Ladies and gentlemen, thank you for standing by. Welcome to the PTC fourth quarter 2020 financial results conference call. At this time, all participants are in a listen-only mode. After the speaker presentation, there will be a question and answer session. To ask a question during the session, you will need to press star one on your telephone. If you require any further assistance, please press star zero. I would now like to hand the conference to your speaker today, Kylie O'Keefe, Head of Investor Relations. Please go ahead, ma'am.
Good afternoon, and thank you for joining us to discuss the PTC Therapeutics fourth quarter and year-end 2020 corporate update and financial results. Joining me on today's call is our Chief Executive Officer, Stuart Peltz, our Chief Financial Officer, Emily Hill, our Chief Development Officer, Matthew Klein, and our Chief Business Officer, Eric Pauwels. Before we start, let me remind you that today's call will include forward-looking statements based on current expectations. Please take a moment to review the slide posted on our investor relations website in conjunction with the call, which contains our forward-looking statements. Our actual results could materially differ from these forward-looking statements, as any and such risk can materially and adversely affect our business and results of operation.
For a detailed description of applicable risks and uncertainties, we encourage you to review the company's most recent annual report, Form 10-K, filed with the Securities and Exchange Commission, as well as the company's other SEC filings. We will disclose certain non-GAAP information during this call. Information regarding our use of GAAP and non-GAAP financial measures and a reconciliation of GAAP to non-GAAP is available in today's earnings release. With that, let me now pass the call over to our CEO. Stuart?
Thanks, Kylie. Thanks for joining us today. In 2020, we have made significant progress in moving our pipeline forward to bring new therapies to patients on all fronts, from research and development through commercial. Despite the COVID-19 pandemic, we initiated five clinical trials, including two registration-directed trials with vatiquinone. Let me begin with the Duchenne Muscular Dystrophy franchise. We continue to see strong global growth and geographic expansion. Our DMD franchise sales in 2020 were approximately $331 million. Specifically, in the U.S., the annual revenue of Emflaza totaled $139 million, which is a 38% increase from last year. The annual revenue for Translarna totaled $192 million and was driven by geographic expansion, new patients, and label modifications. For example, Translarna received marketing authorization in Russia in the fourth quarter. We have also continued to drive additional geographic expansion in Central and Eastern Europe and in Latin America.
Even during the turmoil caused by the COVID-19 pandemic, we secured a Brazilian group purchase order for Translarna. Eric will go into these details shortly. As a reminder, we also recently reported the results from the 045 dystrophin study. We plan to discuss these results along with Translarna's totality of evidence, including existing clinical and real-world data, with the FDA. Our goal has always been to bring Translarna to U.S. patients who have long been waiting for this therapy as quickly as possible. Moving to our splicing platform, the approval of Evrysdi in 2020 was an important milestone for PTC, our partners Roche and the SMA Foundation. Evrysdi is a groundbreaking treatment, which has a benefit for all SMA patients with particular benefits of durability and broad tissue distribution.
As a consequence of the benefits of Evrysdi, it has continued to show a strong uptake, and Roche expects that Evrysdi will become the treatment of choice in the U.S. in 2021. With the near-term expected European approval followed by the Japanese approval, Evrysdi should see continued significant growth this year. Another accomplishment of 2020 with regards to Evrysdi is the royalty monetization deal, which put $650 million on our balance sheet. The structure of the deal allows PTC to receive approximately 60% of the royalty revenue and reverts to 100% once our royalty monetization partner receives $1.3 billion. Our next most advanced molecule from the validated splicing platform is PTC-518 for the treatment of Huntington's disease. As a reminder, PTC-518 is an orally bioavailable small molecule that crosses the blood-brain barrier and reaches all regions of the brain.
Preclinical results demonstrated a dose-dependent reduction in the HTT mRNA and protein in cells of the striatum cortex and cerebellum in the BACHD mouse model. This is critically important as Huntington's disease is a whole brain disease. HTT reduction is clearly titratable based on PTC-518 levels, so the degree of HTT lowering can be tightly controlled. In addition to the whole brain distribution, PTC-518 achieves uniform exposure and HTT lowering in all tissues analyzed, showing a near one-to-one ratio between CNS and blood. This is important because it shows the exposure and effect within the brain is analogous to what we see in the blood cell. PTC-518 is currently in a single and multiple ascending dose phase I trial in healthy volunteers. We will be measuring both HTT mRNA and protein levels in cells within the blood, allowing us to quickly demonstrate drug activity that results in HTT lowering.
This will allow us to select a dose with the desired levels of activity. This same approach was successfully used in the risdiplam program, where proof of concept was demonstrated in the healthy volunteer study. We expect the same trajectory in the Huntington's disease program. We are very excited about this program and look forward to the results that are expected in the first half of 2021. Turning to our virology platform. The second and final stage of the FITE19 registrational trial for COVID-19 has commenced. As a reminder, PTC299 is an oral small molecule with a dual mechanism of action that demonstrates both antiviral and anti-inflammatory effects. PTC-299 inhibits SARS-CoV-2 viral replication and calms the cytokine storm. PTC-299 functions by targeting a cellular enzyme, dihydroorotate dehydrogenase, or DHODH.
The advantage of targeting the cellular enzyme instead of a viral protein is that it's less likely to elicit drug resistance. Though there have been great strides made in the development of vaccines, the lack of effective COVID-19 treatments has significantly hampered our ability to resume normal life, and therefore, the continued focus on developing treatments is key. Now let me turn to our Bio-e platform. We have initiated two registration directed trials with vatiquinone, one in mitochondrial epilepsy and one in Friedreich ataxia, and are enrolling patients in both studies. The global prevalence of mitochondrial epilepsy is estimated at 20,000 patients, and the global prevalence of FA is approximately 25,000 patients. Let me now touch on our gene therapy platform. Our initial focus is to launch our first gene therapy for patients with AADC deficiency, which is expected to occur in Europe during the second half of 2021.
