Equillium, Inc. ($EQ)

And welcome to the Equillium Virtual Investor Event. [Operator Instructions] As a reminder, this call is being recorded, and a replay will be made available on the Equillium website following the conclusion of the event. I'd now like to turn the call over to your host, Stephen Connelly, Chief Scientific Officer at Equillium. Please go ahead.

Thanks, Tara, and good morning, everyone, and thanks for joining today's webinar titled Insights to the Aryl Hydrocarbon Receptor and miR-124 Axis. Now before we start, it probably makes sense to give you a little bit of context on this work from the outset, both the Aryl Hydrocarbon Receptor and miR-124 are two distinct biological molecules. And indeed, therapeutic strategies modulating both of them have been shown to be effective in treating ulcerative colitis. However, data exists from cells, animals and humans that point to a more direct and functional relationship between these two molecules. And despite the biological and therapeutic relevance, the Parador relationship between these two has remained elusive and controversial. So understanding this relationship has critically important implications for both mechanistic interpretation and rational development of therapies targeting the AhR and miR-124 access in inflammatory disease. So presented here today, we'll do a cross-section of work aimed at elucidating the functional relationship between AhR and miR-124, specifically providing a unifying mechanistic framework linking ligand-dependent AhR signaling to miR-124 expression. Now to ensure we have enough time for Q&A, we will go over all of the data in the deck. So please be sure to download the full deck after today's call. So let me just dive straight into it. Just a quick recap on the Aryl Hydrocarbon Receptor as a signaling hub. Now the Aryl Hydrocarbon Receptor is a Zenosensor, it's a ligand-activated transcription factor that resides predominantly in our barrier tissues. And when ligands come into the cytoplasm and buying to AhR, that causes translocation and dimerization within the nucleus. And then the recruitment of cofactors or binding to DNA leads to the induction of multiple pathways responsible for detoxification, immune modulation, anti-inflammatory cytokines, antioxidant proteins and barrier function and repair proteins. Now let's walk you through a few of the top ones in each of the different functional areas. Now historically, the Aryl Hydrocarbon Receptors has been largely known for its ability to initiate xenobiotic responses, so metabolism genes, things that will destroy molecules that have come into the tissues given their signals and then need to be cleared. So things like the SIP 1A, SIP 1A2, SIP 1B1, a number of other genes, including enzymes that glucuronidate molecules, all with the sole purpose of eliciting a response to remove those molecules from the cell and turn off the signal. Now more recently, in the last decade, areas relating to immune regulation and barrier function have been more widely studied. And this has changed the paradigm for how we think about the Aryl Hydrocarbon Receptor. Now here, there are a number of different genes modulated, but also cofactors recruited in a cellular dependent manner. So cofactors recruited in the immune cell typically modulate immune responses like NF-kappaB, ROR gamma t, and then cofactors recruited into the epithelial barriers, like wind, HIF, AP1, et cetera, modulate things like barrier function, repair and regeneration. Now depending on the molecule and depending on the signal and the cell, you can get what we call biased agonism. So not all AhR modulators are created equally. These different gene sets will show a different signature, largely dependent on the ligand itself. Now importantly, XenoBiotic or external unnatural molecules like pollutants or drugs typically elicit very strong xenobiotic responses where dietary ligands, those illicit responses that are more related to the immune regulation barrier function and repair things that are more important from a therapeutic perspective. Now an area of emerging research has been the microRNAs that are induced or modulated by AhR. Now I'm just showing the ones that we know from the literature are induced from AhR activation. But there are a number of mirrors that have been implicated that actually modulate the levels of AhR. And one of those in the literature is miR-124. So let's talk about the AhR miR-124 access. So what's known between this relationship? Now there's a nice set of papers by Zoo. And what they report is that there's an inverse correlation between miR-124 and AhR protein levels in tissue from active celiac disease patients. Over expression in animal and cell models have shown overexpression of miR-124 decreased AhR expression and had a net pro-inflammatory effect, while inhibition of miR-124 increased AhR expression and had an anti-inflammatory effect. Now this suggests there's some sort of negative or regulatory relationship between those where overexpression of miR-124 can then downregulate AhR as potentially a signal termination mechanism. And vice versa when levels of miR-124 are reduced, AhR levels return. And this is meant to be perhaps a way of balancing the anti and pro-inflammatory responses within cells. Now there is another set of data out there. I'd like to turn this to sort of obefasemod paradox. And that's data that would suggest the induction of miR-124 does not illicit increased cytokine. It's actually anti-inflammatory. But the key challenge with this data set would be that we don't really understand from this data, the levels of activation or expression of AhR as a way to reconcile this with the data presented by However, this model can be reconciled, if, in fact, obefasemod is itself an AhR modulator. Now to tackle that, there were three key questions we have to ask. Is ABX464 an AhR modulator? Does AhR modulation itself in miR-124? And what are the functional consequences of AhR blockade on the induction of these downstream pathways? So to tackle this and largely spurred on by questions from the investor and academic community, we look to in silico modeling as a waiter actually just check a priority. Is there a potential binding mechanism here of ABX464 to the PAS biding domain in AhR? And then we actually took a number of AhR modulators and antagonist and tested them in a validated AhR luciferase reporter assay and also look to confirm that these ligands do indeed bind to the past be binding domain. And then largely driven by the pioneering work by Abivax in terms of identifying a regulation of miR-124 and providing a plethora of very, very well-established and validated protocols to monitor miR-124. We essentially copied a lot of this work to show does AhR modulation also similarly induce miR-124? And what is the relationship between these in terms of ligand-dependent AhR signaling and miR-124 expression? Now lastly, in a number of what we think are very well-validated systems of regulatory cells, namely regulatory macrophages, M2 macrophages or regulatory T cells, we use those systems to essentially activate AhR using EQ504 overfasemod and then looked at downstream induction of SIP 1A1, MIR-124, IL-1022 or in cells and model systems where we could look at regulatory effects, epithelial barrier repair and function, as well as regeneration. So let's talk about the first of those, in silico modeling. So let's take a second to think about what the binding domain of AhR looks like. It's shown here on the right-hand side. We call this the AhR PassB binding the makers, where the canonical ligands bind to AhR. Now I think about this as kind of a baseball glove. It's got this large hydrophobic cleft that's able to accommodate the binding of a number of different chemical scaffold, quite a large variety indeed. On the left-hand side, you can see endogenous ligands, natural ligands and Xenobiotics and pollutants where this covers a wide variety of different chemical scaffolds. In fact, there are, as of 2017, 13 approved drugs that we know bind and activate AhR. Now last year, a computational study looking at similarity based screenings of the library, found there were over 77,000 potential AhR binding molecules. This highlights the fact that the AhR binding domain as a xeno sensor taking chemical signals and turning them into biological functions can accommodate a wide variety of chemical scaffolds. So with that in mind, we took the core scaffold of ABX464 and took a look at it for its properties related to AhR modulation. And it does take a number of those boxes in that it's flat planar, confirmational constrained, has extended aromatic or heterocyclic systems and has moderate to high lipophilicity. And then we tested that not just against any compound, but against known AhR modulators. And there are three that will come up here. Laquinimod which is a and active biotech studied molecule. This molecule has been in Crohn's patients, MS patients. This molecule is a potent AhR agonist, but unfortunately, has poor tolerability in patients. Now what you can see here is the overlay of ovefazimod ABX464 behind Lekulamodel. And we can see that the quinolone core, overlaps for elegantly. And then there's also an extended aromatic system to the ringer. And you can see the same with Equilion's-AQ312, which is essentially an analog. And then very interestingly, there's some overlap here with Lilly's recent patent portfolio of AhR modulators, and this has turned compound #68. So again, overlap of that Quinlin core and extended aromatic system and good shape, lipopolicity and hydrophobicity complementarity. So moving forward from this, we actually did some molecular docking. We used move from chemical computing group to do a number of induced fit docking simulation. So we took a number of experimentally solved structures, more specifically the one with indirubin, which is the PDP 7ZUB, identified some of the key elements of the binding pocket and then ran docking simulations with ABX464. Now what we can see from this is that on the left-hand side, indirubin and its experimentally resolved structure binds very much into the -- very well