Design and synthesis of small-molecule inhibitors of two cancer targets :mTor and MDM2/p53

Abstract

Cancer is a disease in which cellular control over growth and differentiation has been lost. Targeted therapies for the treatment of cancer are becoming an increasing area of focus within the pharmaceutical industry and academia, due to the increasing understanding of the biology behind tumourgenesis.

There are 518 protein kinases within the human genome and they play a significant role within cellular signalling. Aberrant signalling of kinases can contribute to the development of cancer, and inhibition of kinase targets can result in either a cytostatic or cytotoxic effort. Kinases have a discrete ATP-binding domain, which presents an ideal target for small- molecule inhibitors. mTOR (mammalian target of rapamycin) is a serine/threonine protein kinase, which forms two complexes, mTORC1 and mTORC2, as part of the PI 3-K/Akt pathway, a growth/survival pathway which has aberrant signalling in a number of cancers.

The development of ATP-competitive inhibitors of mTOR has been based on a series of 2,6- diaminosubstituted pyrimidines, with modest activity against mTOR, as exemplified by NU6027 and NU6227 originally designed as CDK2 inhibitors. Structure-activity relationships for inhibition of mTOR have been explored. The 4-substituent was either modified to a smaller alkoxy group or completely removed, giving reduced activity. At the 5- position compounds with other substituents were then synthesised. Modifications of the 2- and 6-amino groups were also investigated. The pyrimidine heterocycle was also replaced with two pyridine regioisomers. None of the synthesised compounds showed improved mTOR inhibitory activity over NU6227 and NU6027.

The tumour suppressor p53 is activated as a response to DNA-damage, oncogene activation and cellular stress. Once activated p53 acts as a transcription initiator, inducing thetranscription of a number of genes, including those involved in halting the cell cycle, repairing DNA-damage and initiating apoptosis. A further transcriptional target of p53 is MDM2, a negative regulator of p53. Inhibitors of the MDM2-p53 interaction have been reported including the isoindolinones e.g. NCL-00008406.

Structure-activity relationships (SAR) studies for the identification of replacement of the 4- nitro group showed that a 4-ethynyl substituent had a similar level of potency, along with the 4-bromo, 3-fluoro substituents. SARs around the isoindolinone A-ring identified the 6-tert- butyl substituent as equipotent to the parent. Synthesis of an oxetane derivative such as NCL- 00018327 has demonstrated that replacing the cyclopropyl group with a 3,3-oxetane substituent either maintained or improved activity against MDM2.

Synthetic efforts have identified highly potent, low nanomolar, isoindolinone-based inhibitors of the MDM2-p53 protein-protein interaction. Optimal substituents for the benzyl group have been identified, avoiding the use of a nitro group which is toxic within drugs. Synthesis of an oxetane derivative has been shown to be either equi- or more potent than cyclopropyl derivatives, and this modification is predicted to improved the aqueous solubility by reducing the clogP. Synthesis of a tert-butyl analogues using an alternative synthetic route has developed the SAR around the A-ring and resulted in an improved synthetic scheme. Purification of a range of MDM2 mutant proteins has identified a crystallisable form of MDM2 and the recently solved crystal structure of an isoindolinone bound to MDM2 should guide further improvements to potency and aid the incorporation of groups to improve physical properties.

Cancer is a disease in which control over cellular growth is deregulated. Synthetic efforts have been directed toward the development of two targeted anti-cancer agents; the kinase mTOR and the protein-protein interaction MDM2/p53. 2,6-diaminopyrimidine were identified as most inhibitors of mTOR, but demonstrated a flat SAR and no increase in potency was observed. Isoindolinones have been identified as a valuable scaffold to inhibit the protein-protein interaction and have demonstrated excellent potencies.