The BLA submission is also on track for the second quarter of this year. As a reminder, PTC-AADC is a transformative gene therapy that has the potential to produce meaningful changes in AADC deficient patients. PTC-AADC has robust clinical data that demonstrates durability of effect for up to 10 years post-treatment, a crucial consideration in a single-dose gene therapy. Let me discuss our plans for PTC-923. As a reminder, there is an estimated global prevalence of 58,000 PKU patients, and the vast majority are not well addressed by current therapies. Therefore, we are excited about the potential of PTC-923 as a clinically differentiated therapy to address this high unmet medical need. We will start a registrational trial evaluating PTC-923 for treating PKU, called APHENITY, mid this year. Last year, we achieved many important milestones.
We anticipate an exciting year in 2021 that will continue to create substantial value for all our stakeholders. I'll now turn the call over to Matt for key updates on our clinical programs. Matt?
Thanks, Stuart. I want to build on Stu's comments on our development team's achievements in 2020. We have worked hard at PTC to navigate the many challenges of the past year and are excited to continue to deliver on planned development milestones across our multiple platforms. I would like to start with our Bio-e platform. This platform focuses on diseases of oxidative stress by targeting a special class of enzymes called oxidoreductases. Oxidoreductases are a family of enzymes that perform important electron transfer reactions and are known to have important biological functions. vatiquinone, the first compound being developed for the Bio-e platform, targets the oxidoreductase 15-lipoxygenase. 15-lipoxygenase is a key regulator of inflammation and oxidative stress pathways and has been implicated in a number of CNS diseases. As Stu mentioned, we have initiated two vatiquinone registrational trials in mitochondrial epilepsy and Friedreich ataxia.
As a reminder, vatiquinone has extensive safety data, particularly in pediatric patients, with the longest duration of exposure being over 10 years. For the first indication, mitochondrial epilepsy, previous clinical studies demonstrated vatiquinone had a positive effect on seizures and seizure-related morbidity across multiple mitochondrial disease subtypes. These results give us confidence that vatiquinone has the potential to show clinically differentiated improvement for mitochondrial epilepsy patients. The ongoing mitochondrial epilepsy trial, the MIT_E trial, is a randomized placebo-controlled study enrolling 60 children at centers worldwide. The primary endpoint of the study is a reduction in observed motor seizures, with secondary endpoints capturing other aspects of seizure activity and seizure-related morbidity. Enrollment is underway, and data are expected in the third quarter of next year. We are very excited to bring this therapy to children with mitochondrial disease.
The second vatiquinone registrational trial is in Friedreich ataxia, which is a rare, inherited, progressive neuromuscular disease that affects the nervous system and heart. In a previous Phase II trial, vatiquinone treatment demonstrated a significant improvement in disease severity compared to a matched natural history cohort over 24 months. These results support that vatiquinone can deliver a meaningful effect to Friedreich ataxia patients. The Phase III FA trial, MOVE-FA, is a 72-week randomized placebo-controlled study. The primary endpoint of the trial is changed from baseline in the Modified Friedreich's Ataxia Rating Scale, or mFARS. The key secondary endpoint is the change from baseline in activities of daily living as assessed by the FAADL scale. This endpoint strategy was developed in consultation with regulatory authorities in the U.S. and EU. We began trial enrollment in the fourth quarter of last year, and we anticipate results in 2023.
Let me turn to our PTC-518 Huntington's disease program. PTC-518 is an orally bioavailable small molecule developed from our splicing platform that was designed specifically to treat Huntington's disease. Given the need to effectively target every region of the brain, the molecule is designed to cross the blood-brain barrier and avoid efflux, a significant advantage for treating neurodegenerative disease. The phase I healthy volunteer trial is underway and includes both single ascending and multiple ascending dose regimens. As a reminder, this healthy volunteer trial is designed to not only capture key safety and pharmacology data typical of a phase I study, but to also establish proof of splicing mechanism and guide dose selection for future studies. The SAD study includes five dosing cohorts, each with six active and two placebo subjects.
The MAD study is expected to have three to five cohorts, each with six active and two placebo subjects. In the Phase I study, we are monitoring drug concentration in both the CSF and blood and will be measuring levels of HTT mRNA and protein in the cells of the blood. This ability to gain key proof of splicing mechanism data is similar to what we were able to accomplish in the risdiplam Phase I healthy volunteer studies. The data from the PTC-518 SAD and MAD studies are expected in the first half of this year. Turning to our gene therapy platform, we remain on schedule for the CHMP opinion on the PTC-AADC MAA and for the BLA submission to the FDA in the second quarter of this year. We are continuing to progress our FA gene therapy program and expect first in human dosing before year-end.
Turning to our PTC-299 FITE-19 clinical trial, we recently announced that enrollment of the stage one of the study was completed. As planned, the DSMB reviewed the interim safety data and unanimously recommended continuing with the stage 2 of the study. We have already initiated enrollment in the stage 2, and data are expected in the second half of 2021. In 2021, we look forward to advancing additional programs from our pipeline. We are on schedule to initiate the APHENITY phase III trial for PTC-923 in patients with PKU in mid-2021, with data expected by the end of 2022. As Stu mentioned, despite existing therapies, PKU remains a high unmet medical need. To summarize, we look forward to building on the successful execution of our clinical development programs in 2021 and sharing important updates when these programs available.
I'll now turn the call to Eric to provide more detail on our commercial business.
Thanks, Matt. We are very excited with the progress of our late-stage clinical pipeline, which is poised to potentially deliver multiple innovative neurology therapies that we can leverage with our global commercial footprint and existing expertise in rare diseases. As Stu highlighted, the DMD franchise had strong growth in 2020, with both Emflaza and Translarna generating significant revenue. Despite the challenges of the pandemic, we continue to see year-over-year growth of the DMD franchise. For Translarna, the only treatment for nonsense mutation DMD patients ages two and older, we saw revenues of $192 million in 2020. The growth was due to the ongoing expansion of the patient base, high compliance, recent label updates allowing broader access, and continued geographic expansion.
With the recent approval of Translarna in Russia in Q4 2020, we are excited to bring this therapy to nonsense mutation DMD patients and expand the use of Translarna globally, which is now available in over 50 countries. In Latin America, we continue to see good progress. As a reminder, last October, we entered into a purchase agreement with Brazil's Ministry of Health to supply Translarna for both new and existing patients. This order was important given the governmental administrative delays in Brazil hit exceptionally hard by the pandemic. The agreement specified two shipments. We are excited to announce that both shipments were received by Brazil's Ministry of Health last year, including the last shipment in Q4 2020 to ensure continuity of the growing base of Brazilian nonsense mutation DMD patients.
