Integrating ligand-based design and phenotypic screening to generate potent kinase inhibitors of oesophageal cancer

Abstract

Oesophageal cancer is the sixth leading cause of cancer death in the world and the eight most prevalent diagnosed cancer. Despite the improvements in diagnosis and treatment for oesophageal cancer, it remains one of the deadliest cancers worldwide, and its prognosis remains poor. Protein tyrosine kinases play an essential role in the development of different types of cancer, several of which have been found overexpressed on oesophageal cancer. Applying a combination of ligand-based design and phenotypic screening iteratively, novel potent kinase inhibitors that target oesophageal cancer were developed. This novel drug discovery approach, developed at the Innovative Therapeutics lab, served to accelerate the discovery of anticancer lead compounds with optimal drug-like properties.

Firstly, an efficient synthetic route was developed to prepare pyrazolo[3,4-d]pyrimidine derivatives with novel C3 and N1 substituents. Applying this synthetic route, several compounds were produced, followed by phenotypic screens against oesophageal cancer cell lines. This process (design-synthesis-screening) was iteratively applied to generate structure-activity-relationships which informed further series of compound design and future optimisation. A total of 76 novel pyrazolo[3,4-d]pyrimidine derivatives were synthesised and tested in different oesophageal cancer cells. From the design-synthesis-screening rounds, several active compounds were identified and 3 lead compounds with low-micromolar potencies were discovered. Broader biological evaluation of the best compounds included screening in an extended cell panel, cell cycle assay and kinome profiling. The data from these studies identified lead compounds with potent antiproliferative activity, against a wide range of oesophageal cancer cell lines, and can induce cell cycle arrest and apoptosis. The kinome profiling identified Aurora A and PIK3CA/PIK3R kinases as the potential targets of the best lead compound. This finding is consistent with the data from cell screenings and cell cycle assay, AuroraA and PIK3CA kinases are among the pivotal regulatory proteins on distinct stages of the cell cycle and have been linked to the malignancy development of oesophagel cancer. This thesis represents a promising starting point for the development of novel inhibitors of oesophageal cancer, justifying further research efforts to advance the development of the lead compound.