into this hydrophobic pocket termed by the green colors here. And then you can see on the right-hand side, Pheno Alain 295, which is a conserved residue that creates stacking interactions with many known AhR modulators. On the right-hand side, the model of ABX464 also shows the similar binding pattern. And if we overlay the residues here, you can see very strong hydrophobic stacking interactions with this FEED 295, very similar to what we observed with the experimentally resolved structure of indirubin. Now if we take a look here and ask, what are the effects of glucuronidation on ABX464 and its potential binding to the past domain? So if we take a look at the right-hand side here, you can see the 2-dimensional structure of ABX464. On the right -- on the left-hand side, you can see that when you rotate this molecule through 90 and then another 90 degrees, it's very flat, planer and fits very well into the binding pocket. But unfortunately, when you do glucuronidation to this molecule, this planarity has lost. Steric hindrance is introduced and a lot of polar steric bulk is also a potential hindrance to the binding in the AhR past B domain. So we believe that ABX464 would likely be active as an AhR modulator. But the englucaronidated the metabolite form in humans would likely be inactive. So actually test this. We moved on to using the AhR luciferase reporter assay to compare potency of known AhR modulators. This is an assay that provides a highly sensitive quantitative and physiologically relevant readout directly linking ligand binding to transcriptional output. And as you can see on the right-hand side here, tested ABX464, Indigo Naturalist, tapinarof, indirubin, ITE and ABX464 and glucuronidate. Now these molecules are available for purchase. These molecules were ever purchased online and/or in the terms of ABX464, we actually synthesized the englucaronidated version using human liver microsomes to get the metabolite. Now what you can see is the EQ504, as has been disclosed before, is a very potent and selective AhR modulator. And then you see indirubin Indigo at trials to Pinrov. And ABX464 is a moderately potent HR agonist with an EC50 of about 400 nanomolar. ITE, the precursor molecule, the very well-studied AhR modulator derived naturally in human cells, ITE has an EC50 of about 460. Now the englucaronidated version as predicted from the molecular modeling does not bind. Now of course, this is a cellular model and the purpose of glucuronidation to molecules is to typically retard entrants into cells as part of a metabolism and clearance mechanism, so this could be either lack of entry of the glucuronidated form into the cells or if it does get into the cells, a lack of binding to the aryl hydrocarbon receptor for the reasons previously disclosed. Now another important element here is to really prove that the molecules bind to the PaaSB canonical binding domain of AhR. And for this, we use a high affinity AhR antagonist called GNF-351 has an IC50 of about 62 nanomolar. This has been widely used as a research tool to specifically block AhR transcriptional responses in vitro to confirm AhR-specific attribution in multiple cell types. So on the right-hand side, you can see that when we take the highest concentrations of the EQ504 tested or ABX464 tested in the potency assay. And then we add to that 500 nanomolar of GNF and that is a 1:5 ratio of drug to antagonist or a 2:1 ratio of ABX464 to antagonist. You can still see very effective inhibition of the signal, and very effective competition of the AhR modulator and resulting in its lack of transcriptional responses. So a very elegant way to demonstrate that this is not only an AhR modulator that it binds to the PaaSB domain and that it can be completed in terms of signal proliferation. Now let's switch gears. Now we know that AhR modulation can be induced by ABX464, but not by its glucuronidated metabolite. Let's take a look at does miR-124 get induced by activation of the AhR pathways. Now for this, as I pointed out previously, we largely leveraged the Abivax data from the 2022 paper, which does an excellent job of really defining protocols for which you can study the upper regulation of miR-124 in a number of different cell types. And we've selected some data here to show you where it was originally shown to be very strong for ABX464. And then we've also shown you side-by-side EQ504. Now on the left-hand side, what you can see is the level of induction of miR-124 compared to DMSO control, of EQ504 at 5,550 as well as ABX464 and the glucuronidated version of ABX464 again at 5,550. And it's very clear that ABX464 can induce miR-124 that the glucuronidated version here does not induce miR-124. And similarly, with the other AhR agonist that we're testing here EQ504, it is also a strong inducer of miR-124. So new data here that would hint towards miR-124 being a pathway downstream of AhR activation. Now these results correlated actually very well with the reported data in the it paper here shown by Figure 1. Now we also took a look at the cytokines that are inhibited. Again, really leveraging the data from the Aplite paper where IL-17 and IL-6 were strongly inhibited by ABX464 at 5,000 nanomolar. We tested the 50-nanomolar EQ504, which is typically around where we see the EC50 of our molecule across multiple cell types. ABX464 the originally tested concentration and then for a side-by-side comparison, ABX464 also at 50-nanomolar. And what you can see here is a nice inhibition of IL-17 and IL-6 in a statistically significant manner. And again, very well -- very correlated to exactly what was seen in the 2022 paper. Now what's interesting is the dietary ligands of AhR such as the indole, triptofan rivatives are known to be very strong inducers of IL-22. And we've seen this across multiple cell types previously where we see very strong induction of IL-22. Now in this system where you have activated PBMC. So this is not just normal PBMCs. These are PBMCs that have been reactivated, what we can see is that EQ504 is a very strong inducer of IL-22 compared to ABX464 of either 5,000 or 50. And again, this comes back to the idea of biased agonism. And that not all ligands or activators of AhR and list the same transcriptional responses or signatures. And one of the things that we would draw on to EQ504 was it's a very strong inducer of IL-22. So it was nice to see this here once again replicated in the same system. Now narrowing down within PBMCs, the offers focused in on CD4-positive T cells. They were able to sort and isolate CD4 T cells and show that miR-124 could be induced with ABX464. We replicated that experiment. But in this experiment, we also introduced the AhR antagonist. So we used 50, 500 and 5,000 as the concentrations for ABX464 compared to EQ504 at 50-nanomolar, and we kept a steady fixed level of AhR antagonist. Now the reason we do this is because this way we can be sure that the AhR antagonist is not introducing any idiosyncratic effect. But more importantly, what this does allow us to do is create some quasi dose response in terms of the ratio between AhR, modulator and antagonist. And what we can see is very similar to what was previously reported as EQ504, ABX464 at 5,000 or 50 can induce miR-124. But very, very importantly, that AhR antagonism in a dose-dependent manner can actually reduce the levels of miR-124. So this would be the first set of data that really highlights that miR-124 is downstream and dependent on AhR activation. Now we further went on to actually look at the effects on IL-17 expressing cells in isolated CD4 cells. This is again another statistically significant finding from the paper, and we see a very similar commensurate decrease in the IL-17 expressing CD4 cells when either EQ504 or ABX464 at 5,000 or 50. So again, really just demonstrating that there's a lot of complementarity between our experiments and the previously published data. Now moving on into macrophages, a very similar experiment here in terms of inducing miR-124 through AhR activation and then the addition of an AhR antagonist to show the effects on the AhR and miR-124 Axis. So what we can see on the left-hand side is compared to a DMS control of -- we tested EQ504 at 50, ABX464 5,000 and 50, and we introduced GNF at 500 nanomolar. So that represents a 1:10 ratio with EQ504, a 10:1 ratio of drug to an antagonist with the concentration of 5,000 of ABX464 and then a 1:10 ratio when we look at ABX464 at 50 nanomolar. Now very interestingly, in all of these concentrations, we see strong miR-124 induction and then strong inhibition when we -- of miR-124 induction when we block the AhR activation. Now similarly, looking at the cytokines that are induced, two of the cytokines that were highlighted in the paper were TNF-alpha and IL-6, I think we see here a very strong reduction in those cytokines, and we see that map across to our experiments here with EQ504 and ABX464, very statistically significant finding in terms of levels of reduction. Now we went beyond just TNF-alpha and IL-6 and actually looked at reductions in IL-23, IL-12 and IL-1 beta. And we saw a very strong significant decrease with EQ504. And in the context of IL-23, we saw a similar decrease of ABX464. Now it did appear that EQ504 was a more efficient block of IL-12 and IL-1 beta. And again, these could be differences from ligand-dependent bias agonism or signature responses. So -- but this just does highlight the ability of AhR modulation in dampening down multiple pro-inflammatory cytokines. So with that in mind, I think that data does very effectively demonstrate that miR-124 induction is downstream of AhR activation. So the next question we wanted to understand was in cells that are inducers of SIP 1A1, miR-124, IL-10 and 22, what is the functional consequence of blockade of AhR activation on those systems? Now today, we'll just talk about regulatory T cells, but in the larger deck, you'll see the data from the macrophages. And these are two systems that Equillium has published quite extensively on, including more recently at the AAI conference. Now if we take T these are cells that have been differentiated into T regulatory