Targeting the tumour extracellular environment through rational modification of the SNX class of HSP90 inhibitors.

Abstract

HSP90 remains a valuable target for cancer therapy. Unfortunately, targeting intracellular HSP90 has proven not to be a viable chemotherapeutic approach. Compensatory HSR induction and HSP70 overexpression are the main limitations of this approach. A growing body of evidence suggests that targeting the extracellular environment would be of advantage and devoid of the drawbacks observed with intracellular HSP90 inhibition. As a result, the development of extracellular HSP90 inhibitors represents a novel opportunity for cancer therapeutics. In view of this hypothesis, we aimed to design and synthesise extracellular inhibitors and to assay these compounds against HSP90. To develop extracellular HSP90 inhibitors, cell-impermeable analogues of the well-developed benzamide HSP90 inhibitor (SNX 2112) were designed, synthesised and biologically evaluated. The desired target compounds were synthesized using developed methodology, as well as modified methodology. In Chapter 3 we compared and evaluated a variety of reported synthetic methods to deliver the analogues of SNX 2112. Interested in a general procedure for the synthesis of our analogues, we initially attempted to afford both the methyl and the trifluoromethyl containing analogues via a β- triketone mediated procedure. Despite the success observed with the methyl analogues, the instability of a trifluoromethyl containing β-triketone, deemed this procedure not feasible for this class of compounds. Our continued effort towards a general procedure led to the investigation of a tosylhydrazone mediated tetrahydroindazolone condensation; unexpectedly attempts to synthesise the methyl containing analogues via this procedure led to a 1—5 nitrogen to carbon tosyl migration, which was further investigated for varying substrates, and these results are explained in detail in this thesis. It then became apparent that each of the reported methods had its merits and shortcomings, there was no one best method, rather the synthetic approach was mainly determined by the C-3 substituent. The key intermediates were then converted into the desired targeted compounds by tethering the HSP90 pharmacophore to flexible alkyl groups, attached to polar sulfonate and phosphonate functionalities. Hypothetically, introduction of polar alkyl groups, would inhibit cell penetration thus limiting them to the extracellular environment. Based on the goals of our study we were interested in three biological evaluations; to confirm that our modified compounds were still capable of inhibiting HSP90s ATPase activity, to evaluate if our modifications reduced intracellular HSP90 activity, whether they stimulated the pro-oncogenic HSR, and to evaluate their cytotoxicity. Preliminary biological assessment of our compounds was consistent with our hypothesis. Here we showed that our compounds did not inhibit intracellular HSP90, and did not stimulate HSP70 expression, a marker of induction of the compensatory HSR. Furthermore, our analogues displayed cytotoxicity in the nanomolar range against the HeLa cell line. These preliminary data support the feasibility of targeting extracellular HSP90 as a novel anticancer strategy.