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Thank you for the kind of introduction.
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It's my pleasure to be here today.
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So I will talk about our degrader discovery platform and how we use it to support integrated drug discovery for PROTACs and molecular glue.
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And before I start today's topic, I will just have few words about our company.
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So we are Nuvisan, fully integrated science CRO supporting the drug discovery project at different stages from target identification, validation, head to lead and lead optimisation and all the way down to clinic development.
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We have five sites across Europe, four in Germany, one in France.
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The biggest site is Berlin and was taken over from Bayer in 2020 with a fully functional drug discovery unit and supporting lead discovery medicine, chemistry, pharmacology, DMPK and toxicology.
1:01
So we would be happy to support any drug discovery project coming to us.
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So now I will switch to today's topic by a short introduction about targeted protein degradation with PROTAC and molecular glue.
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So as we know recently targeted protein degradation has become a new approach for drug discovery as it can selectively degrade the disease protein and affect both enzymatic and non-enzymatic function of the protein.
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So PROTAC are hetero bi function molecules with three components, the E3 ligase binder, Linker and POI.
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PROTAC can form ternary complex with both E3, and POI induce the POI ubiquitination ultimate degradation.
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Molecular glue is a class of small molecular which also can induce the POI degradation in a similar mechanism compared to PROTAC, but it has a much smaller molecular weight so has a high chance to obtain drug like properties.
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However, the original design of molecular glue is still quite challenging and new sign.
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We have strong capabilities in supporting degrader discovery.
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We have fully integrated degrader platform on one campus site, offering efficient senses, Medcam optimization, state-of-the-art technologies, and assay for the degrader profiling.
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With our established workflow, we can support starting from the binder finding, degrader design and synthesis, DMPK optimization, and all the way down to candidate development.
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So in our degrader discovery platform, we have four important parts.
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The life science chemistry part is supporting the degrader design, faster synthesis, PhysChem profiling.
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We are using biochemical biophysical assay to study the interaction between the protein and degrader using cellular assay.
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Try to understand all the steps which lead to the protein degradation inside of cells such as Ternary, Complex Formation, Ubiquitination, target degradation.
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We're also looking to the selectivity and downstream effect of the degrader.
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And with our DMPK and in vivo animal pharmacology model support, we can further optimise the degrader into drug candidate.
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And in the following slides, I'm going to show you more details for each part.
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Let's start with the chemistry platform.
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So there we have a chemistry toolbox.
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Inside there is a ready to use library containing several E3 ligase binder and around 7K linkers including the commonly used PEC linker, RAGE linker, basic container linker.
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These components can already deliver around 350,000 possible combinations and this library is continuously growing.
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We have smart degrader design to support data mining, compounds enumeration and property calculation.
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We have also established different synthetic methodologies for fast degrader synthesis.
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A unique feature of our toolbox is so-called direct to biology approach where we have used a new approach for fast active protect generation, which I will show you within your case study later in my talk.
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And with our two-chemistry toolbox, we are able to deliver more than 100 degraders per day and when combined with direct to biological approach, we are able to generate the preliminary SAR for this 100 degraders within your time of one week.
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We have selected specific PhysChem assays for the to address the challenges associated with degrader especially for protect such as we use ChromelogD kinetic aqueous solubility measurement and also look into the solubility in bio relevant solutions.
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We look into the chemical stability, measure the hydrolytic stability in different conditions and also do the stability screening.
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We have established assay to determine the EPSA as it has much better correlation with the permeability.
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We also have assays such as XRPD, DSC to study the solid crystal anonymity.
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We also have E3 ligase database containing more than 750 human E3 ligase with all this relevant biologic information inside and this one can be used to support the identification of new E3 ligase.
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And we have several biochemical biophysical assays to study the protein and POR and degrader induction.
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Try to understand the binary ternary complex formation.
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SPR is one of them quite important one.
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We use biocles to measure the SPR for different applications.
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First one is to function as a high throughput screening assay to identify new binder.
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Second, to study the binary ternary complex formation and measure the binary affinity kinetic parameters to fully characterise the binding events.
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So, we regard the SPR as an important assay, provide the rational for preferred ternary complex formation and drive the SAR optimisation.
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We have established ASMS method and has used that first as a high throughput screening method to identify new binder, second to study the ternary complex formation.
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We also have quite strong capabilities for protein production of E3 ligase and POI generate high quality of crystal structure of binary and ternary complex and these are providing greater important information for the degrader optimization.
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To understand the biologic effect of the degrader, we have several biochemical and cellular assays.
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For example, we use TR-FRET assay and NanoBRET assay to study the target engagement and E3 engagement both biochemically and cellular to look into the ternary complex formation.
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We have non operated assay and biochemical HTRFSE and we offer non operated assay tube assay protect plate bases to check on the POI ubiquitination and to study the protein degradation inside cell.