cells then treated with either EQ504 or ABX464 and then with the addition of an AhR antagonist, Here, we're looking at the relative responses to SIP 1A1 and miR-124. And as is the signature of AhR activation, we can see induction of SIP 1A1 in these T regulatory cells, and we can also see induction of miR-124. Now when you add to this the antagonist, GNF at 500, which represents a 1:10, 10:1 and 1:10 ratio across these concentrations. You can see a nice clear inhibition of induction of SIP 1A1 induction at an activation marker as well as a reduction in miR-124, again highlighting that miR-124 is a downstream effect of AhR activation. Now when we do the same experiment, and this time, we actually look at the effects of induction of IL-10 and 22, we see again a very similar response. In the induction of IL-10 and IL-22 is AhR dependent. And it happens with both EQ504 as a known AhR modulator as well as ABX464 and the addition of GNF at concentration of 500, again, representing 1:10, 10:1, 1:10 ratio, you can see a nice inhibition of induction of these anti-inflammatory cytokines. Now again, we won't go into necessarily the macrophage data of this. The short story is there, it shows exactly the same relationship. So I urge you to maybe download the deck later and take a look at that. Now what about the induction of SIP 1A1 on miR-124 by AhR activation in intestinal epithelial cell lines? Now previously reported by Abivax in the 2022 paper is that miR-124 induction is not seen in epithelial cells. And that's what's replicated here on the right-hand side. Regardless of the concentration, we show here a nice dose response, we do not see activate induction of miR-124 by either EQ504 or ABX464. But you do still have AhR modulation as demonstrated by induction of SIP 1A1 on the left-hand side. So a nice dose response with EQ504. And you see at the higher levels of ABX464 induction of SIP 1A1. Now moving forward, we take the 50 -- 100 nanomolar concentration here from EQ504 because this is largely where the molecule tends to plateau well. And then we took the 1,000 nanomolar concentration, which is the maximal concentration here tested for ABX464. Now if you look at those cells, again, using 100 of EQ504 or 1,000 nanomolard of ABX464. And add to this 2,500 nanomolar of GNF. We use a lot more inhibitor here needed to be optimized, likely because the inhibitor has poor transit into these epithelial cells. But what you can see from a perspective of cause and effect is the activation of AhR and induction of SIP 1A1 happens with EQ504 and ABX464. And then the addition of GNF and AhR antagonist commensurately decreases that. So with EQ504, that's a 1:25 ratio and with ABX464 that's a 1:2.5 ratio, and that's why we see a sort of quasi dose response here. Now what are the functional consequences of AhR antagonism in epithelial cells. So we're not seeing miR-124 induction, but we are seeing activation of SIP 1A1. So what does that do functionally within these cells? Now again, there's two sets of data, but I'll be just presenting the wound healing data here, but I do urge you to take a look at the barrier integrity data by assay at a later date. So this is an experimental model that we published quite widely on, including more recently at the American Academy of immunology, where we actually tested this against indirubin another clinically validated AhR modulator. Now as a wound healing asset, you take these layers of cells, you propagate them and then you score them to create a scratch. And then you monitor the healing rates over time. And what you can see here on the right-hand side are the percentage wound area compared to the DMSO control after 4 days. And we can see after 4 days, the DMSO vehicle has about 40% wound area. But EQ504 or ABX464 at 100 and 1,000 nanomolar, respectively, significantly improve wound healing by about double of that. But when you add an AhR antagonist into this experiment, you worsen wound healing, suggesting that the wound healing via AhR is also AhR dependent and is worsened when you inhibit the AhR dependent pathways. Now this is shown in this data specifically when we look at the biomarker for activation of AhR SIP 1A1. So we take these 284 cells that have been scored. And at the end of those 4 days, we take a look at the induction of SIP 1A1 compared to the DMSO control. And as we've shown in immune cells, EQ504 or ABX464 to 100 or 1,000 nanomolar, respectively, induce SIP 1A1 as a biomarker of activation. And similarly, when we add in the antagonist at either a 1:25 for EQ504 or a 1:2.5 ratio for ABX464, then we see a very strong inhibition of induction of SIP 1A1 again, highlighting the AhR dependency of these molecules. Now lastly, one of the important biomarkers for epithelial barrier function and repair is clouding too. Elevated levels of protein to have been associated with leaky gut or impaired gut barrier function. It decreased -- its increased, sorry, in ostericolitis and DSS colitis models. AhR activation has been associated with reduced chlordinib levels in intestinal models. And decreased claudin 2 may reflect restoration of epithelial barrier function following AhR agonist treatment. Now a lot of this data is already published and those publications are listed there in the footnote. And what you see here is similar to what happens with wound healing and SIP 1A1, the marker of downstream induction -- is induced, sorry, is reduced when you add an AhR modulator like EQ504 or ABX464. So you can see those nice reductions, statistically significant reductions in the levels of cordoning 2. And then when you add an AhR antagonist, those levels shoot up. Now interestingly, those levels get even worse suggesting that AhR has sort of tonic signaling properties in modulating barrier function and further inhibition of AhR activity worsens barrier function. Now that's an observation that's been replicated elsewhere. We know that if you knock out AhR, barrier tissues actually become leaky. -- claudin levels rise. And so this is a very important finding here in terms of looking at a functional readout for AhR modulation via either EQ504 or ABX464. Now it also goes a long way to perhaps shedding light on why Abivax's ABX464 is effective in the treatment of ulcerative colitis, because not only it's modulation of immune cells, but it's modulation of barrier function, repair and regeneration in epithelial cells. So I think this is a new, really important finding and one that I think further expands the value proposition of -- value proposition of AhR modulation and the miR-124 axis. Now let's just quickly go through the summary and conclusions. Is ABX464 and AhR modulator? Well, our results would suggest, yes. It's a moderately potent AhR modulator. We have a potency of about 400 nanomolar that binds to the canonical PassB domain of AhR. But the ABX464 and glucuronidated version does not bind nomodulate-AhR. Now again, this could be a lack of entrance by the cell, given that these polar glucuronidation are meant to inhibit cell transfer or it could be that if it's in the cell, that polysteric bulk and lot of planarity precludes its binding and activation of AhR. The next question was, does AhR modulation induce miR-124? So again, using a lot of the pre-published work by Abivax in the 2022 paper, HR modulation by EQ504 or ABX464, but not by ABX4 does induce miR-124 and decreases pro-inflammatory cytokines in multiple cell types. So that's very interesting that here, this data would also suggest that it's AhR dependent signaling leading to the induction of miR-124. Whether you use an antagonist or a modulator like ABX464 that does not induce AhR activation, then you do not see the induction of miR-124. Now there is no miR-124 induction in epithelial cell line, that's a previously reported finding. So it seems that miR-124 is contextually important just for immune cells. Now from a functional perspective, we know that AhR modulation is effective at inducing downstream pathways, including miR-124 in immune cells and epithelial cells. But what was really nice here was that using functional models of inflammation, tissue barrier function and regeneration, we should AhR antagonism inhibits the induction of SIP 1A1, miR-124, IL-10 and IL-22 in T-regulatory cells and macrophages, and it also inhibits SIP 1A1 and Cloudant 2 in epithelial cells worsening the healing. Now again, not shown here, but I urge you to take a look at the barrier function assays included in the larger deck is that we also see the AhR antagonism reduces and inhibit SIP 1A1 in epithelial cell lines in worsening the barrier integrity. So that sort of moves us on to really this last slide, which I think moves us towards a more unified mechanistic framework for how we think about ligand-dependent AhR signaling and miR-124. So this is an agglomeration of work that's published by ourselves, by Abivax, but also the academic community. And that would suggest that when AhR modulators bind to AhR and activate it, it induces downstream pathways. It induces modulation of immune and barrier function proteins. It induces expression of xenobiotic enzymes that terminate the signal and remove the ligand from the system via SIP 1A1 or glucuronidation. And then it also induces a number of microRNAs shown here specifically for today's context, miR-124. We know that other mirs, such as MiR, I think, it's 355, are also related to T-effector cells and IL-10 signaling. So the fact that mirs are downstream markers of ligand activated transcription in the AhR pathway should not be a surprise. Now what's interesting is that the SIP 1A1 or glucuronidation negative feedback mechanisms leading to ligand metabolism or the induction of miR-124 that leads to lowering of AhR signaling could be parts of the regulatory framework that help modulate AhR activation in cells. But overall, what this shows is that modulation of the AhR pathway via AhR modulators such as EQ504 or ABX464, reduce inflammation and fibrosis and increased barrier function and repair mechanisms. So with that, I'd like to thank the key acknowledgment here of the Equillium research team and pass this back to the moderator for Q&A.

