A Unique Small Molecule Inhibitor of Enolase Clarifies Its Role in Fundamental Biological Processes

Jung, Da Woon; Kim, Woong‐Hee; Park, Si-Hwan; Lee, Jinho; Kim, Jinmi; Su, Dongdong; Ha, Hyung‐Ho; Chang, Young‐Tae; Williams, Darren R.

doi:10.1021/cb300687k

Cited by 81 publications

(71 citation statements)

References 51 publications

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“…When surface ENO1 expression was downregulated by transfection with ENO1-specific siRNA (online supplementary figure 1), production of proinflammatory cytokines by RA PBMCs in response to apoB stimulation decreased (figure 2B). Furthermore, apoB-induced IL-1β and IL-6 production was suppressed in the presence of ENOblock, a non-substrate analogue that can directly inhibit ENO1 activity12 (figure 2C). Therefore, treatment of PBMCs with apoB induces proinflammatory response, and this response depends on expression of functional ENO1.…”

Section: Resultsmentioning

confidence: 99%

Apolipoprotein B binds to enolase-1 and aggravates inflammation in rheumatoid arthritis

et al. 2018

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ObjectiveImmune cells from patients with rheumatoid arthritis (RA) express more enolase-1 (ENO1) on their surface than those from healthy subjects, and they elicit an enhanced inflammatory response. This study is aimed to identify the ligands of ENO1 that could promote inflammatory loops in vitro and enhance the arthritis severity in vivo.MethodsENO1-binding proteins in RA synovial fluid were identified by mass spectromety, and affinity to ENO1 was evaluated by means of a ligand blotting and binding assay, surface plasmon resonance and confocal microscopy. Proinflammatory response by the interaction between ENO1 and apolipoprotein B (apoB) was tested in vitro and in vivo using peripheral blood mononuclear cells and a K/BxN serum transfer arthritis model and low-density lipoproteins receptor (LDLR) knockout mice.ResultsApoB in the synovid fluid of patients with RA was identified as a specific ligand to ENO1 with a higher affinity than plasminogen, a known ENO1 ligand. ApoB binding to ENO1 on monocytes elicited the production of tumour necrosis factor-α, interleukins (IL)-1β and IL-6 through both p38 mitogen-activated protein kinase and NF-κB pathways. In the K/BxN serum transfer arthritis model, administration of apoB increased the production of proinflammatory cytokines and exaggerated arthritis severity. The severity of K/BxN serum transfer arthritis in LDLR knockout mice was comparable with wild-type mice.ConclusionsA key component of atherogenic lipids, apoB, aggravated arthritis by potentiating the inflammatory response via its interaction with ENO1 expressed on the surface of immune cells. This suggests a novel mechanism by which lipid metabolism regulates chronic inflammation in RA.

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Section: Resultsmentioning

confidence: 99%

Apolipoprotein B binds to enolase-1 and aggravates inflammation in rheumatoid arthritis

et al. 2018

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“…Thus, the expression levels of NSE and key inflammatory mediators were examined in vehicle treated SCI in rats and compared those with SCI animals treated with a novel small molecule inhibitor of enolase ENOblock (C31H43FN8O3). ENOblock is a novel small molecule which is the first, non-substrate analogue that directly binds to enolase and inhibits its expression and activity (47). It is the first reported enolase inhibitor that is suitable for biological assays, and ENOblock treatment has been shown to target cell-surface-bound enolase to modulate cell growth through downregulation of AKT and BCL-xL.…”

Section: Introductionmentioning

confidence: 99%

Targeting Enolase in Reducing Secondary Damage in Acute Spinal Cord Injury in Rats