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We also have several cellular assays such as immune fluorescent degradation assay, fluorescent intensity degradation assay, and hybrid assay.
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Some of the assay has also been adopted into as a primary screen assay using your high throughput fashion.
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Of course, we also have other assay available such as western blot, capillary electrophoresis and so on.
8:30
Besides looking to the POI ubiquitination and degradation, we also have a Protonmix platform help us to understand the selectivity of the degrader and identify the nail substrate.
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So and also a good understanding of the in vitro, in vivo DMPK would be important for the degrade optimization.
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We have a set of assays to study the compounds and make the ranking and also selection of the candidate that can be used for the PKPD study and human PK prediction.
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And this assay are quite flexible, can be used to meet a specific project needs.
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We also have a broader range of animal pharmacology model, which is more than 200 type of animal model in mice and rats, especially in the field of oncology and immune oncology.
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So we can study tolerability, exposure, efficacy.
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I think that this one is now efficacy PKPD and also establish new animal model for specific disease.
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So this animal models are would allow in depth characterisation of the degrader molecular and answer critical questions for further optimisation.
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So that's a general information regarding our platform and now I will switch to several case studies and show you how we use this platform to support integrated and degrader discovery.
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The first one was a collaboration with Bayer.
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We were together to develop molecular glue for CDK12.
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So starting with phenotypic screening, identify inhibitory compounds and later with biochemical and cellular profiling, we found such compound was functioning as a molecular glue induce the degrading degradation of CCNK and CDK12.
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With further optimization, we have generated candidate compound with quite good antitumor efficiency in animal study.
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The second one was also collaboration with Bayer to develop PROTAC for MetAP2, starting with a binder for MetAP2 extended with linker and its * binder and with first optimisation generated molecular called BAY-277.
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It turns out to be a potent and selective MetAP2 degrader was quite good in vivo and tolerability and efficiency.
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The third one was collaboration with Bayer and University Upsala, and we were using AMR to study the molecular cheminonicity and we found under different biochemical environment for the same protect molecular, it can adopt to different confirmation and contribute to the permeability.
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And the last example is what I just mentioned before the direct to biology approach which is quite new.
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And there we have combined the pool synthesis together with AS-MS Ternary complex assay for active protect generation.
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So the workflow looks like this.
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We use fast solid support synthesis with a split and pool strategy to generate protect pool as a mixture.
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This pool has been submitted for AS-MS Ternary complex assay.
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In this assay that if a protag from this pool forms ternary complex after washing, the enrichment of the compound would be higher than the compounds only forms binary complex or does not bind at all.
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So based on this assay, we were apt to identify protags which can form ternary complex with POI and E3 ligase resources of this protects and testing in relevant biochemical and cellular assay to check on the protein degradation would allow us to obtain active PROTAC.
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Based on this idea, we have done a small case study for fast generation of new active BRD4-VHL PROTACs starting with computer modelling and selection of five linker using solar phase synthesis to generate protect pool containing five PROTACs.
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Here we have choosing to protect one which was a report literature reported BRD4 PROTAC called Mz-1.
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We use this one as opposed to control both for the chemical synthesis and biochemical assay.
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So the whole synthesis was straightforward on solar support.
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7 steps with 21% overall yield.
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There was no need of purification in between and afterwards and the purity of this protect pool was more than 70%.
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Then we put this pool into AS-MS Ternary complex assay and interestingly we found all five PROTACs from this pool formed somehow Ternary complex.
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For example, here in present of BRD4 and VHL, the enrichment of PROTAC 1 is much higher than the case if only BRD4 was present.
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The difference between these two indicate a Ternary complex has been formed.
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Encouraged by these results, we have done resources of this 5 PROTAC and put into the cellular study.
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When the K562 cell line which is a CLL cell line treated with this 5 PROTACs for 48 hours, we found PROTAC 1,2,4,5 they were inducing the BRD4 degradation.
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You know dose dependent manner and surprisingly PROTAC 3 didn't do anything for BRD4, although it was forming the ternary complex in the biochemical assay.
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And then in the proliferation assay, we noticed that all 5 PROTACs were having anti proliferation effect in which PROTACs 1,2,5 had similar * and PROTACs 3,4 were less active and had similar * compared to the negative control system Z1.
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So based on these results, we conclude that with our direct to biological approach, we were able to faster generate new active BRD4 PROTAC as a similar or comparable degradation efficiency as Mz-1.
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And that will bring almost to the end of my talk.
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I hope I could convince you about our capabilities in supporting the integrated degrader discovery and our platform is quite flexible, can be extended within other emerging degrader technologies and also go beyond degradation.
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And we believe the chemically induced proximity field including the target protein degradation is quite fast and growing with a huge promise.
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And we are excited to part of it and looking forward for further development.
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With that, I would like to thank you for your attention and happy to answer any questions.