Great. Thank you, Stephen. Yes. So at this time, we will be conducting a question-and-answer session. So please hold for a brief moment while we poll for questions. Our first question comes from Steve Seedhouse at Cantor Fitzgerald.

Great. Thanks so much. Very interesting studies. Thanks for hosting this. Wanted to work through a couple of interesting questions I had here. Just first regarding the mechanism behind miR-124 induction. Do you think it's transcriptional? Because the working hypothesis for obofasimod, I think it's around splicing regulation, but it seems just more likely that it would be transcriptionally induced. I'm just curious what you think?

Yes. We're not experts at miR-124, so all I can do is give you my sort of opinions and some hypothesis here. When AhR is activated, it can bind to DNA elements, right, which is where we'd likely see genomic elements like me as be generated, but it can also induce co-factors. I think what we can tell from this is that it's very clearly AhR modulated. Is that the only mechanism? It seems like from this data would be a very much a dominant mechanism here. Now is it transcription or is it something related to AhR activating something and that leading to modulation in miR-124? I don't think these experiments go to that level of detail. I think from our perspective, the key question was modulate AhR, does that modulate miR-124? The exact mechanism here, I believe, is transcriptional. But I think further work would be required. I think the alternative mechanism here and that it binds to the cap binding domain, which has been demonstrated and that changes splicing. It would be very interesting to identify ways to be able to blockade that pathway and actually look at the impact on sales. That would be a way to sort of tease out relative contribution there. But I think we can all appreciate that using an AhR antagonist that then blocks downstream miR-124 induction does very, very strongly suggest this is a downstream induced market of AhR activation.

Yes. Great points. Just on the -- on the negative regulatory activity that miR-124 has on. So in your experience that you've shown here, did you look at AhR expression over time? Like what is happening there? Is it getting down regulated -- and then what do you think will happen in vivo? In other words, what are the implications here for the durability of the efficacy like in a maintenance setting in IBD, for instance?

Yes. So we've looked at this in immune cells. And that data is in the broader deck. We did this in the T regulatory cells. We see a reduction in HR expression by about maybe 20% to 30%. So it doesn't cause the wholesale loss of of the receptor. Now we should maybe think about how the ZAL research was completed. They very, very heavily overexpressed miR-124. So we can't really reconcile the concentrations of miR-124. Now we've already known in the literature that when you activate the ligand and the receptor, the ligand gets metabolized or shuttled out of the cell and moved away. There are represses that turn off the signal and the protein of AhR can be moved on to the proteasome for degradation. So lots of AhR after activation has been previously reported. Now the miR-124 Axis is sort of interesting because it would be another element that is sort of a regulatory element in response to activation and that it's generated and then it sort of dampens the signal. So we do see a reduction in AhR expression, total protein expression, but this doesn't, we believe, result in a meaningful drop in protein available for signaling. You also see this that in Crohn's patients, there does appear to be a reduction in AhR, whereas ostaive colitis patients, there does not seem to be much of a change in the level of -- change in the levels of AhR expression. And then lastly, I'd say that we've done animal models that have dosed for prolonged periods of time, and we can continue to see sustained levels of activation and efficacy, suggesting that there's plenty enough AhR still present as it sort of is made used, recycled made used recycled, et cetera, that would allow us to continue to stimulate that pathway in a durable fashion fourth, as you point out, maintenance dosing in ulcerative colitis.

So just as a follow-up on that, just the last question for me today. The -- do you think that -- so given the miR-124 may have this negative regulatory effect, but it doesn't seem to be sort of fully abolishing what AhR agonism is doing that's constructive. Do you think miR-124 separately has its own independent anti-inflammatory effects separate from that regulatory action such that maybe on net its induction is helpful? And just because looking at the -- I'm just curious if the degree of miR-124 induction is relevant here for the efficacy translating into the clinic. And if -- or if you just be better off having an AhR agonist that doesn't induce miR-124 at all?

