Design and Synthesis of Novel Pteridine Derivatives as Aurora Kinase Inhibitors for the Treatment of Cancer

Abstract

World-wide report of cancer risk and its mortality suggested that cancer has 2nd highest death rate after metabolic disorder. Anti-mitotic drugs are well known from the 1980s as chemotherapeutic agents. These molecules perturb the cell division process and proved to be effective in anticancer therapies. Aurora kinase recently became the most interesting target for development of potential kinase inhibitors. Some crucial functions of Aurora kinase are centrosome maturation, spindle formation, segregation and proper alignment of chromosomes. Furthermore, inhibition of Aurora kinases led to cell cycle delay and G2/M arrest followed by cell death. Many small molecules are already under the clinical development but still not able to make its way to market. In this study, design, synthesis and pharmacological screening was performed for the hit identification. Diversified eleven molecules with pyrimidine common core were used to developed pharmacophore using GALHAD module of Sybyl X. Unique pharmacophore, consisting of eight features, was validated by GH scoring and ROC analysis. Validated pharmacophore was utilized for virtual screening process to identify sub structural similarities using NCI database. From the results of Qfit value, obtained by NCI database, pteridine was selected as a scaffold for the development of novel derivatives. Apart from this, 3D-QSAR study was also performed on 56 thienopyridine derivatives to identify the various substitutions required around the scaffold. Knowledge based SAR study was applied and 2-(4-hydroxypteridin-2-ylthio)acetamide core designed as Aurora-B inhibitors. Different substructures having pteridine scaffold were docked on Aurora-B (PDB ID: 4C2V) and studied for their binding modes. Best designed series from the results of molecular docking were synthesised and confirmed by spectroscopic techniques like 1H & 13C NMR, FT-IR, Mass and purity was checked by HPLC. All molecules were screened in-vitro over seven different cancer cell lines and in Aurora-B enzymatic assay. Results of the cytotoxicity on cancer cell lines showed IC50 in range of 0.2 μM to 100 μM. Toxicity of all molecules were also checked using normal (VERO) cell line and most of compounds were found to be non-toxic (>100 μM). Further screening of most active inhibitors were evaluated by colony forming assay where inhibition was visualized as proof of concept. Best four molecules were further estimated in cell signaling inhibition using cell cycle analysis (PI staining) and found G2/M arrest in cancer cells. Apoptosis assay was also performed using Annexin-V/FITC staining and produced early apoptosis in low μM range. Best four molecules were also screened in in-vivo animal models. Preliminary those molecules were checked in p388/D1 murine luekaemia model for increment of life span study and best two molecules were found most active with increase lifespan of greater than 60% compared to disease control group. Those two molecules were evaluated in early stage subcutaneous animal model where cancer cells injected in the Balb/C mice and found greater than 55% tumor growth inhibition in both molecules compared to disease control group. Serological parameters (GGT, LDH, CK-MB) were also evaluated as reduction or less cancerogenic condition in biological system. After histopathological examination of tumors, it was found less tumorigenic condition with slow tumor growth in treatment group compared with disease control animal tumors. At the end, developed hit molecules can inhibit cancer progression with better pharmacological activity and can be improved pharmacokinetic efficacy in future.