Background:Heat shock protein 90 (HSP90) has a key role in the maintenance of the cellular proteostasis. However, HSP90 is also involved in stabilisation of oncogenic client proteins and facilitates oncogene addiction and cancer cell survival. The development of HSP90 inhibitors for cancer treatment is an area of growing interest as such agents can affect multiple pathways that are linked to all hallmarks of cancer. This study aimed to test the hypothesis that targeting cysteine residues of HSP90 will lead to degradation of client proteins and inhibition of cancer cell proliferation.Methods:Combining chemical synthesis, biological evaluation, and structure–activity relationship analysis, we identified a new class of HSP90 inhibitors. Click chemistry and protease-mass spectrometry established the sites of modification of the chaperone.Results:The mildly electrophilic sulphoxythiocarbamate alkyne (STCA) selectively targets cysteine residues of HSP90, forming stable thiocarbamate adducts. Without interfering with the ATP-binding ability of the chaperone, STCA destabilises the client proteins RAF1, HER2, CDK1, CHK1, and mutant p53, and decreases proliferation of breast cancer cells. Addition of a phenyl or a tert-butyl group in tandem with the benzyl substituent at nitrogen increased the potency. A new compound, S-4, was identified as the most robust HSP90 inhibitor within a series of 19 derivatives.Conclusion:By virtue of their cysteine reactivity, sulphoxythiocarbamates target HSP90, causing destabilisation of its client oncoproteins and inhibiting cell proliferation.

e14591 Background: AT-101 is an orally active pan Bcl-2 inhibitor currently under clinical development in Phase II trials. We investigated the effect of AT-101 alone or in combination with chemotherapies or targeted kinase inhibitors in NSCLC both in vitro and in vivo. Methods: Growth inhibition was measured by WST assays. The CalculSyn method was used to assess drug interaction by calculating the Combination Index (CI) value. The ability of AT-101 to potentiate the anti-cancer effect of chemotherapies or kinase inhibitors was evaluated in xenograft models. Results: A panel of 11 NSCLC cell lines with overexpression of Bcl-2, Bcl- XL or Mcl-1 proteins was treated with AT-101, docetaxel, pemetrexed, erlotinib, sorafenib, sunitinib, rapamycin, as a single agent and in combination. AT-101 inhibited the growth with IC50 at 3–9 uM. When treating cells simultaneously with both agents, AT-101 demonstrated strong synergy with those agents in A549 or H460 cells with CI values < 1.0. Protein analysis results indicated that AT-101 caused apoptosis by a time- and dose-dependent induction of Noxa expression in those cells. The expression of Bcl- XL was not influenced by AT-101. Pro-Caspase-3 was reduced with increasing doses of AT-101. In vivo, combined treatment of AT-101 with docetaxel, erlotinib, or sorafenib synergistically suppressed subcutaneous NSCLC A549 cells tumor growth compared with treatment with either agent alone. The synergist effects with the ErbB1 inhibitor Erlotinib is associated with the overexpression of the target protein ErbB1 in those cells. Only combination therapy resulted in significant tumor growth delay and no significant toxicities were observed. Conclusions: Our results demonstrate that AT-101significantly enhances the anti-tumor activity of chemotherapy and targeted agents and may represent a promising new anticancer agent with a novel molecular mechanism. Molecular targeted therapy with AT-101 may improve the outcome of current chemotherapy for NSCLC with Bcl-2, Bcl-xL and/or Mcl-1 overexpression. [Table: see text]