Yes. So we've tested other working backwards. We've tested a number of different AhR modulators, indirubin, et cetera, to -- and these do also induce miR-124, but we focused in on EQ504 for this presentation. So it does seem that AhR modulation broadly does induce miR-124. So then your question is, does miR-124 on itself induce an anti-inflammatory effect? Kind of like if you induce IL-10, induce IL-22, these are all functional proteins claudin I mean there are a laundry list of things that are regulated by AhR that contribute to therapeutic efficacy here. So this study, and we should be fair to what this study helps us answer and what it doesn't help us answer is it shows that miR-124 is induced by AhR, but it doesn't actually levy the contribution in terms of anti-inflammatory effects from miR-124 or other components of the AhR pathway. I think that experiment one would want to just add in miR-124 in the absence of an inducer like an AhR activator, et cetera. And that would be a key experiment where one could really ascribe some anti-inflammatory, antifibrotic tissue barrier function effects to the molecule itself miR-124. The challenge we have in our experiments, of course, be it the original Aplite paper or our work here today is that you're always giving an HR modulator. So it's very hard, if not impossible, to tease out the relative contribution. How I generally see it is that if it does have effect, which is great, it's induced by AhR activators, right? So it comes back down to the original point here, which was -- when we set out to do these studies, it was to ask a lot of questions from the academic and investment community is, what is the relationship between AhR and miR-124. And that miR-124 seems to be induced by AhR in an AhR dependent fashion. But it doesn't necessarily tell you what level of contribution that has to therapeutic efficacy. What I would say in the last point is that people have taken AhR modulators, put it into animal models and used anti-IL-10 antibodies, anti-IL-22 antibodies and they see a very strong reduction in the efficacy in those models by neutralizing IL-10, IL-22 and other pathways. So I'd say that if it is contributing, I would posit that it's probably a minor contribution compared to the others. But again, caveated but this is not a set of experiments meant to tease that out.

Our next question comes from Thomas Smith at Leerink.

Thanks so much for putting together this really comprehensive event. Really interesting data sets. Just a couple for us. Just on the wound healing assay, it looks like you aren't seeing miR-124 induction in epithelial cells. You have an AhR dependent relationship on a junction proteins. Wondering what do you think is driving that? Have you looked at other tight junction proteins beyond

We have, in other systems, we've done this in barrier function tissue tear assays. I think we've looked at the a variety. We can certainly follow up with that information at a later date. There are a number of those known in the literature -- now miR-124 was previously disclosed to not being upregulated in epithelial tissue. This was from the Aplite 2022 paper. So we were not necessarily surprised to find that it wasn't induced, but we do see the economical activation marker of AhR. Now when you look at the other mirrors that are related to AhR activation, they tend to be very contextual, meaning that they show up only in certain cell types. So I think the miRs here, as we think about the part of the downstream transcriptome of AhR would be cellularly contextual, right? So it comes back to Steve's question is, do we think it's driving efficacy, maybe in a certain cell type or its contribution is limited to a certain cell type that's reflected in this data here. So I think learning from what we've seen with other miR molecules is that it tends to be maybe important to certain cell types. So here it could be that it's a very important regulator of the AhR activation pathway in immune cells because it seems to be very predominant in immune cells, but it doesn't seem to have an effect in epithelial cells. Now conversely, the levels of induction of SIP 1A1 are much, much higher in epithelial cells than you see typically in immune cells unless you really hone in on a specific immune subset. So it could be that depending on the cell, they have their own specific intrinsic mechanisms for regulating downstream AhR activation.

Got it. That makes sense. And then maybe I could just ask -- maybe taking a step back, given what you're seeing mechanistically here, you've done all of this mechanistic work now. Maybe talk a little bit about how you expect that to translate clinically relative to And then maybe another corollary to that, obafasimad hasn't got restricted. I mean are there any implications, I guess, to your approach here with gut-restricted AhR modulation that we should be thinking about?

Yes. So super important question and one that we're early in our sort of clinical journey, right? So a lot of this will be better answered in humans. But I'll give you my opinions and hypothesis here on sort of the relative differences here maybe from a molecular perspective and then potential advantages through EQ504 and Equillium approach. Now we should first talk about the pharmacology. When you agonize something, you can decouple PK and PD. When you antagonize you largely want to be an IC90 sit on the receptor because every time you come off that receptor, it can signal. When you agonize you should think about this as you're pushing a ball down the hill. Once it's going, it's going, which is why you often see a disconnect between the PK/PD relationship, but you also see cumulative responses in which that the downstream markers have durable pharmacodynamic effects even when the AhR receptor is no longer occupied. Now that's important because it sort of gets at one of the key questions I think we get asked a lot, which is if the englucaronidated version is not an active AhR modulator, but it has been hypothesized to be the active component in patients the parent molecule, which is an AhR modulator is in a minor component. So how do we reconcile that? How would you reconcile that? And there are three main ways we can think about this. I think the first one is that, again, with pulse agonism, you don't need to be on a receptor all the time. It's not total exposure sensitive system. It's really time on the receptor to turn it on and then once it's on, it's doing its thing. So you don't necessarily need high levels of continuous exposure to elicit those pharmacodynamic responses. So perhaps even just the shorter pulses of obafasemod before it becomes englucaronidated and accumulates as what we believe is a non-AhR modulating loyalty. Maybe that's enough. The next one is that the end glucuronidation can be deconjugated in the intestinal tissue. There's macrophages, neutrophils, and they do produced beta-glucuronidase. And I think that's likely to be a minor contribution here is that the molecule would find its way out into those tissues, BD glucuronidated and then become active in those tissues, it's certainly plausible. I think perhaps the most interesting point to think about here is enterohepatic recycling. Now the concentrations of the englucuronidated version of ovefasmod appear to be a bit of a biological and pharmacological outlier in that they are 5 to 20 micromole of concentration. This is going on. Unfortunately, the sort of scan data that I can that I can glean from the literature. I think this might be a better question post Abivax is that those very high levels of molecule. If even a fraction of that is excreted into the bile, which is a very natural mechanism for glucuronidation, remember, adding this polar group is meant to excrete the molecule, the fact that we're seeing very large an extended PK times does suggest that there's some intrahepatic recycling. So even if a fraction of that gets endrohopatically recycled gets pushed into the bile into the colon, where the long residence time in the colon allows deconjugation from the beta glucarolidays is there to the parent AhR activating molecule and memory absorption. That does give you a potential mechanism for gut enrichment, whereby you're actually shuttling the molecule out of the peripheral blood into bile, into the colon, deconjugated reuptake. And when that activates AhR lists its pharmacologic response as that moves back into the plasma, it gets glucuronidated and that cycle continues. Now we don't in the PK data that I've seen, see a classical intrahepatic recycling second peak as they call it. But the likelihood is that there's such high concentrations in the peripheral blood that I'd point out, even if a fraction of this is enterohepatically recycled, it's pharmacodynamically meaningful, but it may not show as a sharp peak. It may just show more as a blended sort of tail to the PK. I think what would be really interesting is understanding the levels of ABX464 as its parent molecule in intestinal tissues. And answer the question, how much of AhR are we actively modulating in those tissues. And then you could understand is this fully saturated. Is this as much efficacy that we can get out of the molecule? Or is there more to be had, which is why our approach has been largely to get the molecule directly to colon tissues because it's the epithelial cells, the resident laminaproprias cells that we're modulating those are the pharmacologically relevant cells. And to avoid systemic exposures by creating these high local low systemic ratios.

Got it. That makes a lot of sense. And maybe last question on my end. Can we get an update on some of the clinical preparation formulation work that's ongoing? And just remind us what are the outstanding gating factors to getting 504 in the patients here?

Yes. No problem. Thanks, Tom. Well, look, the short of that is there's been no change. I'd refer you to the corporate deck there. The guidance still holds that we expect to initiate the study here in midyear, which we believe it brings up to Q3. So there'll be future updates pending on that as and when they're available, but I'll defer to the corporate deck for all of the deferred guidance.

