Novologues Containing a Benzamide Side Chain Manifest Anti-proliferative Activity Against Two Breast Cancer Cell Lines

The heat shock protein 90 (Hsp90) folding machinery is essential for the maturation of nascent polypeptides into their biologically active three-dimensional-structures and for the rematuration/clearance of misfolded proteins that form under cellular stress.1–3 As a prosurvival chaperone, Hsp90 overexpression is commonly observed in transformed cells, which is required to sustain the hostile tumor micro-environment associated with nutrient deprivation and hypoxia. Pharmacological inhibition of Hsp90 has been shown to induce the degradation of oncogenic proteins associated with all six hallmarks of cancer that rely upon Hsp90.4–8 Consequently, Hsp90 represents a highly sought after target for the treatment of cancer. In fact, 17 small molecules that bind competitively to the N-terminal ATP-binding pocket are under clinical evaluation against various cancers.9,10 However, heat shock factor 1 (HSF-1), the master regulator of the pro-survival heat shock response also binds Hsp90. Ultimately, Hsp90 N-terminal inhibition results in HSF-1 release, and upon phosphorylation, trimerizes and translocates to the nucleus wherein it binds the heat shock elements to activate the pro-survival, heat shock response (HSR). The HSR serves to expand the cellular buffering capacity and to assist in the maturation of mutated and oncogenic substrates.11 This concomitant heat shock response is detrimental to the treatment of cancer and may lead to drug resistance and tumor metastasis.12 Recent studies have demonstrated that allosteric modulation of the Hsp90 C-terminus can separate the pro-survival heat shock response from pro-apoptotic, client protein degradation.13–20 Two classes of small molecules derived from novobiocin 1, (Figure 1) the first identified Hsp90 C-terminal inhibitor, were discovered via the structure-activity relationship studies. KU-32 (2), which lacks a 4-hydroxyl, the 3’-carbamate, and contains an acetamide in lieu of a prenylated benzamide, represents a lead compound that induces the heat shock response at concentrations much lower than that needed for client protein degradation.2,21 Consequently, this class of analogues has been evaluated as neuroprotective agents to refold protein aggregates.22–24 In contrast, KU-174 (3) contains a biarylamide side chain in lieu of the acetamide, and induces Hsp90 client protein degradation without induction of the heat shock response.25–26 Therefore, this class of novobiocin analogues manifests optimal properties for the treatment of cancer, as no HSR is observed with such compounds

Modulating Molecular Chaperones Improves Mitochondrial Bioenergetics and Decreases the Inflammatory Transcriptome in Diabetic Sensory Neurons

We have previously demonstrated that modulating molecular chaperones with KU-32, a novobiocin derivative, ameliorates physiologic and bioenergetic deficits of diabetic peripheral neuropathy (DPN). Replacing the coumarin core of KU-32 with a meta-fluorinated biphenyl ring system created KU-596, a novobiocin analogue (novologue) that showed neuroprotective activity in a cell-based assay. The current study sought to determine whether KU-596 offers similar therapeutic potential for treating DPN. Administration of 2–20 mg/kg of KU-596 improved diabetes induced hypoalgesia and sensory neuron bioenergetic deficits in a dose-dependent manner. However, the drug could not improve these neuropathic deficits in diabetic heat shock protein 70 knockout (Hsp70 KO) mice. To gain further insight into the mechanisms by which KU-596 improved DPN, we performed transcriptomic analysis of sensory neuron RNA obtained from diabetic wild-type and Hsp70 KO mice using RNA sequencing. Bioinformatic analysis of the differentially expressed genes indicated that diabetes strongly increased inflammatory pathways and that KU-596 therapy effectively reversed these increases independent of Hsp70. In contrast, the effects of KU-596 on decreasing the expression of genes regulating the production of reactive oxygen species were more Hsp70-dependent. These data indicate that modulation of molecular chaperones by novologue therapy offers an effective approach toward correcting nerve dysfunction in DPN but that normalization of inflammatory pathways alone by novologue therapy seems to be insufficient to reverse sensory deficits associated with insensate DPN

Absolute Stereochemical Determination of Asymmetric Sulfoxides via Central to Axial Induction of Chirality

The absolute configuration of chiral sulfoxides is determined by means of host–guest complexation that leads to the induction of axial chirality in an achiral host. The central to axial induction of helicity is rationalized by a simple recognition of the relative length and size of the substituents attached to the S-center. This technique is used to determine the absolute configuration of chiral sulfoxides, requiring micrograms of sample, without the need for prefunctionalization

