Inhibitors of Sulf-2 and ERK5 :modulators of cell signalling pathways as potential cancer therapeutics

Abstract

The majority of modern targeted cancer therapeutics are directed against components of cell signalling pathways. In this thesis inhibitors of two enzymes involved in cell signalling are investigated. Sulfatase-2 is a heparan sulfate processing enzyme which has been implicated as a tumour promoter in multiple cancer cell lines, with high Sulf-2 expression being an indicator of a poor clinical prognosis. A monosaccharide sulfamate inhibitor 25, based on the structure of the endogenous substrate and having an IC50 of 130 M against Sulf-2, has been published. Multiple strategies were pursued in parallel in order to attempt to identify compounds with an IC50 against Sulf-2 of < 100 M for use as chemical tools in target validation studies. These strategies included the preparation of 25, and of a set of analogues of the generic structure A, with variation of substituents at R1, R2, R3 and R4. 25 A 64 Sulf-2; IC50 130 M Access to these targets was facilitated by the development of a procedure for regioselective sulfamoylation of the O6-position of the template. Analysis of this reaction on a model substrate resulted in an increased understanding of the factors which affect the success of sulfamate formation, enabling the reaction to be used with greater efficiency on complex intermediates, and longer reaction sequences to be undertaken to provide the desired test compounds. Sets of phenolic, benzylic, and cyclic aliphatic sulfamates such as 64 were also designed in an attempt to identify a non-saccharide scaffold which could mimic the spatial arrangements of groups believed to be important for binding of the endogenous substrate to Sulf-2. Preliminary sulfatase inhibition data has been generated, indicating that analogues of 25 with superior inhibition of Sulf-2 have been identified. iv ERK5 is a soluble serine/threonine protein kinase component of a non-classical MAP kinase pathway, which has been implicated in tumour development. Breast cancer patients with high ERK5 levels have decreased disease-free survival, and ERK5 has been shown to have a role in cellular invasion and metastatic spread through affecting cell migration and attachment to the extracellular matrix. A pyrrole carboxamide series of ERK5 inhibitors was optimised from lead compounds including 212, which suffered from high clearance and low membrane permeability in vitro, and translated to poor oral bioavailability in vivo. 212 ERK5; IC50 199 nM 305 ERK5; IC50 7 nM The focus of optimisation studies was to improve ERK5 inhibitory activity and pharmacokinetic parameters to deliver compounds suitable for in vivo target validation studies. Analysis of medicinal chemistry design tactics and physicochemical properties, which had previously led to the development of successfully approved kinase drugs was used to guide compound design. The amide substituent was explored and developed into three sub-series. The first analogues prepared suffered from instability in human and mouse liver microsomes and activity against the hERG cardiac ion channel. These flaws were successfully overcome leading to compounds such as 305, which achieved an ERK5 inhibition improvement of over 20-fold, with good microsomal stability and low hERG activity. Docking of compounds into a recently published co-crystal structure of a small molecule inhibitor bound to ERK5 has provided information on the possible binding mode of the pyrrole carboxamide series.