We continue to see further growth coming from new patients in the region and expect the next Brazil order in the second half of this year. Moving on to Emflaza, which is the first and only corticosteroid approved for all DMD patients ages two and older. We saw revenues of $139 million in 2020, which is a 38% year-over-year growth, driven primarily from new patient starts, a reduction in bridge and PAP free of charge programs, an increase in compliance, and lower treatment discontinuations. Importantly, we continue to see strong new prescription growth into 2021, supported by publications of Emflaza's real-world clinical benefit over prednisone, which is now driving patients to seek switching treatment from their healthcare providers. We expect the DMD global franchise growth to continue in 2021 with geographic expansion for Translarna and new patients for both Emflaza and Translarna.
Based on this, our revenue guidance for the DMD franchise for 2021 is $355 million-$375 million. Switching to Tegsedi and Waylivra. We continue discussions with CMED for pricing of Tegsedi in Brazil. During this process, we continue to provide medical education, genetic testing, and patient program support as needed. For both Tegsedi and Waylivra, we continue to engage in patient finding in Latin America with ongoing success in these programs. We also continue to engage in early access programs in the region as we await a decision on the Waylivra ANVISA filing in Brazil, which is expected in Q3 2021. Moving on to AADC. PTC-AADC is a transformative gene therapy that has the potential to produce meaningful changes in AADC patients. As a reminder, AADC deficiency is a highly morbid and fatal pediatric neurological disorder. There are currently no approved disease-modifying therapies available.
In clinical trials, PTC-AADC gene therapy demonstrated significant and durable neurological and neuromuscular improvement shown to continue for up to 10 years after treatment. PTC is currently preparing for our first gene therapy launch for patients with AADC deficiency, which is expected to occur in Europe during the second half of 2021. As part of these efforts, identification and preparation of expert pediatric neurological centers of excellence is underway throughout the U.S., Europe, and Latin America. Patient finding activities are also accelerating, with over 60 screening programs in over 20 countries to identify 300 patients by the time of launch. I continue to take pride in our global customer-facing teams as they ensure continuity of access to PTC products for rare disease patients in need.
We continue to expand our commercial expertise with the upcoming launch for AADC deficiency in neurology and build on our success in translating groundbreaking science to transform the lives of rare disease patients worldwide. Now, let me turn the call over to Emily for a financial update.
Thanks, Eric. In 2020, PTC saw strong continued revenue growth and progress across multiple platforms of our pipeline. We are executing on a number of fronts to deliver on many potentially value-creating milestones this year. The press release issued earlier this afternoon summarizes the details of our fourth quarter and year-end 2020 financial results. I will take a few minutes now to review these financial results and our 2021 guidance. Please refer to the press release for additional details. Starting with our top-line results, we reported $380.8 million in total revenue for the full year 2020, compared to $307 million for the full year 2019. This increase was driven primarily by three factors, Emflaza growth due to both new patient starts and high compliance, Translarna driven by broader access, geographic expansion, and label updates, as well as Evrysdi, driven by royalties and milestones associated with the U.S. approval and launch.
Revenue growth was due primarily to our global DMD franchise. Translarna net product revenues were $191.9 million for the year, compared to $190 million for the full year 2019. For Emflaza, we reported net product revenues of approximately $139 million for the full year 2020, which compares to $101 million from the prior year. This represents a 38% year-over-year growth. The total DMD franchise net product revenue was $331 million for 2020. Our 2021 DMD franchise revenue guidance is between $355 million and $375 million, and this guidance does not reflect any other anticipated revenue contribution. The royalty purchase agreement with RPI has allowed PTC to diversify its market risk of having a future royalty stream currently tied to one product by transforming its potential future cash flows into a $650 million cash asset.
This has created real value for PTC's financial position, as the cash is being invested to support PTC's research and development platforms and patient care initiatives. By retaining the majority interest in the future royalties due from Roche, capping the potential payout to RPI at $1.3 billion to retain future upside, and the rights to receive the remaining potential regulatory and sales milestones, PTC has retained its ability to receive consistent cash flows in future periods. We recognized $42.6 million in collaboration revenue in 2020, an increase of $26.9 million from the prior year. The increase is primarily related to three regulatory milestones that were triggered from Roche in 2020. We also recognized $4.8 million in royalty revenue in 2020 due to the FDA approval of Evrysdi in August, as we are entitled to royalties on worldwide annual net sales.
Non-GAAP R&D expenses were $438.9 million for the full year 2020, excluding $38.7 million in non-cash stock-based compensation expense, compared to $236.6 million for the full year 2019, excluding $20.8 million in non-cash stock-based compensation expense. This increase in R&D expenditures reflects costs associated with advancing the gene therapy, splicing, and Bio-e platforms, increased investment in research programs, and the advancement of the clinical pipeline. Additionally, the increase in R&D expenses includes one-time charges of $53.6 million related to the acquisition of Censa Pharmaceuticals and $41.4 million related to the MassBio agreement for commercial manufacturing of our lead gene therapy program in AADC deficiency. Non-GAAP SG&A expenses were $213.6 million for the full year 2020, excluding $31.6 million in non-cash stock-based compensation expense, compared to $181.2 million for the full year 2019, excluding $21.3 million in non-cash stock-based compensation expense.
We anticipate non-GAAP R&D and SG&A expenses for the full year 2021 to be between $725 and $755 million, excluding approximately $100 million in estimated non-cash stock-based compensation expense. Cash, cash equivalents, and marketable securities totaled $1.1 billion as of December 31st, 2020, compared to $686.6 million as of December 31st, 2019. I'll now hand the call over to the operator to start our question and answer session. Operator?
Thank you. As a reminder, to ask a question, you will need to press star one on your telephone. To withdraw your question, press the pound key. Please stand by as we compile Q&A roster. Our first question comes from Eric Joseph with JPMorgan.
Good evening. Thanks for taking the questions. Just a couple from me. First with PTC-AADC, assuming a positive CHMP decision, perhaps you could just sort of walk us through the initial launch strategy in Europe. Of the 300 patients that you're anticipating to identify at launch, what countries Excuse me. I guess, how should we be thinking about their regional distribution? What proportion are in Europe? Also, I guess, in just thinking about the risk-benefit, just thinking about the age of the juvenile patient population here as well, can you talk a little bit about the risk-benefit profile as it varies by age? Is there any challenges with stereotactic delivery in older juveniles compared to what's been sort of described to date in infants? Thanks.