Our next question comes from Alex Thompson at Stifel.

Great. Appreciate the presentation. Maybe to follow up a little bit on Tom's question. As you're thinking about 504, I think restriction, in particular, is there a rate or do you expect 504 to be glucuronidate or metabolize more rapidly or less rapidly than opefazimod? And is it really the time to modulate AhR that matters here? Or is it the way that you are modulating AhR kind of as it relates to kind of the downstream signal that you're seeing in sales at this point that might matter clinically?

Yes. So each AhR modulate has its own unique properties, right? And I think by and large, what you want to induce is less of the xenobiotic more of the immune, right? Inducing SIP 1A1, there are 13 approved drugs. -- inducing SIP 1A1 on itself is not a toxic attribute of the molecules or anything else that. So things that we've seen with the dioxane or even the signals of Indigo Natalist, largely idiosyncratic to the molecule and unlikely to be a class effect of AhR modulation. So when we think about giving a molecule directly to the colon, we're not thinking about this as got restricted. I tend to avoid using that term because I don't want to give the impression that this has some active restriction mechanism that keeps a molecule in the GI tract and for bids from going out anywhere else. I think that would probably be an overreach. We know that when molecules get into GI tissues that some will get out into the plasma. Now in our toxicology studies and in the literature, there's no specific predefined level for which you want to stay under. This is really just about tipping the scales in our favor, which is -- the cells that we want to activate are the epithelial cells, the immune cells that are in the and appropriate. We don't believe we're getting any bang for our buck from modulating cells in the peripheral blood. Sometimes you can see changes in the peripheral blood because of what you've changed in the in the colon. So that's always been an area of controversy. So we want to deliver the molecule directly to the colon tissues. And the same properties that allow you to get into the tissues are likely the same properties that are going to get you into the plasma. But we've seen it with steroids, multiple other different molecules is that if you can deliver the molecule to GI tissues directly, you see high tissue concentrations and low systemic. That allows us to drop the total DUCs. And in the modeling, which we can give an update on later in the year, our modeling, as would be expected, shows that we can get high tissue concentrations, lower systemic concentrations. And again, we believe we're going to be well below the level of important activation of AhR and the peripheral, but certainly very much adequate in those colon tissues to activate AhR and get all of our signal of efficacy. So I think we'll learn more out of the Phase I in terms of how much those ratios pan out. But I think it's hard to say anything here about comparative efficacy to overfasma. That certainly wasn't the intention of these studies, it was really about what is the relationship between AhR and miR-124. I just think the advantage we have is a molecule de novo designed to modulate AhR. It's based off the naturally occurring strong immune and barrier function modulating endogenous AhR ligand, and we're going to deliver this and monitor its levels in colons to make sure that we're maximizing all of the efficacy on target for AhR. Now if anything would be one of the blind spots with the ovafasemod and ABX464 data set is we just don't know how much AhR would be modulating in tissues.

Great. And then maybe as a follow-up, as we think about the Phase I study and you've kind of framed it as a proof of mechanism study. What kind of biomarkers PD or otherwise, should we be thinking about in that sort of data set?

Yes. So I'd say a win for us would be getting tissue concentrations into the levels that we expect for EC50, EC90. The next one is SIP 1A1 induction. That's a conomical biomarker. The reason that's so widely used is because it happens in all cell types -- differential levels, but it happens in all cell types. It's very predictable. And it's a very clear signal above the background noise really -- it's a very transient biomarker. And so that allows us to really look at dose response. The other downstream markets, which we may or may not see in normal healthy volunteers because you typically require activated cells, right? So it's typically more important for cells in the gut or cells that are activated is things like IL-10, IL-22, changes in other biomarkers. So we will do some pretty broad transcriptomic analysis on an exploratory data basis. But what we know from all of the translational work completed by ourselves and others is that activation of SIP 1A1 is indicative of downstream translation -- transcription, sorry, of important genes to modulating the immune and barrier function. And this is meant to really bridge what we know about the atrial experience where they've dosed it to many hundreds, potentially thousands of patients. And in those data sets, they've shown on target activation of AhR by induction of SIP 1A1. So I think that's how we think about this from a proof of mechanism perspective.

So our next question comes from Jacob Herbert at Raymond James.

And I really appreciate how thorough you were on the slides today. So I guess just a quick follow-up to one of the last questions. In the event that you do measure microRNA 124 and the Phase 1 healthies. Would you be able to help frame for us if there's a full level of induction that you would see as derisking?

For miR-124, we're not the experts there, right, again, caveat with all of these experiments are really meant to tease out the relationship, right, mechanistic pathway profiling less perhaps comparative pharmacology here. Now what I would say is we would most likely now knowing that EQ504 modulates miR-124. And that it's a good biomarker in blood, but perhaps less so in epithelial tissues, which we'll be collecting is that I think we'd be it behooves to collect that and take a look at it. Now the increased levels of miR-124, they did not appear to correlate dose responsively with SIP or even the downstream market. So that's where I would caution is that we just don't know enough perhaps in terms of how they correlate to other markets. That's why SIP 1A1 is so widely studied and so widely relied on is that it's just a very well-characterized cause and effect system for pharmacodynamics of HR modulation. So I'd say we'd collect it, try to make sense of it and perhaps between now and getting that clinical data read, we can perhaps look to get a better understanding of those levels. But I would say that it -- so far, because it looks like it's a market that might accumulate so multiple doses lead to accumulation and then you see a plateau those types of biomarkers typically don't give you the best read on dose response because depending temporarily on when you look for them, we see that a little in our data today. We're looking so late down the line is that you've saturated and accumulated all of your response. So I think markets like SIP 1A1, which are more transient and happened earlier and in a more dose responsive fashion, might just be more enriching from a dose selection perspective than miR-124, which may accumulate and persist saturating the signal, so you might lose your ability in dynamic range.

That's very helpful. And then I guess just one follow-up on the glucuronidation I apologize if I missed this detail. Is EQ504 glucarona data? And if not, would you expect that to be an advantage versus obafazimod in terms of potentially having a longer duration of action?

Yes. I'm not sure there's -- we can never call it advantage or disadvantage at this stage, because we don't know really the effects of humans. Typically, glucuronidation should cause the rapid excretion of a molecule. So to see the accumulated suggest either high plasma binding high plasma protein binding, so it just sort of sits around in the plasma, you create this larger sync. There's perhaps little to no benefit of that happening. If it's not the active species, right, which is what the data here would suggest. It doesn't mean that it can't then release it as a sync to become pharmacologically active elsewhere. But it's hard to really tease out whether that's an advantage or disadvantage for us in our molecule. What I can tell you about our molecule is that we don't have extended persistent accumulation in the plasma. We have good tissue retention in epithelial cells. And then we have fairly rapid plasma clearance. And that's likely a contribution of sip enzymes, glucuronidation et cetera. But from our early animal models, we have not seen our toxicology PK models a mechanism like obafazimod, where it is almost entirely or predominantly glucuronidated and then that changes the pharmacodynamic profile. We have a very well-behaved typical sort of decay curve again in animals of EQ504. And that's essentially what we're looking for is activation within the tissues. It gets out to the plasma. It gets metabolized, no active metabolites, no quirkiness, no additional accumulation mechanisms. What would be interesting is if what you see with ovafasemod, which I do believe is an advantage for the molecule is that you don't really want AhR activation systemically. Ideally, it just happens locally. And that's largely just to avoid over metabolizing your molecule. So by actually modifying the molecule to not be an AhR modular. It might just be a protective masking mechanism, which again is inherent to AhR as a pathway. So there you could argue perhaps could be an advantage. But it's not a prerequisite. That's not what is absolutely required. You don't need to necessarily design that in I think what you want to see is rapid clearance from the plasma, rapid uptake in the GI tissues and this asymmetry in tissue versus plasma concentrations that leads to activation in the tissue, but no activation in the plasma.

