by Lauren Hoffer
I have been meaning to write a blog explaining what I understand in regards to the progress being made to ensure that all people with Angelman syndrome (AS), including those with the UPD/ICD/mosaic genotypes, will have access to clinical trials for various disease modifying therapies, and what FAST is doing to de-risk this potential, knowing that paternal activation may be associated with more unknowns in this genotype that others. The recent FAST grant to Dr. Keung at NC State spurred me to get this out to our community and my fellow UPD/ICD families as many of us struggle as we wait to know when will the time be ripe for our children with the more unique genotypes.
For those that don’t know me, my name is Lauren Hoffer. My husband, Greg Willis, and I have two boys, including a 4-year-old, Ben, who lives with Angelman syndrome, UPD. Ben has two very long runs of homozygosity on the 15th chromosome (30 and 17 MB). I don’t know about any of you, but in our diagnosis appointment, our doctor looked at my husband and me and said, “You two are not blood related, right?” (Another way that you can have each of the chromosome pair matching.) Um, no.
I am also on the Board of Directors for the Foundation for Angelman Syndrome Therapeutics (FAST), and have been for over a year. I have taken part of this blog from the summary written by Dr. Jim Daley, a scientist and AS brother who is also a FAST scientific advisor, and Dr. Allyson Berent, FAST’s Chief Science Officer, with Allyson also serving as editor to make sure I get the scientific nuances accurate. It is not lost on us that UPD, ICD, and the mosaic genotypes have not been invited to any of the ASO clinical trials yet. Most of us understand why, but for anyone who is new: in a typically developing person, UBE3A is only expressed from the mom’s copy (maternal) of chromosome 15, as the father’s (paternal) copy is silent in neurons based off of a phenomenon known as “imprinting.” This is a normal phenomenon in all of us, and only occurs in neurons (brain cells). Individuals with AS do not have a functional copy of UBE3A being read from the maternal copy for various reasons.
Antisense oligonucleotides (ASOs), which are currently in clinical trials, as well as several other strategies in the works: CRISPR based approaches, shRNA, miRNA, etc. (collectively “paternal activation strategies”), are attempting activation of the normally silenced paternal copy of UBE3A. By stopping the imprinting, or inhibiting the antisense transcript that causes the imprinting, UBE3A can be expressed. This is often referred to as “stopping the stop.” Individuals with AS – UPD have two paternal copies – with two copies of UBE3A silenced. Individuals with AS – ICD have a maternal and paternal copy, but the maternal side is behaving like a paternal and is also silenced. An Angelman–mosaic diagnosis likely means you have some cells that are functioning normally with a maternal copy that is appropriately being read, but other neurons do not have a maternal side expressing UBE3A. This is most commonly due to ICD of the maternal copy on some percentage of neurons. This percentage is variable, as is the clinical presentation in this population.
If paternal activation works, UPD, ICD, mosaic genotypes would potentially have two copies of UBE3A unsilenced. This could potentially cause more UBE3A to be expressed than a typically developing person has. Overexpression of UBE3A has been linked to autism, a condition known as Dup15q Syndrome. People with Dup15 and Prader Willi UPD likely have overexpression of UBE3A, and those individuals are not neuro typical. The issues related to Dup15 and Prader Willi have been linked to other genes or proteins in the region that are also over- or under-expressed. A research abstract was presented at the FAST Translational Research Symposium in December 2020 showing that in cellular and animal models if these neurons go to a 200% expression, which is what could happen if we get full biallelic expression in individuals with UPD/ICD or mosaic AS, it could be safe. Further investigation is still needed, but so far things are looking promising.
There is also the possibility that individuals could get close to typical UBE3A expression, as Dr. Art Beaudet pointed out in his FAST Science Summit talk a few years ago. It is not clear whether we can get all neurons to express UBE3A and get full brain coverage with any one specific medicine. One thing we know about UBE3A – a little bit goes a long way. People with AS mosaicism – with just 5% of what they should have of UBE3A – generally have less developmental delay and seizures than kids without that 5%. We are hoping for much more for everyone with AS, but having double expression could allow individuals with UPD/ICD to get closer to typical UBE3A expression, even if some cells are overexpressing the protein and others are not expressing enough.
A majority of scientists think that paternal activation will work well for UPD/ICD/mosaic individuals, and most think it is a question of dosing to get enough, without overexpression. We may need to find a sweet spot. There are two possible scenarios for how dosing could work in paternal activation in UPD/ICD/mosaic. Those with UPD/ICD or mosaicism may need half the dose of individuals with a deletion because they have twice as many paternal alleles being unsilenced; or they may need double the dose to get enough drug into each neuron to address both alleles being that the amount of medicine entering each neuron would be split between the two copies and there could be a threshold that is needed to turn on that paternal gene. The current trials have focused on individuals with deletion or mutation (null) for many reasons – the most important one being that they are null, have only one silent copy, and they only need one working copy to get back to a more typical cell. This has a lot less guessing that would need to be done. In addition, with the current trials include the deletion and mutation genotypes, which is over 90% of the AS population, and the deletion and mutation genotypes are a more homogenous group individually so that endpoints and changes can be more easily compared. The added risk of overexpression in UPD/ICD/mosaic is the main reason to not include these individuals in the first round of safety, tolerability and exploratory endpoint studies. The current trials, with participants who are deletion and mutation positive will help doctors and scientists understand the dosing better for this 90%. The ASOs wear off. This has pros and cons, but should we have an issue – there is an upside to the medicine wearing off, and from the current trials, doctors are getting an understanding of how long that takes. Likely, with UPD/ICD/mosaic, the dosing will start very slowly.