Yeah. Thanks, Eric. Thanks for the call. Maybe I'll start. Then I'll turn it over to Eric, who could talk to you more about the commercial planning. I thought I'd start just by saying, just to remind everyone that the AADC treatment that we have for gene therapy actually is, we think, is really a transformative gene therapy that really can make very meaningful differences in the lives of patients with AADC deficiencies. That is, just to remind you, it's an ultra-orphan, highly morbid, and a pediatric disorder.
Much like when you think about SMA, in the severe form of these patients, they really are developmentally arrested and are unable to hold their head up, roll over, sit, stand, and move, and they could die within the first few years of life. There's really no approved therapies and nothing really for disease-modifying therapy that's available for them. I think in the clinical trials, we showed that there was really transformative changes in these patients, where patients that were developmentally arrested like this, they all improved, and we had patients that went from not being able to move to being capable of rolling and sitting, standing, and walking. We saw substantial progression as a consequence of this.
Really highly transformative results, clinical data for five years showing improvement, up to 10 years of which we've continued to follow them, and we saw this in all pediatric age groups that we measured, including older children and adolescents. We saw a very nice benefit as a consequence of it. We didn't see anything from the standpoint of surgeries, so challenges related as a consequence to the surgeries. We think it's a highly valuable product, we believe, because of we think it's one of the best packages, clinical packages, because of durability, the ability to see a biomarker. We can see changes in dopamine levels that we continue to see. I think that is really powerful. What we've been working on in terms of the launch is obviously getting ready and identifying patients, getting centers of excellence.
Eric, why don't you talk through a bit about the potential launch and what we're doing for launching that.
Sure. Eric, thanks for the question there. First of all, most importantly, we have a number of key activities that Stu outlined in preparing that regional rollout. Those included an increase in disease educational programs that were driving genetic testing. We now have 60 screening programs now in over 20 countries, the vast majority in Europe as well as LATAM, in other central and eastern Europe, and other places where we know there will be access and reimbursement for gene therapy products and ultra-rare disease products. We've done a number of things in terms of global country and specific webinars and virtual symposiums. The level of the preparation in the marketplace has been extremely, I would say, aggressive in terms of how and where we're preparing in terms of our patient-finding activities and education.
As Stu mentioned, one of the key areas as well is making sure that those sites, those centers of excellence, are prepared to not only treat the patients, but also follow them, and we've been doing that in many of the key areas. The regional rollout is going to be specific, as you would expect, with first commercial launches in Germany. We will immediately roll out a number of early access programs and taking full advantage of the mechanisms by which European, as well as Latin American, and other areas of the world will have once the final opinion and approval comes from Europe. We would anticipate Germany, of course, and then early access programs in countries like France, Italy, Spain, Northern Europe, and Scandinavia, and others. We'll take full advantage of early access programs in Latin America as well based on the European approval.
We're very confident at this point, and most importantly, I would say I'm excited that we've seen an acceleration of patient finding since we've implemented a large number of these programs, and we're anticipating to have 300 patients addressable at the time of launch. These will be in countries where we have access and reimbursement to gene therapy.
Great. That's helpful. I'll hop back in queue, thanks.
Thank you. Our next question comes from Gina Wang with Barclays. Your line is now open.
Hi. How are you guys doing? This is David on for Gena. Thank you for taking my questions. I have a few questions around for the Huntington's disease program, PTC-518. First question is just around the dose. Can you discuss what dose levels you're testing in healthy volunteers, and what's the half-life in dosing frequency?
Yeah. Thanks for that. Just to remind everybody that we have PTC-518, and the role of this is to cause splicing to induce an intron into the RNA so that there's a premature stop codon so that it would get prematurely stopped, so you don't make the protein, and the RNA is rapidly degraded. We designed this. It's an orally bioavailable small molecule that crosses the blood-brain barrier. It reaches all regions of the brain, and I think most importantly is as well, one of the major characteristics is it's not an efflux out of the brain. Just for everyone, the reason that that is so critical of a property is that many things can pass the blood-brain barrier, but the brain protects itself and moves out many things that they think are potentially toxic to it. That efflux, so there's many pumps that do that.
We've made sure that PTC-518 can pass the blood-brain and stay within that. That's a critical property, so that therefore you know the level of what you see in the blood is what you see in the cells within the whole brain. What we've shown in animal models that HTT reduction was clearly titrated based on its exposure level, so that the degree of HTT lowering can be tightly controlled. I think that really distinguishes 518 because it achieves uniform exposure and that the huntingtin lower is in all cells of the tissues analyzed. In all cells within the brain. Therefore, it shows a near one-to-one ratio between the PTC-518 levels that are observed in brain cells, and again, all parts of the brain, and what we see in the cells of the blood. That's because it passes the blood-brain barrier.
We see the same levels in blood and brain. We have a very good measurement to show that's indeed the case. When we think about the trials that we did, it was based on obviously working on what the levels that we saw in the mouse, rat, and non-human primate. Based on our understanding of the experience that we've seen in splicing, that what we did is obviously screen the molecules for specificity and biodistribution. We selected really based the molecule on that. We moved into the clinic, and the levels that we did is based on the results that we saw in the animal models and the predictions of what we would see in people is how we chose the levels of drug that we would begin with.
I think that's how we did this, where we identified what we think is the right dose, start low, and continue to move up to be within regions that we would predict to see HTT lowering, and choose it based on that. That's how we chose the doses to go into humans and look for what levels were changed. David, did that help you?
Yes, that's very helpful. Just to follow up on that, Stuart, I guess the question is, what level of HTT reduction are you looking for in healthy volunteers to enable you to select a dose for the Huntington's disease patients in your next trial?
Yeah. The way we think about this is we're going to do a single ascending dose and multiple ascending dose. We'll be able to know what exposure gives us a targeted number to be able to move forward on. That's the beauty. It's really great that we're capable of identifying, knowing what the dose is, characterizing the exposure, and then measuring the level of changes in the RNA over time, and both doing that in a single ascending dose and a multiple ascending dose. We get a very good characteristic of what's the dose that gives a steady state level of reduction. I think that's one of the major advantages that we have.