Our next question comes from Leland Gershell at Oppenheimer.

Great. Thank you, Stephen. This has been a terrific presentation, and we appreciate the updated data. A few questions from us. First, I wanted to ask, I guess, teeing off the point you made earlier about with the sort of an agonist condition. I need to have the molecule on the receptor for very long. It can have durable downstream effects. Have you looked at the time course between obefazimad and EQ504 in terms of the time course of action and expression of miR-124? And also wanted to ask your opinion, obviously, you've gone a long way here in terms of the science delineating the pathways between HR and miR-124. Do you think that this negative feedback mechanism is sufficient to sort of help us explain and resolve the conflicting evidence between some of those preclinical data and the clinical data we've seen from

Yes. So working backwards, I'm not -- again, not the expert of all of that data in hand. I'd say we really set out this to sort of do mechanistic attribution and pathway profiling. -- there are some disadvantages or limitations, deficiencies in using cellular models. And that is when we add the drug, it's on the cells, right? And then actually in the Aplite paper, the drug is added over multiple times. So you assume in all of those cellular models, drug is always on the receptor because it's always in the system. That doesn't reflect what happens in humans. The drug is dosed, it gets on the receptor, then it's cleared and cleared from the system. So it doesn't necessarily reflect the cellular models well in that way. So we should just acknowledge the deficiencies, limitations of cellular models. We have done experiments where we compared this to indirubin, which is the relative induction of SIP 1A1 versus IL-10 and IL-22. Again, this isn't about how much time is required on the receptor because in all of these cellular models, the molecule is always on the receptor essentially in those scenarios. What we did see is that indirubin induces a lot more SIP 1A1 per, call it, pound of or whatever metric you want to use of IL-10 and 22. And that's sort of known in the literature as I the precursor molecule for EQ504 is a very strong inducer of IL-10 and 22. It's less of an inducer of SIP 1A1. But in both of those experiments, the IL-10 and 22 or both of those molecules for indirubin and EQ504, the IL-10 and 22 goes up and it stays up for quite a long period of time where SIP 1A1 comes up and comes back down pretty quickly. So I'd say that, that demonstrates the sort of temporal effects that when you agonize the pathway, the downstream effects can be durable, but the limitation would be that there's no we'd have to wash molecule off or clear that system of molecule to then look at really a time decay there. So I really can't answer a sophisticated fashion about what it is the specific time one wants. I think from a clinical perspective, we'd be looking to see that we could durably induce SIP 1A1 in the tissues in a dose-responsive way. And we'll be looking for choosing doses that were either on the curve or into that plateau, and then we'd be moving that on into efficacy studies. So I think there's limited we can do on the bench with these agonists to tease out trying to thread the time on receptor loop.

And lastly, just a question as you have all this wonderful work you've been doing on the science and looking at different ligands of -- have you looked at or do you plan to perhaps look at the formation bio compound, KMR 301, which has been reported in the literature under, I think, a different name, but that's also a small molecule miR-124 inducer, may be more potent the nobofesmod has based on the same kind of core structure. Just wondering if you looked at any mechanistic activity of that compound?

Yes. And again, I think this comes down to sort of what our objectives internally is, I think uncovering the relationship between miR-124 and AhR is really important. I think that moves the needle. It helps us rationalize therapeutic effects. It can help you think about the design of molecules that can help you think about biomarkers as we've discussed on the call, looking at miR-124 now, given that's a clinical biomarker, right? There's a lot of value in that. I don't think what we're interested in doing is making analogs of other compounds. I think we're very confident in that we spent a lot of time looking for HR modulators. We settled on the analogs of IT again because they're naturally derived, right? So the parent molecule is the molecule that circulates in the lungs and guts of humans. It's a very strong and nontoxic agonist of AhR. We've developed analogs of that. So I think that we believe that we have a potent selective modulator with good drug-like properties. It has all the attributes that we're looking for. We now know that it can induce miR-124. And similarly, we now know that ABX464 is an AhR modulate. I think that relationship has been strengthened. I think from here on in, it really doesn't change what we do with EQ504. We have our plan. We understand what our molecule does. We understand how we want to deliver it. We're currently formulating that for the Phase I. So generating other molecules or whatever, I think has limited value to us and our development programs, I think from here on out, we'll just be focusing on what downstream pathways such as miR-124, for instance, and how that relates to clinical efficacy, how that might inform our own drug development program. Expanding formation Bio has a very similar molecule to ABX461. I'm not aware of any other molecules developed and sort of marketed necessarily as pure miR-124 inducers. But I think from today's presentation, one could assume that if you're an AhR modulator, you're also miR-124 and do so.

Our next question comes from William Wood at B. Riley.

I really appreciate the thorough data here today, Steve. Maybe just thinking about -- we've discussed a number of cytokines today in which you presented here. And then priorly, you -- a lot of focus has been on IL-10, IL-22, but also Abivax has been more focused a bit more on IL-17. IL-6. With just sort of the number of families here and the number of cytokines that you presented and discussed even just today, how should we think about what combinations or even what individual specific cytokines we should be thinking about possibly even families of cytokines, which may provide the best support for downstream clinical benefit? And then I have a follow-up.

Yes. So there's probably buckets, and that's why I put the top 4 because in reality, there's a lot more than -- the other part of this is that if I promised 50 things to show you 50 things move, you'd hold to it, right? So we've really tried to distill this, I think, to IL-10 and 22. And I'll walk you through the reason why. -- naturally derived HR ligands from the tryptophan metabolites in dolls, right? They're very strong inducers of IL-10 and 22. Those are typically referred to as hallmark cytokines of HR modulation. They're also very widely studied. ITE and indirubin, two naturally occur in one in one exogenous AhR model too much widely studied AhR modulators in the literature. Very strong induction of IL-10 and 22. And when you block IL-10 and 22, you lose efficacy, suggesting they're pretty dominant, right? But they're also play atrophic cytokines. So there's things that happen downstream. They're also very proximal to AhR activation. So we think about IL-10 and 22 as being directly transcriptionally derived from AhR activation. Whereas inhibition of pro-inflammatory cytokines is probably a little more distal, right? You're probably modulating NFKB and that's modulating other things and T cell differentiation. That's further downstream. I like to pick biomarkers that are very proximal to the activation event and they are really important to think about clinical efficacy. We know that if you knock out IL-10, IL-22 in animal models, they undergo a lot of gastroenteritis. There is some conflicting data with both of those. But I think in the context of looking at the barrier function and immune regulation effects, it's very clear, they're very important. We also know that in patients who have IBD, they have lower levels of IL-10 and 22. And when treated with molecules that derive efficacy, even SKYRIZI, I think there's some data here, too. is that they tend to improve levels of IL-10 and 22 because the system is starting to repair and restore. So I tend to think about things is that we have IL-10 and 22 because they seem to be centrally important to mucosal, biology and health and immune homeostasis and barrier function. But then you've got things like clouding, which are clearly very much regulated by the AhR access. People have studied those very widely. I think that's the second set of tissue markers would be looking at. And then you've got your phenotypic changes. Do you reduce levels of Th17 cells? Well, that wasn't a direct consequence of HR modulation. That's a consequence of the molecules or proteins that were transcribed by HR and a little bit further distal from the activating event. So I think really here, IL-10 and 22 are things that we'll focus on very heavily in the clinic. But in today's realm of being able to do transcriptomics and get tissues from the colons, we'd probably look to expand that set because it would always be more helpful if we had put more biomarkers to think about efficacy or patient population target selection -- but you can think about IL-10 and 22 is the sort of central access for how we think about AhR modulation and its therapeutic value proposition.