Absolute Stereochemical Determination of Asymmetric Sulfoxides via Central to Axial Induction of Chirality

The absolute configuration of chiral sulfoxides is determined by means of host–guest complexation that leads to the induction of axial chirality in an achiral host. The central to axial induction of helicity is rationalized by a simple recognition of the relative length and size of the substituents attached to the S-center. This technique is used to determine the absolute configuration of chiral sulfoxides, requiring micrograms of sample, without the need for prefunctionalization

Point-to-Axial Chirality TransferA New Probe for “Sensing” the Absolute Configurations of Monoamines

A host molecule, capable of freely adopting <i>P</i> or <i>M</i> helicity, is described for molecular recognition and chirality sensing. The host, consisting of a biphenol core, binds chiral amines via hydrogen-bonding interactions. The diastereomeric complex will favor either <i>P</i> or <i>M</i> helicity as a result of minimizing steric interactions of the guest molecule with the binding cavity of the host, resulting in a detectable exciton-coupled circular dichroic spectrum. A working model is proposed that enables non-empirical prediction of the chirality of the bound amine

Development of Noviomimetics as C‑Terminal Hsp90 Inhibitors

KU-32 and KU-596 are novobiocin-derived, C-terminal heat shock protein 90 (Hsp90) modulators that induce Hsp70 levels and manifest neuroprotective activity. However, the synthetically complex noviose sugar requires 10 steps to prepare, which makes translational development difficult. In this study, we developed a series of “noviomimetic” analogues of KU-596, which contain noviose surrogates that can be easily prepared, while maintaining the ability to induce Hsp70 levels. Both sugar and sugar analogues were designed, synthesized, and evaluated in a luciferase reporter assay, which identified compound <b>37</b>, a benzyl containing noviomimetic, as the most potent inducer of Hsp70

Modulating Molecular Chaperones Improves Mitochondrial Bioenergetics and Decreases the Inflammatory Transcriptome in Diabetic Sensory Neurons

We have previously demonstrated that modulating molecular chaperones with KU-32, a novobiocin derivative, ameliorates physiologic and bioenergetic deficits of diabetic peripheral neuropathy (DPN). Replacing the coumarin core of KU-32 with a meta-fluorinated biphenyl ring system created KU-596, a novobiocin analogue (novologue) that showed neuroprotective activity in a cell-based assay. The current study sought to determine whether KU-596 offers similar therapeutic potential for treating DPN. Administration of 2–20 mg/kg of KU-596 improved diabetes induced hypoalgesia and sensory neuron bioenergetic deficits in a dose-dependent manner. However, the drug could not improve these neuropathic deficits in diabetic heat shock protein 70 knockout (Hsp70 KO) mice. To gain further insight into the mechanisms by which KU-596 improved DPN, we performed transcriptomic analysis of sensory neuron RNA obtained from diabetic wild-type and Hsp70 KO mice using RNA sequencing. Bioinformatic analysis of the differentially expressed genes indicated that diabetes strongly increased inflammatory pathways and that KU-596 therapy effectively reversed these increases independent of Hsp70. In contrast, the effects of KU-596 on decreasing the expression of genes regulating the production of reactive oxygen species were more Hsp70-dependent. These data indicate that modulation of molecular chaperones by novologue therapy offers an effective approach toward correcting nerve dysfunction in DPN but that normalization of inflammatory pathways alone by novologue therapy seems to be insufficient to reverse sensory deficits associated with insensate DPN

Diverging Novobiocin Anti-Cancer Activity from Neuroprotective Activity through Modification of the Amide Tail

Novobiocin is a natural product that binds the Hsp90 C-terminus and manifests Hsp90 inhibitory activity. Structural investigations on novobiocin led to the development of both anti-cancer and neuroprotective agents. The varied pharmacological activity manifested by these novobiocin analogs prompted the investigation of structure–function studies to identify these contradictory effects, which revealed that modifications to the amide side chain produce either anti-cancer or neuroprotective activity. Compounds that exhibit neuroprotective activity contain a short alkyl or cycloalkyl amide side chain. In contrast, anti-cancer agents contain five or more carbons, disrupt interactions between Hsp90α and Aha1, and induce the degradation of Hsp90-dependent client proteins