We have a small molecule to where we know the exposure in the blood and in the brain, and we've shown that what you see in the blood is equal to what you see in the brain. We've seen that across all of the animals, including non-human primates. We're very comfortable knowing that. As what we're doing is, as we measure in people, we'll be measuring the levels of drug and then the reduction of HTT RNA in the blood. We'll have a very good dose response level that we can titrate, that we can define what dose we want. We're probably starting to think about looking at probably a 50% reduction within that. We'll look for that in terms of clinical benefit.
I think there's results out there when you look at clinical data in terms of some snips that you see in patients where if you lower it 50%, they have a substantial improvement in terms of nine years without disease demonstration. There's pretty good clinical data that suggests that a 50% will have a benefit. We also have the capability that as we learn more, should we go lower or not, we could titrate that. I think we can go lower if we think it's correct to do so. We're starting to think to look at the reduction at looking at a 50% level. We'll be able to identify if we can go lower and even further, what would be the dose that We'll be able to define that dose. Did that help you, David?
Yes, absolutely. Very helpful. Thank you, Stu. I'm going to hop back on the queue.
Thank you.
Thank you. Our next question comes from Alethia Young with Cantor Fitzgerald. Your line is now open.
Hey, guys. This is Lee Ahn for Alethia . Thanks for the update. Wanted to follow up on Huntington's program, the phase I study. Can you just give us a quick update on where you are in the trial now, and have you moved to the next portion of the study?
You want to know. Right. As we've talked about in terms of the trial, we've obviously been doing the trial. We're looking at both the SAD, the single ascending dose, and then going into the multiple ascending dose trial. We've obviously been doing both of those. We said that we'd have it by within the first half of the year, so it's underway, and that we've been moving forward. Just to remind everyone, it's designed to capture, obviously, the safety and pharmacological data. What I said is in the single ascending dose study includes up to six cohorts, each with six patients who have active, six with active drug, 2 with placebo. The MAD study is expected, where we'll have, it could be up to iv cohorts, again, with six subjects that could have active drug, while two would have placebo.
things have been dosed and moving forward without any issues. We're actually pretty excited about it. just to remind everyone that in the phase I study, what we're monitoring is obviously the drug concentration in the blood, the levels of the HTT messenger RNA, and ultimately protein in blood cells. We will have one cohort that will measure PTC-518 blood levels as well as blood levels in the CSF. as a reminder, these are healthy volunteer studies, and the subjects don't have Huntington's disease. we don't expect to have, in a sense, the proteins of interest that people have talked about in the CSF. The purpose is to do exploratory PK measurement studies in the CSF and to be able to correlate that with the blood.
At the end of the day, I think what you'll see is that we're going to show proof of splicing mechanism, determine the pharmacokinetic characteristics of PTC-518, be able to define the dose and be able to say what dose exposure levels that allow us to target the level of HTT reduction that we're shooting for, and to know if we wanted to go higher, what dose we would need. I think these results are going to be really analogous to what we were able to accomplish when we did the SMA trial, the risdiplam, in the healthy volunteer subjects, which worked out really beautifully. We're on track to share the data for PTC-518 for both the single ascending dose and the multiple ascending dose, and expect to have that within the first half of this year.
Of course, we're planning to have a deep dive that we will talk further with you so you can see much of the data of the science and the data that we have in the preclinical studies ahead of that.
Okay, got it.
I'm excited.
Okay. Thanks for the color. My second question is just on AADC. Just wondering, for you to file BLA in second quarter, what are the remaining steps and, in terms of the cannula study, can you just give us an update on where you are right now?
Sure. I just want to remind you, obviously, we've already talked a bit about it. We think this is a transformative gene therapy, and that we've seen what it does for patients already both in the durability and the 10-year post-treatment follow-up, we know about this quite well. Just to remind everyone in the EU, the MAA is moving forward. We expect a CHMP opinion in the second quarter of this year. Matt, you want to talk a little bit about where we are right now?
Yeah, sure.
For the-
Thanks, Stuart. As we've talked about before, one of the key gating factors to the submission of the BLA was conducting additional surgeries with the cannula we intend to use commercially. Now that cannula is CE marked in Europe for the delivery of gene therapy, which is why, obviously, that wasn't an issue with the MAA submission. It's been used in many gene therapies before, and it was just a matter of us getting experience demonstrating the safety of the cannula delivering our specific gene therapy to our intended patient population. We've actually completed two of those surgeries already. The procedures went well. There were no complications from the surgery, and recovery is on schedule.
Again, just as a reminder, these surgeries are to evaluate the safety of the cannula, given that we already have extensive safety and efficacy data that Stu mentioned accumulated over a number of years with the gene therapy product. We plan to conduct one additional surgery. Once those data are collected, we'll align with the FDA and move forward with the BLA submission.
Okay, great. Thank you.
Thank you. Our next question comes from Robyn Karnauskas with Truist Securities. Your line is now open.
Hi. Thanks, everyone. I'll take another question. This is Min on for Robyn. Hey, Stu, back on Huntington. When you present the data, are we going to get data from all the dose cohorts, or will it be whatever the trial has enrolled and that's what you can present, so maybe possibly just a few cohorts from the SAD study? I guess, just the first question around there.
I think what we've said in the past, we're going to have both. Obviously, what we have, we're on track to share the data for both the SAD and MAD studies within the first half of the year. What we were planning to have is a deep dive ahead of that. If we get to that point, when we get to that point, we would have, by the end of the year, both the SAD and the MAD data on that. We'll share the dose levels with you at that time.
I see. I guess last question. Do you have an idea or a sense where the therapeutic dose may be and where that may fall in? Sorry, one more question as well. Yeah, I guess that one first. Any idea for where the therapeutic dose may fall within the MAD or SAD study?
Yeah, I think, again, based on the data and based on what we're doing, we had a pretty good indication of what we thought that would be in terms of defining the dose. It's been moving forward. I think we'll be in pretty good shape where we'll be able to have a dose response curve in terms of measuring both the splicing and where the dose is. We'll be able to give you a pretty accurate prediction of what dose gives a particular exposure that leads to, in the sense of if we want to say a 30% or 50% or 70% reduction of HTT. I think we'll be in a pretty good position to be able to tell you that and to be able to move forward based on that.
