orphan-receptor gamma (ROR-γ). Most studies on ROR-γ usually relate it as an orphan due to low or zero possibility to identify potential inhibitor for this receptor. Amazingly, promising inhibitors of ROR-γ and their therapeutic potential were currently identified and evaluated experimentally, among which inhibitor XY018 has appreciable bioactivity. However, molecular understanding of the conformational features of XY018-ROR-γ complex is still elusive. Herein, we provide the first account of conformation details of XY018-ROR-γ using multiple computational approaches. Comparative molecular dynamics (MD) simulation of XY018-ROR-γ and hydroxycholesterol bound ROR-γ (HC9-ROR-γ) were carried out. This was widened to binding free energy calculation (MM/GBSA), principal component analysis (PCA), root mean square fluctuation (RMSF), radius of gyration (RoG) and ligand-residue interaction network. In addition, the in silico study was optimized to predict toxicity and biological activity of the identified ligand. Findings from this study revealed that: (1) hydrophobic packing contributes significantly to binding free energy, (2) Ile136 and Leu60 exhibited high hydrogen-bond in both systems, (3) XY018-ROR-γ displayed a relatively high loop region residue fluctuation compared to ROR- γ bound to natural ligand HC9-ROR-γ, (4) electrostatic interactions are potential binding force in XY018-ROR-γ complex compared to HC9-ROR-γ, (5) XY018-ROR-γ assumes a rigid conformation which is highlighted by a decrease in residual fluctuation, (6) XY018 could potentially induce pseudoporphyria, nephritis, and interstitial nephritis but potentially safe in renal failure. In vivo examination of UM-164 as a bioactive moiety against Src/p38 kinase was recently reported in literature. This ligand is a promising lead compound for developing the first targeted therapeutic strategy against triple-negative breast cancer (TNBC). However, the conformational features of UM-164 in complex with Src remained poorly explored towards the rational design of novel Src dual inhibitor. Similar to XY018-ROR-γ investigation, a comprehensive account on the conformational features of Src-UM-164 and the influence of UM-164 binding to the Src using different computational approaches was also provided. This was carried out through MD simulation, principal component analysis (PCA), thermodynamic calculations, dynamic cross-correlation (DCCM) analysis and ligand-residue interaction network profile, as well as toxicity testing. Analysis of results from this investigation revealed that: (1) the binding of UM-164 to Src induces a more stable and compact conformation on the protein structure; (2) UM-164 binding to Src induces highly correlated motions in the protein; (3) high fluctuation exhibited by the loops in Src-UM-164 system support the experimental evidence that UM-164 binds the DFGout inactive conformation of Src; (4) a relatively high binding free energy estimated for the Src-UM-164 system is affirmative of its experimental potency; (5) hydrophobic packing contributes significantly to the drug binding in Src-UM-164; (6) a relatively high H-bond formation in Src-UM-164 indicates enhanced drug-protein interaction; (7) UM-164 is relatively less toxic than Dasatinib, therefore, is potentially safer. Furthermore, a mutant form of Src was also investigated due to its drug resistivity character. Thr91 mutation was found to induce a complete loss of protein conformation required for drug fitness in c-Src. Computational studies were carried out on this mutant enzyme in complex with UM-164 as described in Src wild-type. A notable observation from binding free energy analysis results is that, a reduction in binding affinity up to -13.416 kcal/mol was estimated for this mutated candidate compared to the wild-type-UM-164. This entire work provides an invaluable contribution to the understanding of dynamics of the orphan nuclear receptor (ROR-γ) and non-receptor tyrosine kinase (Src) which could largely contribute to the design of novel inhibitors to minimise the chances of drug resistance in castrated resistance prostate cancer and triple negative breast cancer, respectively.