et al. 2017

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Spinal cord injury (SCI) is a complex debilitating condition leading to permanent life-long neurological deficits. The complexity of SCI suggests that a concerted multi-targeted therapeutic approach is warranted to optimally improve function. Damage to spinal cord is complicated by an increased detrimental response from secondary injury factors mediated by activated glial cells and infiltrating macrophages. While elevation of enolase especially Neuron Specific Enolase (NSE) in glial and neuronal cells is believed to trigger inflammatory cascades in acute SCI, alteration of NSE and its subsequent effects in acute SCI remains unknown. This study measured NSE expression levels and key inflammatory mediators after acute SCI and investigated the role of ENOblock, a novel small molecule inhibitor of enolase, in a male Sprague-Dawley (SD) rat SCI model. Serum NSE levels as well as cytokines/chemokines and metabolic factors were evaluated in injured animals following treatment with vehicle alone or ENOblock using Discovery assay. Spinal cord samples were also analyzed for NSE and MMPs 2 and 9 as well as glial markers by Western blotting. The results indicated a significant decrease in serum inflammatory cytokines/chemokines and NSE, alterations of metabolic factors and expression of MMPs in spinal cord tissues after treatment with ENOblock (100μg/kg, twice). These results support the hypothesis that activation of glial cells and inflammation status can be modulated by regulation of NSE expression and activity. Analysis of SCI tissue samples by immunohistochemistry confirmed that ENOblock decreased gliosis which may have occurred through reduction of elevated NSE in rats. Overall, elevation of NSE is deleterious as it promotes extracellular degradation and production of inflammatory cytokines/chemokines and metabolic factors which activates glia and damages neurons. Thus, reduction of NSE by ENOblock may have potential therapeutic implications in acute SCI.

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“…A recent report by Jung et al . described an altogether different glycolysis inhibitor: a heterocyclic, cell permeable inhibitor of Enolase that apparently binds outside the active site [7]. Jung et al .…”

Section: Introductionmentioning

confidence: 99%

“…Jung et al . identified (N-2-2-2-aminoethoxy)ethoxy]ethyl]4-4-cyclohexylmethyl)amino]6-4-fluorophenyl)methyl]amino]1,3,5-triazin-2-yl]amino]benzeneacetamide hydrochloride) also known as AP-III-a4 as a compound selectively toxic to cancer cells under hypoxic conditions [7]. Further experiments led to the conclusion that AP-III-a4 exerts its effect by direct inhibition of the glycolytic enzyme Enolase with a reported IC 50 of ~0.6 μM, hence it was dubbed ‘ENOblock’.…”

Section: Introductionmentioning

confidence: 99%

ENOblock Does Not Inhibit the Activity of the Glycolytic Enzyme Enolase

Satani¹,

Lin²,

Hammoudi³

et al. 2016

PLoS ONE

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Inhibition of glycolysis is of great potential for the treatment of cancer. However, inhibitors of glycolytic enzymes with favorable pharmacological profiles have not been forthcoming. Due to the nature of their active sites, most high-affinity transition-state analogue inhibitors of glycolysis enzymes are highly polar with poor cell permeability. A recent publication reported a novel, non-active site inhibitor of the glycolytic enzyme Enolase, termed ENOblock (N-[2-[2-2-aminoethoxy)ethoxy]ethyl]4-4-cyclohexylmethyl)amino]6-4-fluorophenyl)methyl]amino]1,3,5-triazin-2-yl]amino]benzeneacetamide). This would present a major advance, as this is heterocyclic and fully cell permeable molecule. Here, we present evidence that ENOblock does not inhibit Enolase enzymatic activity in vitro as measured by three different assays, including a novel 31P NMR based method which avoids complications associated with optical interferences in the UV range. Indeed, we note that due to strong UV absorbance, ENOblock interferes with the direct spectrophotometric detection of the product of Enolase, phosphoenolpyruvate. Unlike established Enolase inhibitors, ENOblock does not show selective toxicity to ENO1-deleted glioma cells in culture. While our data do not dispute the biological effects previously attributed to ENOblock, they indicate that such effects must be caused by mechanisms other than direct inhibition of Enolase enzymatic activity.

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A Unique Small Molecule Inhibitor of Enolase Clarifies Its Role in Fundamental Biological Processes

Cited by 81 publications

References 51 publications

Apolipoprotein B binds to enolase-1 and aggravates inflammation in rheumatoid arthritis

Apolipoprotein B binds to enolase-1 and aggravates inflammation in rheumatoid arthritis

Targeting Enolase in Reducing Secondary Damage in Acute Spinal Cord Injury in Rats

ENOblock Does Not Inhibit the Activity of the Glycolytic Enzyme Enolase

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