Appreciate that. And then I know we've been looking forward a little bit on our side, thinking about maintenance data coming up and then also there their data later this year in Crohn's. In terms of what you'll be looking for in that data set, those data sets that may give you confidence either in UC or possibly even Crohn's. What specifically might you be looking for, whether at the top line or digging down into the biochemical or biomarker data sets that really think you will really support your advancements going forward?

Yes. Well, first, there's a lot of kudos owed to here and those data sets. And the nature of ABX464, I think is game changing for ulcerative colitis, specifically, the maintenance data. Now what's interesting to us about the maintenance data is it's reflected in the Indigo Nitriles treatment data sets as well. So in the Indigo Naturalis treatment, they had very strong induction of remission rates that we gate. I think it was 40% to 50% depending on which one of those Phase II studies you look at. That's very strong. And that was still in a patient population that was mixed almost 50-50 between biologic-naive and biologic exposed. So pretty reflective of a contemporary population. When they did the longer-term studies of they saw deepening of those responses. They have more clinical remission in those patients, whereas typically for biologics or some of the other small molecules, what you get around a week 8 and 12 may not necessarily get better. So I think the maintenance data is super important for ulcerative colitis patients from something like Abivax because you see that if you can stay on drug longer, you can see those results improve. I think that's coming from things like IL-22, and that was demonstrated in the data sets, where R22 was correlated with mucosal healing, but it's a long-term value proposition, which continues to drive barrier function repair, regeneration, immune homeostasis -- so you'd expect something like IL-22 to have the properties of allowing the tissues to heal. When you use a JAK inhibitor and you're just blunt all of the immune responses, right? I mean I think there's diminishing returns from that -- from an efficacy perspective. And I think with something like IL-10 and 22, you're talking about restoring to a natural balance to mucosal health. So that is very intuitive to see why results get better over time, right? We use week 8 and 12 often, I wouldn't say arbitrarily, but that's how we test things in ulcerative colitis. But the week 52 clinical remission data is game changing for fast. And I would expect that given the data that we've seen today, knowing that it's modulating HR is I would expect that that's likely coming from pathways modulated by the HR. So I hope to see that they are successful in demonstrating very, very deep maintenance data. And I would expect that part of the value proposition of EQ504 going forward is that strong induction of early remission rates at week 8 or 12. I think that's where we can win. There's still obvious room for improvement. About 16% for oblafasmod Indigo naturals in the week 8 to 12 was getting 40% to 50%. So there's room for improvement there. But I think having that continued improvement to mucosal healing endoscopic remission and clinical remission by week 52. I think is the thing that really changes the game for patients and physicians.

So our final question comes from Adam Walsh at ROTH.

Stephen, amazing presentation. I appreciate all the detail. A couple here. The first, is there a publication strategy around what you've been presenting today and what would be the time line there? And kind of follow-on to that, is there an IP strategy that can be derived from this in terms of composition of matter, method of use or anything like that?

Yes. Well, -- last but not least, appreciate you joining, Adam, was good to hear from you. The kudos goes to the team here. I feel like this is a little bit like thanking the salesman for building a great car. It really goes down to the group here who did all this fantastic work. From a publication strategy, we will look to publish this data through additional conferences, potential publications around our own molecule where we might compare against other AhR modulators like -- and now we can add to that ABX464 and that's important because it's a molecule that's shown great efficacy at Phase III. IP strategy, we don't really ever discuss IP strategy here for obviously good reason. We're always very aware of what we can do from an perspective. I think we're more focused on our molecule than generating IP around anybody else's molecule. And I think largely, what we're doing is cooperating and further expanding work that's already out there in the public domain. People have tested overfasmod as an AhR modulator. People have shown that relationship. I think what we sought to do was really solidify that with some good mechanistic data orthogonal approaches antagonism, over to just sort of really understand the relationship. And now we can learn from clinical data, mechanistic profiling pathway analysis of other molecules, which I think is to the benefit of patients and general drug development. So I think that's all I can say at least from the publication strategy and IP. And remind me if I missed anything else in your question.

No, that's totally fair. That's great. Appreciate that. And then just one final one. In terms of the Phase I trial, you mentioned that SIP 1A1 is kind of the key induction to look at there with the downstream IL-10 and IL-22 potentially being more seen in UC patients, -- does that impact your decision point on whether to include Part B UC cohort? And will we -- and when will we hear information on whether that Part B UC cohort will be included or not?

Yes. So maybe working backwards on that. We are still working with our advisory board that we more recently put together with Bruce Sands, Get to Hans, Florian Reader, Simon Travis and Vipal Jira, we think a sort of really experienced team. We've engaged them early. We've got plenty of time to think about design. So I think you'll hear from us probably sometime in and around the Phase I, what we would be doing afterwards. Now you raised a point which is would we learn anything in the Phase I that may change Phase II. Again, I think that's something that we'll have to carefully consider as part of the long-term development plan. what we want to achieve in the Phase I is that we have a molecule that's safe, well tolerated, activates HR and colon tissues. And from a SIP 1A1 prospective, this really isn't about hitting a certain level because every AhR modulate it actually induces to a different level, it's levels of SIP 1A1. So we shouldn't see this as we have to hit the same as another molecule, really what you'd be looking for is that plateau. When you hit that plateau, you fully occupy the receptor, you fully turned on all the transcriptional machinery. And from our translational work in the lab, we know that correlates to maximized induction of all of these other downstream markets. So I think that's a win for us is showing that we can save, well tolerate, deliver drug to the colon, get to good drug concentrations of the drug, so we can engage SIP 1A1. And then I think it will be about selecting dose doses and moving that forward into an ulcerative colitis patient population. So I think more to come there. We're laser-focused on getting that Phase I up and running. We have at least received very strong feedback from the ulcerative colitis community that should this get into the realm of treating ulcerative collates patients. This would be a mechanism of high interest to them, given the experience they've seen and/or had with Indigo Naturals, which is the natural botanical HR modulator. And then lastly, I think we recently brought on Singhal, who recently came from Ventec. She was previously at me arena. She really adds to the experience, expertise and the brain trust era Equillium as it relates to ulcerative colitis trial. So super excited to have brought her on and she'll be leading the clinical development. So I certainly don't want to take any of her thunder this early in the game.

Great. Thanks for the question, Adam. So this concludes today's Q&A session. I'll now turn it back to you, Steve, for some closing remarks.

Well, thank you for everybody who joined and perhaps stayed on 20 minutes past. We do very much appreciate everybody's time. Please do visit the website in the next few hours. The presentation will become live. There will be some additional data. Should you wish to engage in any of the conversations that have been had here, please do reach out via the appropriate channels to Equillium, and we'll be more than happy to take additional questions. But once again, thank you, and good day.