Between the SAD and MAD, we're going to have a very good picture in terms of both safety as well as the dose, the exposure, and the level of reduction at the end of the first half of this year.
Got it. That's helpful. Then, sorry, last question. Regarding your comments around titration, I guess, will you have enough sense to know whether there's more safety risks that would come on with going with a greater than 50% reduction? I guess, how do you know there won't be any longer-term safety risks that this trial may not be able to measure should you push beyond 50%?
Well, I think what we're shooting for right now is around that. The way I think about this is that there's plenty of data in terms of people who have either one allele or as a consequence of that, there are no safety findings that we see of that. I don't think there's going to be a risk in terms of that. In terms of it going lower, there isn't really a lot of data beyond that. We clearly know that it's important during early embryonic development, there isn't a lot of data that says after that, how low can you go?
I could tell you from, as a geneticist point of view, when you look at the level of most things that when you say how much do you normally need, if you have some of the factors, if you have 10% of them, you're pretty much normal. The question is at what level, how far can you go where you don't see any effect? I think there are some ways where we're thinking of being able to test that in animal models that have human HTT, where we can actually use titration of our compound to reduce it to the level of interest, and we might be able to do some animal studies to be able to define that.
The reason we would be able to do that with 518 with a humanized HTT mouse is that we're really capable of achieving uniform exposures that continually reduce levels. For instance, if we want 50% down, we could see 50% down in virtually every tissue in the body and certainly within the brain. That would include the cortex, striatum, cerebellum, and every other tissue within the brain. We know that from the work we've done, we've been working with the groups to be able to build that mouse. If we're able to do that, we can actually at least get some data that would suggest can we go lower or not. We're obviously shooting for 50% right now, and I think that's a target that I think we'll certainly be able to achieve.
Great. Thank you so much, Stuart.
Thank you. Our next question comes from Joel Beatty with Citi. Your line is now open.
Hi, thanks for taking the questions. First, are there ways to assess using biomarkers in the clinical data for PTC-518 if you're generating uniform knockdown throughout the body, or how important is it to assess that clinically?
I think clinically, the biomarker that we're choosing is looking at a reduction of Huntingtin levels within the blood, which we've done extensive analysis in both mouse, rat, and non-human primates. We've been able to show that the reduction there goes along with the reduction that we've seen in other tissue types. That's going to be a pretty good biomarker. Obviously, just so everyone knows that HTT is an intracellular protein, right? When we do the analysis, we're looking directly within the cells, and the blood cells are a good marker for the level of reduction that we've seen there, we've seen in all other tissues when we looked in mouse models. We know we'll be able to do that direct correlation based on the blood and what we see, the reduction in protein and RNA levels. Does that help you, Joel?
Yeah. Great. Maybe switching gears for Evrysdi, could you remind us what additional potential milestones remain?
Any what?
Milestone payments.
Oh, milestone. Oh, yeah, sure. Emily thought she wasn't going to talk today, but I'll pass it to Emily.
Thank you so much. Thanks, Joel, for the question. I appreciate it. You are correct. We have retained all of our milestones for Evrysdi after the royalty monetization, and we do have about $300 million in milestones remaining, largely sales-based and regulatory. The most real-term, near-term milestones upcoming are about $55 million in total, potentially for 2021. $20 million would be in the first commercial sale in the EU, $10 million for the commercial sale in Japan, and then a potential $25 million sales-based milestone upon Roche hitting $500 million in sales.
Great. Thank you.
Thank you.
Thank you. Our next question comes from the line of Brian Abrahams with RBC Capital Markets. Your line is open.
Hi there. Thanks so much for taking my questions. A few on the Huntington's program. Given what's known or not known about HTT protein turnover in the context of the timeframe of the study, I was wondering how you guys are thinking about looking at mRNA versus protein changes. What's going to guide your decision as to whether to move into those additional two MAD cohorts versus just running three? As you sort of look past this study, how do you think about how the blood-brain barrier properties in Huntington's patients might impact the overall plasma to CSF ratio, and how you might be thinking about dose adjusting when you go into patients? Thanks.
Thanks, Brian. When we think about the drug concentration within blood and the levels that we'll see and the reduction that we'll see in the HTT mRNA and protein within the cells, we're also taking the CSF that what we anticipate is the level that we see in blood and the CSF levels would be equal, and that's similar to what we've done in non-human primates. We think that's going to be very telling to us. We'll be able to just confirm the ratio that we've seen previously. I think that's good. At least we'll have a potential to see that. That's what's going to help us in terms of defining what the exposure is and what that would be.
That's the way one could look at that in terms of to make sure that you see the correct level, and that could be done elsewhere as well. I think that's a really important way for us to be able to be sure in terms of the PK and then equating the PK to reduction of RNA and protein. You bring up actually a pretty good point in terms of when you think of RNA and protein, a lot depends on the half-life of the protein and the RNA. We'll be able to get a pretty good handle on that in the multiple ascending dose and looking to what happens with both the RNA level. What we're obviously trying to do is to, in the multiple ascending doses, to reach steady state and then to be able to monitor the HD RNA and proteins.
We know that in some ways, HD has a leaky blood-brain barrier. I think that's what you're referring to. I think that's something that we'll have to take into consideration as we monitor the PK of them in patients.
Got it. Thanks so much.
Yeah. Great. I would just make the other point is that we don't have issues with efflux, so that shouldn't cause any sort of issues for us.
Thanks, Stuart.
Yeah.
Thank you. Our next question comes from the line of Vincent Chen with Bernstein. Your line is open.
Thank you very much for taking the questions, and congrats on all progress. A couple more for you on Huntington's disease. The first is, in your preclinical RNA-Seq experiments, how many other RNA splicing events were identified that were modulated by PTC-518? Which ones and how much were they modulated? The second question is, thinking about the ongoing study, how many doses do the MAD healthy volunteers get, and what's your sense for how much run-out is needed to assess the risk of off-target toxicities?
Yeah. Could you just say the first question one more time?
Sure. In the preclinical RNA-Seq experiments that you did, how many other RNA splicing events were identified that were modulated by PTC-518? Which ones were they, how much were they modulated?
