Q: Are PTOTAC soluble or insoluble?
A: Yes, PROTAC can be made soluble.
Q: You mentioned not knowing where to target in E3 ligases, why is targeting the substrate and E3 ligase interaction domain not plausible?
A: Yes, it is possible to develop E3 ligand by targeting the substrate-ligase interaction. In fact, that is how VHL ligand was developed. This is one consideration we discussed in our recent review article.
Q: Is there a reason why HECT E3 ligases are not considered in PROTAC design?
A: No specific reason as to why HECT ligases can’t be developed for PROTAC. One reason this has not happened yet may be that HECT are very large proteins and are not studied as well as CRL or other RING-type E3.
Q: Do you think the major challenge of lack of typical active pockets for CRLs being a large multi-unit protein family without typical active pockets to facilitate the drug design?
A: Yes, one major challenge to target/inhibit CRL is the lack of a well-defined active site, even though they act as an enzyme, like other enzymes such as kinases. But one could still target them by other means such as PPI inhibitor, albeit technically more challenging.
Q: How to understand the relationship between degradation activity, selectivity and possible off-target effects based on different targets, different cell lines, and different animal models?
A: I am afraid this is a $1M question and involves multiple aspects and I can’t responded with a simple answer. If you are asking for the selectivity, this is an advantage for the PROTAC since one could gain the view of selectivity of a given degrader at proteome scale by a simple proteomic analysis, which would be much more feasible than finding out how many kinases are affected by a given inhibitor.
Q: Do you think the Post translational modification of the E3 ligase complex is an important factor to be consider when designing PROTAC? Since PTM like Nedd will change the Cul flexibility and conformation. And how can we study this?
A: Neddylation is critically important for the activity of any CRL-based PROTAC. Other PTM such as phosphorylation, acetylation, methylation, so far do not seem to play critical role in the regulation of CRL as needylation.
Q: Did you test in any thermal assays of pain (got plate, tail flick etc.)?
A: No, we have not tested yet, but we do not have any reason to think that TRK degrader would not work for thermal pain.
Q: How to establish a scientific and credible PK/PD evaluation system as traditional methods cannot accurately assess the PK and PD properties of PROTACs?
A: There is no fundamental difference between PROTAC and traditional small molecule drugs when optimizing the PK, PD properties, although PROTAC molecules are larger, tend to be more unstable, have more hot-spots to be fixed and need more efforts to make into orally active.
Q: What are the advantages and disadvantages of N-degron pathway, over currently available E3 ligands.
A: Very little study on the feasibility of N-degron E3 for PROTAC at present. It’s difficult to comment, but I wonder whether high degree of substrate selectivity (specific amino acid sequences at particular location in the substrate protein) might be somewhat less compatible with the need of high degree of flexibility of PROTAC to target ‘any’ given proteins.
Q: This is a new area for me, so please excuse the basic question: what is the evidence that PROTACs (warhead-linker-E3 binder) small molecule is not subject to metabolism or degradation inside cells, and thus can be recycled?
A: PROTAC molecules ARE subjected to the intracellular metabolism and degradation. In fact, improving the metabolic stability of PROTAC is a major aspect of PROTAC medicinal pharmacology. The evidences supporting the recycling of degrader include the high potency, lasting effect and rapid kinetics.
Q: How to quickly and effectively screen for target protein ligands that can be used in PROTACs, especially those targeting protein−protein interactions
A: There are multiple ways to screen small molecule ligands/binders for proteins, but those binders are not very useful if not functional, until now. But there is no effective way right now to screen these initial binder-hits for their suitability as ligands for PROTAC. Such an assay, if developed, would be very useful for the TPD industry.
Q: What needs to happen before PROTACs, and molecular glues become a standard/accepted treatment?
A: Good phase 3 results, especially the ones that target a previously undruggable disease-causing protein.
Q: I believe PROTAC will decompose in few hours. So why we call it as catalytic?
A: We call the degrader catalytic because it connects an enzyme (E3 ligase) to a substrate and promotes a catalytic reaction (ubiquitylation). We don’t call the degrader catalyst, though. Yes, the degraders will be eventually decomposed, like the enzymes will be eventually degraded.
Q: For the TRK study, did you use a focused library to identify the degraders?
A: We discovered TRK degrader by linking and testing existing TRK ligands to a E3 ligand. This led to the discovery of potent degrader and we did not need to screen any library.
Q: What are your thoughts on the ADME optimization of PROTAC molecules for orally bioavailable drugs, since your company is optimizing their degraders?
A: Optimizing PROTAC into orally active compounds is not as challenging as a year or two ago. Although not routing yet, we have developed multiple orally active degraders targeting different proteins now.
Q: After targets ubiquitinated, they are always degraded by proteasome or they can be degraded by other mechanism, for example, autophagy?
A: Ubiquitin also tags proteins for lysosomal degradation, not just proteasome. This can be determined simply by using inhibitor specific to proteasome, lysosome or autophagy.
Q: Do you have any comments on the target selection for PROTAC drug discovery, for example, to differentiate from well-established inhibitors?
A: We think the future of PROTAC is to target previously undruggable non-enzyme proteins, not to compete with inhibitors.
Q: What do you think of slow vs. fast degrader? Would you go after slow degraders? (E.g. a degrader that start degrading the substrate after 20h)
A: We don’t see obvious reason for the need of a slow or delayed degrader. No one has attempted yet to developed a delayed degrader, but this need can certainly change for different target or indications.
Q: If you can’t see in vitro differentiation of a PROTAC from a SM inhibitor at a given target, do you think you can see differentiation from in vivo studies?
A: Yes, there are multiple situations an in vivo differentiation can occur. These include the ability of degrader to overcome the drug resistance, to achieve more potent efficacy and perhaps most importantly selective degradation based on tissue-selective E3 expression.
Q: Where can I learn more about BioDuro-Sundia's PROTACs drug discovery activities?
A: Clients considering a service contract can learn more about our PROTAC Drug Discovery Platform. Our recent webinar also details BioDuro-Sundia's PROTACs-related service capabilities.
