The promiscuous protein binding ability of erythrosine B studied by metachromasy (metachromasia)

2015

CDT

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The tumor necrosis factor (TNF) superfamily (TNFSF) contains about thirty structurally related receptors (TNFSFRs) and about twenty protein ligands that bind to one or more of these receptors. Almost all of these cell surface protein-protein interactions (PPIs) represent high-value therapeutic targets for inflammatory or immune modulation in autoimmune diseases, transplant recipients, or cancers, and there are several biologics including antibodies and fusion proteins targeting them that are in various phases of clinical development. Small-molecule inhibitors or activators could represent possible alternatives if the difficulties related to the targeting of protein-protein interactions by small molecules can be addressed. Compounds proving the feasibility of such approaches have been identified through different drug discovery approaches for a number of these TNFSFR-TNFSF type PPIs including CD40-CD40L, BAFFR-BAFF, TRAIL-DR5, and OX40-OX40L. Corresponding structural, signaling, and medicinal chemistry aspects are briefly reviewed here. While none of these small-molecule modulators identified so far seems promising enough to be pursued for clinical development, they provide proof-of-principle evidence that these interactions are susceptible to small-molecule modulation and can serve as starting points toward the identification of more potent and selective candidates.

Section: Ox40–ox40lmentioning

confidence: 65%

“…Planar or partially planar aromatic and hydrophobic residues are indeed over-represented at PPI interfaces [26, 27]. Recently, even promiscuous small-molecule PPI inhibitors have been identified [28, 29]. The ability to target PPIs could considerably enlarge the number of feasible therapeutic targets.…”

Section: Introductionmentioning

confidence: 99%

TNF Superfamily Protein-Protein Interactions: Feasibility of Small- Molecule Modulation

Song¹,

2015

CDT

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“…This sensor cell line was generated by stable transfection of HEK293 cells with the human CD40 gene and an NF-κB-inducible SEAP construct, which consists of the SEAP reporter gene under the control of the IFN-β minimal promoter fused to five NF-κB (and five AP-1) binding sites. As before [23], cells were cultured and assayed for activation as recommended by the manufacturer. Briefly, the cells were cultivated in Dulbecco’s Modified Eagle (DMEM) media supplemented with 4.5 g/L glucose, 10% v/v FBS, 50 U/mL penicillin, 50 μg/mL streptomycin, 100 μg/mL Normocin, and 2 mM l -glutamine.…”

Section: Methodsmentioning

confidence: 99%

“…We are using these readily available cells as a standard cell-based model to assess ligand-induced receptor activation for selected members of the tumor necrosis superfamily (TNFSF) in our work focusing on small-molecule costimulatory modulation [22, 23]. Here, we evaluated the suitability of this model to quantitate glucocorticoid activity both in a detailed full dose-response format as well as in a single-dose format.…”

Section: Introductionmentioning

confidence: 99%

Effects of representative glucocorticoids on TNFα- and CD40L-induced NF-κB activation in sensor cells

Cechin

2014

Steroids

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Glucocorticoids are an important class of anti-inflammatory/immunosuppressive drugs. A crucial part of their anti-inflammatory action results from their ability to repress proinflammatory transcription factors such as nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) upon binding to the glucocorticoid receptor (GR). Accordingly, sensor cells quantifying their effect on inflammatory signal-induced NF-κB activation can provide useful information regarding their potencies as well as their transrepression abilities. Here, we report results obtained on their effect in suppressing both the TNFα- and the CD40L-induced activation of NF-κB in sensor cells that contain an NF-κB–inducible SEAP construct. In these cells, we confirmed concentration-dependent NF-κB activation for both TNFα and CD40L at low nanomolar concentrations (EC50). Glucocorticoids tested included hydrocortisone, prednisolone, dexamethasone, loteprednol etabonate, triamcinolone acetonide, beclomethasone dipropionate, and clobetasol propionate. They all caused significant, but only partial inhibition of these activations in concentration-dependent manners that could be well described by sigmoid response-functions. Despite the limitations of only partial maximum inhibitions, this cell-based assay could be used to quantitate the suppressing ability of glucocorticoids (transrepression potency) on the expression of proinflammatory transcription factors caused by two different cytokines in parallel both in a detailed, full dose-response format as well as in a simpler single-dose format. Whereas inhibitory potencies obtained in the TNF assay correlated well with consensus glucocorticoid potencies (receptor-binding affinities, Kd, RBA, at the GR) for all compounds, the non-halogenated steroids (hydrocortisone, prednisolone, and loteprednol etabonate) were about an order of magnitude more potent than expected in the CD40 assay in this system.

“…A number of studies suggested various scaffolds, many of them based on flat aromatic structures, as appropriate for design of effective SMPPIIs [108–110]. Notably, along these lines, we have identified certain xanthene-based organic dyes, such as erythrosine B and rose bengal, as the first promiscuous SMPPIIs [111, 112]. While their inhibitory mechanism is not entirely clear, it might be related to their rigid flat structures, since planar or partially planar aromatic and hydrophobic residues are over-represented at PPI interfaces [113], and they may make the binding of such structures here particularly likely.…”

Section: Small-molecule Ppi Inhibitorsmentioning

confidence: 96%

Toward Small-Molecule Inhibition of Protein–Protein Interactions: General Aspects and Recent Progress in Targeting Costimulatory and Coinhibitory (Immune Checkpoint) Interactions

Bojadzic

2018

CTMC

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Protein-Protein Interactions (PPIs) that are part of the costimulatory and coinhibitory (immune checkpoint) signaling are critical for adequate T cell response and are important therapeutic targets for immunomodulation. Biologics targeting them have already achieved considerable clinical success in the treatment of autoimmune diseases or transplant recipients (e.g., abatacept, belatacept, and belimumab) as well as cancer (e.g., ipilimumab, nivolumab, pembrolizumab, atezolizumab, durvalumab, and avelumab). In view of such progress, there have been only relatively limited efforts toward developing small-molecule PPI inhibitors (SMPPIIs) targeting these cosignaling interactions, possibly because they, as all other PPIs, are difficult to target by small molecules and were not considered druggable. Nevertheless, substantial progress has been achieved during the last decade. SMPPIIs proving the feasibility of such approaches have been identified through various strategies for a number of cosignaling interactions including CD40-CD40L, OX40-OX40L, BAFFR-BAFF, CD80-CD28, and PD-1-PD-L1s. Here, after an overview of the general aspects and challenges of SMPPII-focused drug discovery, we review them briefly together with relevant structural, immune-signaling, physicochemical, and medicinal chemistry aspects. While so far only a few of these SMPPIIs have shown activity in animal models (DRI-C21045 for CD40-D40L, KR33426 for BAFFR-BAFF) or reached clinical development (RhuDex for CD80-CD28, CA-170 for PD-1-PD-L1), there is proof-of-principle evidence for the feasibility of such approaches in immunomodulation. They can result in products that are easier to develop/ manufacture and are less likely to be immunogenic or encounter postmarket safety events than corresponding biologics, and, contrary to them, can even become orally bioavailable.