Yeah. Maybe it's worth talking a little bit about how we optimize the compounds in general. I think that's actually an important point. The way we do the optimization and looking first in selectivity. Selectivity is a pretty important point for us. We begin, when we do the high-throughput screening with the small molecules, we screen them in the Huntington, in fibroblasts for them. We optimize those molecules for efficacy and potency, which also includes selectivity and pharmaceutical properties. We look very much so to select for molecules that may start out to be less selective but build in as much selectivity as we can.
At the end of the day, we've identified these molecules and obviously spent a lot of time to show that they really do have favorable properties, a safety window that we've seen in the animal models, strong potency, and a pretty selective HTT that alters the select. We learn a lot about the mechanism and the specificity of that. Just contrast that with, for instance, the competitor molecule that was for one molecule and then the others. I can tell you that, for instance, the specificity of our molecule versus SMA is substantially different than the selectivity that's built in. It's far more selective for Huntington's disease than SMA. That was a consequence of really spending the time to build in the selectivity within the molecule to be able to do that.
That's really a consequence of both understanding the specific sequences, the differences between them, understanding how the U1 there works, so where we can build in the selectivity in the RNA sequence. I think, in the deep dive that's coming up, we'll spend some more time on the question that you're asking, and probably we'll be discussing that more during the deep dive time.
I see. That's very helpful. I guess the second question was just simply, how many doses do the healthy volunteers in the multiple ascending dose study get? What's your sense for how much run-out you would need to assess the risk of off-target toxicities? If I think about splicing toxicities, how quickly would you expect those to show up, and how much run-out do you need?
Obviously for the multiple ascending dose, that's a 14-day dosing period followed by 28 days of observation. That's what we do in terms of defining the dose. Jim, you got to remember also that we're also completing the long-term safety toxicity studies that will help us. I don't want the folks that are listening to think there's something different about splicing toxicities versus any other toxicity, whether it's off-targets for splicing or some other function. The key is, we spent a lot of time looking for selectivity and specificity, we look at toxicity. It could be for off-target, it could be for whatever the reason is. That's why we do those studies to look for the off-target toxicities.
I don't think you can make the argument there's something unique about these toxicities versus any other drug toxicity. That's why we do these experiments, the safety toxicology experiments, to be able to identify the toxicities, understand the risks, to be able to move forward. I think this is a very standard and stepwise approach and the targeting of splicing is no different than doing kinases, phosphatases, transcription factors, other things that other people do. I wouldn't try and make it to something special here versus other things.
I see.
Does that help answer?
Yes. Very helpful. Thank you for the color, and congrats again on all the progress.
Thanks a lot. I appreciate it.
Thank you. Our next question comes from the line of Raju Prasad with William Blair. Your line is open.
Hi there. This is Sammy on for Raju. Thanks for taking my question. I was curious if there's any update on your Angelman syndrome gene therapy. I believe previously you said that the Angelman program and Friedreich ataxia gene therapy programs were delayed because of COVID. I'm just wondering if that's still the case or if there's any other behind-the-scenes things going on. Just secondly, how are you thinking. In terms of the FDA's recent guidance for gene therapies for CNS diseases, how are you thinking about that guidance influencing those trial designs? Do you plan on using the same endpoints that will be used for the vatiquinone trial for Friedreich ataxia? Thank you.
Sure. Yeah, you're right. We've talked about this in the past. I do think one of the things that happened during the pandemic is that there are really COVID-related delays that push programs out. We're really working hard, and we're progressing on the FA gene therapy program, and this affects the human dosing before year-end. Similarly, we're pushing hard on the Angelman syndrome. I don't think we actually gave particular timelines on that, but we're moving forward on that, and we'll probably talk more about that as we get closer. We'll talk through over time as we do this. Maybe, Matt, do you want to add any other color to this?
Yeah, sure. Thanks for the question, Sammy. On the, I think you had some questions about our Friedreich ataxia gene therapy program and the recent FDA guidance. Maybe I'll just tackle the guidance first. Quite frankly, not a lot of surprises there. It's sort of read very standard FDA guidance for the development of therapies for neurodegenerative diseases. I think that they're obviously communicating the need to have rigorous control groups, even for gene therapy, and they obviously also wanted to spend a lot of time talking about the importance of device and device comparability and making sure that your product works well with the device and having consistency throughout the development program.
All things that's, one, not really different from general guidance around CNS drug development, and then second, with specifics to gene therapy, it's all things that we've learned and become quite familiar with along the way with our experience doing gene therapy development. I think the other thing that we obviously have in our programs, which is very important, is we're taking a targeted CNS approach, and a few benefits to that. One, is that we're able to deliver the gene therapy product to the specific anatomic areas where the pathology for the disease really results from. For example, in our AADC program, we're delivering our gene therapy products to the putamen, which is really the key area for dopaminergic neuron function. In Friedreich ataxia, we're delivering our gene therapy precisely to the dentate nucleus of the cerebellum.
Obviously, the cerebellum is a key component of ataxia, which is obviously a key component of the Friedreich ataxia disease. By doing targeted gene therapy, not only does it allow us to have lower doses, which obviously lowers risk and exposure to the vectors, obviously it also lowers our manufacturing burden. Switching now to your questions regarding the Friedreich ataxia gene therapy program, I think we're actually incredibly excited to be able to be developing two therapies for Friedreich ataxia. We have our oral small molecule, the vatiquinone which has broad distribution through CNS and other organ systems, as well as our gene therapy program, which is delivering to ataxia targeted dentate nucleus. I think these are really complimentary approaches.
I think while Friedreich ataxia has a great deal of pathology resulting from the dentate nucleus, it's also a whole brain disease and a whole body disease, as the heart, for example, is a significant source of disease morbidity and mortality risk in Friedreich ataxia. We view our approaches as incredibly complimentary, and we're excited to be able to offer both of these therapies in a complimentary way. In terms of the development itself and endpoint selection, I think one of the great advantages of looking at Friedreich ataxia is the well-established natural history. The community is well aggregated, has a very rigorous natural history, particularly around the disease rating scale that was developed by the community, as well as the activity of the daily living scale.