As with any experimental therapeutic, there is risk; however, rest assured that FAST is ensuring that the best researchers are engaged in order to de-risk, as much as possible, any potential therapeutic for UPD/ICD and mosaic genotypes, so that this is a reality for ALL.
A key bottleneck in de-risking paternal activation in UPD/ICD/mosaic individuals has been the lack of models to study these genotypes.
Do we need a UPD/ICD/mosaic mouse? For anyone following closely – the AS mice are mostly small gene deletion or mutation mice – they have solely UBE3A deleted; although FAST is now funding the creation of a large deletion mouse model to actually mimic the human disorder. First, it is really hard to make a UPD/ICD mouse that can live. We have discussed this with numerous experts who create rodent models, including those who were mostly responsible for the AS models, and while the mice are great at showing a proof of concept there was very little interest by pharma partners or academia in feeling that this would accelerate drug development for these genotypes, while costing a tremendous amount of money. There actually was a UPD mouse at one time, but due to the difficulty in maintain it, it does not exist currently. Therefore, we decided not to spend our limited resources on another mouse, but we still felt there needed to be an elegant way to understand the differences that exist for UPD and ICD individuals and help to further de-risk therapeutic options.
FAST is funding research that focuses on the creation of novel cell lines in a really exciting way. FAST funded a robust stem cell project with Dr. Yong-Hui Jiang at Yale University to develop numerous human stem cell lines, which will be differentiated into neurons, for all genotypes, including UPD and ICD. Then, these lines will be transported to Dr. Keung’s lab at North Carolina State University to create these amazing landing pads to explore what happens with gene activation and expression including UBE3A and outside of UBE3A, which can be a problem with UPD/ICD genotypes. From there Dr. Keung is developing organoids (or mini-brains) to really evaluate the neuron connections and see what different treatments do to these connections. We are so grateful to so many of you who have graciously volunteered to donate blood to make these lines!
Let’s talk about this “landing pad.” What is this and how will it help those with UPD/ICD/mosaicism? This is an explanation from Drs. Daley and Berent:
“This newly funded FAST grant to Dr. Albert Keung of North Carolina State University will address this question of UPD/ICD/mosaic models in a series of elegant cellular models. Dr. Keung’s lab will epigenetically modify human stem cell lines to mimic the UPD or ICD epigenotype of chromosome 15, in which methylation of the DNA reflects paternal patterns on both chromosomes. These cell lines will be a valuable, globally available tool for evaluating therapeutics for UBE3A activation. …
[T]his project aims to provide the Angelman syndrome research community a set of cell lines that can be used to efficiently model the biology of ICD and UPD, as well as organoids that model the mosaic genotype, and understand the impact of gene overexpression in these different regions. This work will create valuable resources which will be shared with the AS research community and will help to drive AS research forward with the ultimate goal of accelerating drug development in Angelman syndrome for each and every genotype.”
As a UPD parent, I am incredibly excited and grateful that FAST has funded this work. I won’t take full credit for pushing hard on UPD/ICD genotype evaluation as this is within FAST mission through and through, but the UPD/ICD/mosaic voice is loud and clear at the table! This work may also be able to help us understand things like why individuals with UPD/ICD are more prone to food seeking. Thank you to the FAST team, particularly Allyson, for seeking this out and for brainstorming what else we could do for individuals with UPD/ICD/mosaic Angelman syndrome to ensure that not a single child or adult with AS is left behind. This is what FAST lives for every single day.
There are several strategies in the works that are adding a working copy of UBE3A, not through paternal activation. These strategies should work equally for ICD/UPD/mosaic as it does for point mutation and deletion. One of these FAST-funded strategies – through Hemopoietic Stem Cells Gene Therapy (or HSC-GT), was just published in a peer reviewed journal by both Dr. Joe Anderson and Dr. Jill Silverman’s labs at UC Davis. Dr. Anderson reported at the 2020 FAST Scientific Conference this past December that they are in the final stages and have started their IND enabling studies. This could really be a reality for all of our kids in the near term!
And lastly, downstream therapeutics, such as NZ-2591, through Neuren Pharmaceuticals, hopeful to start in 2021, could potentially benefit all genotypes.
In the end, I am told that after further de-risking the ASOs through the current trials with participants who are fully null and have only one copy to activate (e.g. deletion positive), the trials will likely be opened up to our loved ones with UPD/ICD/mosaic AS. I do not know exactly when this will be, but I have a lot of faith it won’t be long.
My hope is high. Very high. While I wish Ben had gotten a working medicine yesterday; I remain hopeful that all individuals with AS are well positioned to get help soon.
Please do not hesitate to speak up. Ask questions of the FAST board and researchers with regard to any science being funded, genotype being (or not being) discussed, and I am always here to talk. If I don’t know the answer to your question, I will do my best to get it.
And I leave you by asking to you continue to fundraise for FAST. There is not a medicine in our loved one’s bodies yet. We are throwing as many balls down the lane at one time as we can think of and that we can afford. But as we learn more, we have so many new options, which could be better than the last, and we learn where we need to pivot, but to do that – we need money. Our fundraising dollars are hard at work, but we cannot take our foot off the gas until we have a working medicine in the bodies of ALL individuals living with Angelman syndrome.