Enabling Protein Degradation Drug Discovery

Ubiquigent – your expert partner in ubiquitylation and protein degradation drug discovery

At Ubiquigent, we have applied our ubiquitin system expertise to develop a comprehensive suite of assays to support your drug discovery programmes.  So, regardless of whether you’re interested in identifying or characterising deubiquitylase (DUB) enzyme inhibitors or modulators of ubiquitin ligases, or wish to exploit the ubiquitin proteasome system (UPS) to trigger the degradation of proteins – including across previously ”undruggable” targets – via PROTAC or molecular glue type approaches, we have the knowledge, assays and capabilities to support and advance every step of your drug discovery programmes.

The central role of ubiquitylation in diverse cellular processes makes the enzymes responsible for ubiquitylation and de-ubiquitylation very attractive drug targets to address a range of human pathologies.  In the decade or more since the Nobel prize was awarded for the discovery of ‘ubiquitin-mediated protein degradation’, and more recently the clinical approval of proteasome inhibitors, there has been growing interest in exploiting other com­ponents of the ubiquitylation machinery as therapeutic targets, such as DUBs.

The development of selective DUB inhibitors has been limited by insufficient understanding of DUB biology, difficulties in establishing robust biochemical assays suit­able for compound screening, limitations in cellular and in vivo models to assess DUB activity or inhibition, and the pleiotropic nature of various small-molecule DUB inhibitors (Harrigan et al., 2017).  Over the last 10 years, Ubiquigent scientists have focused on building a suite of biochemical and cellular assays to identify and characterise screening hits, eliminate false positives, confirm cellular target engagement, and explore the mechanism of action of the molecules with the objective of supporting client drug discovery programmes.

Through our network of academic partners, collective experience in drug discovery and deep and extensive experience in ubiquitin system assays and medicinal chemistry, our scientists can complement your disease-specific expertise to assist you in identifying genuine drug candidates.

Scroll down to explore how our Drug Discovery Screening Platform can be applied to:

  • Target identification and validation
  • Identification and characterisation of DUB inhibitors
  • Target engagement and cellular selectivity profiling
  • Mechanism of action studies
  • PROTACs and molecular glues
  • E3 ligases

Target identification and validation

Our PROTEOMEprofiler and DUBprofiler-Cell™ multiplexed tandem mass spectrometry enabled platforms are powerful complementary methods to support target identification and validation objectives.

Changes in protein expression and homeostasis in response to disease states or other experimental stimuli (eg cytokine addition or stress conditions) can be revealed through the PROTEOMEprofiler service.  Since the entire proteome of the cell or tissue is examined, this is an entirely unbiased method for identifying up or down regulated targets of interest including – but not limited to – the ubiquitin pathway.  The workflow is highly flexible and provides for your cell line of choice to be examined, furthermore the multiplexed procedure enables the direct comparison of multiple conditions.  Follow-up experiments utilising isogenic lines, in which candidate DUBs (or ligases) are genetically deleted, could offer further confirmation of the relevance of these targets (and associated pathway modulation) to disease, providing robust target validation.

The DUBprofiler-Cell™ platform focuses on active DUBs within a cell or tissue. Using this assay, we can investigate the activity of individual candidate DUBs or the entire active “DUBome” in therapeutically relevant cells and tissues or compare the active DUBome between isogenic cell line pairs carrying genetically defined mutations or treated with various stimuli.  As with the PROTEOMEprofiler platform the DUBprofiler-Cell™ workflow is highly flexible and enables your cell line of choice to be examined.

Identification and characterisation of DUB inhibitors

Ubiquigent’s DUBprofiler service is well established as the leading platform for the identification and characterisation of novel DUB inhibitors and has been employed by leading groups in support of their drug discovery programmes.  Use the DUBprofiler in vitro platform to rapidly reveal chemical starting points from your library or utilise the service to support your Q-SAR medicinal chemistry cycles of compound optimisation to establish potency and selectivity across our extensive panel of DUBs.

One of the unique selling points of our services is our flexibility; the majority of our clients wish to use our standard, validated conditions for DUBprofiler, but we are happy to work with you if you require the assay to be tailored to your requirements, e.g. adjusting pre-incubation parameters, or screening any number of DUBs from our extensive panel at any concentration(s) of compound.  We are also able to quickly determine IC50 values for any compounds of interest arising from a primary screen using the same assay format.

We are happy to follow up with further analysis of your compounds by any of the following approaches:

  • Confirmation of hits in an orthogonal assay (e.g. DUBprofiler-MALDI™)
  • Determination of reversibility, competitive/non-competitive nature of compounds (including but not limited to use of the DUBadduct™ assay)
  • Elimination of false positive redox cycling compounds (REDOXprofiler)

Target engagement and cellular selectivity profiling

The DUBprofiler-Cell™ assay allows you to quickly confirm test compound-DUB target engagement in a relevant cellular context and delivers relative cellular EC50 values versus every DUB engaged by the compound.  As such, it is hugely complementary to DUBprofiler, with the added feature of it being highly relevant to the disease context since you can interrogate your cell line of choice (eg one in which you see a relevant phenotypic/functional/therapeutic/PD effect of the compound).

Mechanism of action studies

Treatment of disease-relevant cells with candidate compounds within our PROTEOMEprofiler service provides invaluable information about the mechanism of action of candidate compounds. Analysis is entirely unbiased and proteome-wide.  Time-course and dose response studies can be efficiently incorporated into a study, alongside inactive analogues or tool compounds.  Use PROTEOMEprofiler to confirm the primary target in cells, reveal full pathway and proteomic alterations and identify potential biomarkers to assist and accelerate clinical development.

PROTACs and molecular glues

The use of proteolysis-targeting chimeras (PROTACs) or molecular glues are a new modality that “hijacks” the ubiquitin-proteasome system to trigger the degradation of specific target proteins implicated in disease, that might otherwise be chemically intractable.  One of the key features of PROTACs is their selectivity; however it has been shown by the PROTAC MZ1, for example, that predictions about selectivity are not always borne-out in cells.  Thus, one essential element of any targeted protein degradation drug discovery programme – using the PROTAC, molecular glue or similar modalities – is to be able to demonstrate selectivity in a disease-relevant, cellular context, by understanding the down – and potentially up – regulation of individual members of the cellular proteome.

PROTEOMEprofiler is a new service that reports changes in relative proteome abundance in response to any cellular input and provides relative quantification across the range of conditions tested.  The platform could be employed to establish the selectivity of individual PROTACs/molecular glues (for example as part of MOA studies or due diligence on in-licensing opportunities), or to prioritise different PROTAC/molecular glue designs for future development.

Stay posted for our PROTACprofiler in vitro Q-SAR assay platform, coming soon.

E3 ligases

Ubiquigent has been approached by several clients to use our knowledge of the ubiquitin pathway and assay development expertise to develop novel ubiquitin system target assays in support of their drug discovery programmes.  As always, we are flexible with our client’s requirements; validated assays can be screened against client compound libraries at Ubiquigent or transferred to the client laboratories.  As one example, IDOL has been identified as a target of significant interest in Alzheimer’s disease and in the regulation and turnover of the LDL receptor (LDLR) in lipid homeostasis.  We have developed an HTS-compatible IDOL assay for screening compound libraries.

For further details on any aspect of our Drug Discovery Screening Platform, follow the links on the left or Contact us directly to discuss your requirements.