As we think about gene therapy, the gene therapy development program, obviously, we're going to be looking to those where there's significant natural history, so we can really appreciate the impact of a gene therapy on the long-term trajectory of the disease. Obviously in our first-in-human studies, we'll also be exploring other endpoints that are both biomarkers, biochemical mediators, imaging modalities, and also other clinical endpoints that we know to be important to the disease and that would certainly capture the ability of a gene therapy to have a significant effect on disease trajectory.
Great. Thanks for the clarity.
Thank you. Our next question comes from the line of Danielle Brill with Raymond James. Your line is open.
Oh, hi, guys. This is Danielle Gatullon, on for Danielle. Thank you for taking the question. I had a question on slide 1-8. How uniform was the HTT reduction in animal models for a given dose? Is the reduction affected by the length of nucleotide repeats, or any other factor that you could identify?
Sure. Thanks for the question. Actually, that's the beauty, the fact that one thing that I think really distinguishes PTC-518 is that it really achieves uniform exposure. When we look, for instance, for HTT lowering in these cells and tissues analyzed, if we see a 50% reduction in blood, we saw in animal models a 50% reduction within the brain, and in particular, a 50% reduction in the cortex, striatum, cerebellum. That's amazingly uniform. That's good, right? That's a demonstration of really, as I said, we had a one-to-one ratio in terms of what we saw lowering in the blood within the brain. That's true within all tissues as well.
That actually turns out to be, I think, very important because, I know people have talked about Huntington in terms of striatum and cortex, but I think when you look, you see that Huntington's disease is a whole-brain disease. It's very important to emphasize that based on our preclinical results, that we'll see a dose-dependent reduction of the HTT mRNA and protein in all cells within the brain. That includes the striatum, cortex, and cerebellum. It gets everywhere. Again, I think the really critical point is that, and this was true for SMA, I think it's going to be true here as well, is that we have the ability to measure the reduction based on exposure since we can determine that within the blood, and that that same ratio that we see in the blood occurs in other tissues as well.
Right. Thank you. That's helpful. In terms of the HTT gene itself and the variation in the nucleotide repeat within HTT, does that have any effect on HTT reduction-
No. No, it doesn't
For a given dose?
Yeah, that's in a different location. The way this is working is it's within an intron of the pseudoexons, the piece of the RNA within the intron that normally doesn't get spliced in but because of the molecule, it does get spliced into the mRNA, and that leads to, again, premature termination and rapid degradation of the RNA. That's not in the CAG repeat, and it doesn't matter as a consequence of that. There is no issue with that, and that's the beauty of this. This will be good for all patient types. Obviously, in the long run, what you can see, because it's orally bioavailable, that the advantage here is that we would anticipate over time that it would be for patients.
We'll be looking initially at those that are manifesting symptoms, but you can imagine over time, we would be able to do that for patients before they have symptoms because it's so easy to take. I think that's important. At the end of the day, the answer to your question is that pseudoexon incorporation occurs regardless of the degree of HTT expansion.
Okay. Thank you very much.
Sure.
Thank you. Our next question comes from the line of Joseph Thome with Cowen and Company. Your line is open.
Hi there. Thank you for taking my question. Just one on the Bio-e platform. Curious with PTC-857, after you see these healthy volunteer data, maybe what are you looking for, and when can we see that advancement into a potential GBA Parkinson's study? As 857 and vertiquinone both target 15LO, can you tell us what are the differences that make 857 maybe more amenable to the Parkinson's disease indication? Thanks.
Yeah, sure. I think part of what we're doing, it does have different properties in terms of that probably would be an advantage for something that we're taking long-term for a lot of people. Again, it's an orally bioavailable molecule. For indications like GBA, what you said, Parkinson's, obviously it's in a different number of patient population versus vatiquinone and where that's going now. We thought it'd be best to have a different molecule with perhaps different properties. That's sort of in the sense it has a different value for the size of the patient population. We thought it was important to have another molecule. Matt, you want to talk a little bit of where we're at in terms of the trial and what we're thinking about?
Joseph, thanks for the question. As you mentioned, we've been studying 857 in the phase I study, which was a single ascending and multiple ascending dose studies, fairly standard healthy volunteer studies, where the focus has really been safety and pharmacology. We completed the dosing in those studies, and we're in the process of doing the analysis. Really what we're looking for here is understanding the pharmacology, making sure that the molecule's behaving in humans as we've been able to model it in the preclinical studies. Also identifying the dose level that gets us the exposures that we saw to be efficacious in all the preclinical work that we've done.
We want to walk away from this study with the dose level that we know is safe, that has predictable pharmacology, and would be consistent with delivering the exposure that's necessary to achieve the preclinical effects we've seen. As a bit of background. AP-857 targets 15-lipoxygenase, which is a key governor of a number of pathways which independently are known to be important in Parkinson's disease pathology, such as microglial activation, alpha-synuclein oxidation, aggregation, glutathione depletion, and lipid-based oxidative stress. What we're able to do by targeting 15LO is affect simultaneously all four of those pathways. We do have an extensive amount of preclinical work demonstrating effect.
Again, what we want to see in the phase I study is identifying that dose level that brings us that exposure that matches up with what we've seen in the preclinical studies, and then we'd be in a position to move forward with the next stage of development.
Yeah. Thanks, Matt. I think the really important point here as well is that oxidative stress, really extra electrons that cause oxidative stress, there's multiple diseases that we can go in, as Matt has said. Having another molecule to go into other indications with different properties, we think is going to be valuable.
Great. Thank you so much.
Thank you. I am not showing any further questions. I will now turn the call over to Stuart Peltz for closing remarks.
Awesome. Well, thanks a lot for joining us today. As many of you may be aware, the Rare Disease Day is upcoming this Sunday, and this is oftentimes where we take stock of where we've been and where we're going as a company. It serves as an important reminder to us that we do this work ultimately to benefit the patients. I hope that you see just how strong our execution has been in 2020 and how this sets up for us for many value-creating milestones in 2021. We're excited about the programs that we have, and we look forward to, in particular, I know everyone's been interested, you can see by the questions, the interest in the phase I Huntington's disease readout, and we look forward to sharing that with you within the second quarter. Again, thank you for joining. Have a good evening.
Ladies and gentlemen, this concludes today's conference call. Thank you for your participation. You may now disconnect.